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Long-chain Inulin (lc-inulin) prepared from chicory-derived inulin. Inulin is used in term infant formulas, toddler formulas, and medical foods. Ingredient is exempt from premarket approval requirements of the federal food, drug and cosmetic act.

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0% found this document useful (0 votes)

215 views126 pages

Ucm370435 PDF

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rupinder pal singh

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GRAS Notice (GRN) No.

477
http://www.fda.gov/Food/IngredientsPackagingLabeling/GRAS/NoticeInventory/default.htm

ORIGINAL SUBMISSION

000001

(b) (6)

000002

June 7, 2013
Via Express Courier
Ms. Paulette Gaynor
GRAS Notification Program
Office of Food Additive Safety (HFS-200)
Center for Food Safety and Applied Nutrition
Food and Drug Administration
5100 Paint Branch Parkway
College Park, MD 20740-3835
Re:

GRAS Notification for Long-chain Inulin (lc-inulin)

Dear Ms. Gaynor:

On behalf of Danone Trading B.V. (the Notifier), ENVIRON International Corporation is pleased to
submit this Notification of the Generally Recognized as Safe (GRAS) Determination for the
ingredient, long-chain inulin (lc-inulin) prepared from chicory-derived inulin for use in term infant
formulas, toddler formulas, and medical foods. This Notification contains 1) the Notifiers GRAS
Exemption Claim; 2) the Expert Panel Report on the GRAS Status of the ingredient; and 3) the
Safety Evaluation report.
Sincerely,
(b) (6)

Gavin Thompson
Principal
GT:gs
28-26968C

ENVIRON International Corp. 1702 East Highland Avenue, Suite 412, Phoenix, AZ 85016
V +1 602.734.7700 F +1 602.734.7701
environcorp.com

000003

GRAS Exemption Claim for

Long-Chain Inulin (lc-inulin)
Danone has determined that the use of long-chain inulin (lc-inulin) prepared from chicoryderived inulin in term infant formulas, toddler formulas, and medical foods is exempt from the
premarket approval requirements of the Federal Food, Drug and Cosmetic Act because Danone
has determined such use to be Generally Recognized As Safe (GRAS). This determination is in
compliance with proposed Sec. 170.36 of Part 21 of the Code of Federal Regulations (21 CFR
170.36) as published in the Federal Register, Vol. 62, No. 74, FR 18937, April 17, 1997.

A. Name and Address of Notifier

Mr. Flemming Morgan
President
Danone Trading B.V.
Schiphol Boulevard 105, Tower E, 1118 BG Schiphol Airport,
The Netherlands

B. Common or Usual Name of GRAS Substance

The substance that is the subject of this GRAS determination is long-chain inulin (lc-inulin).
Other chemical names for this lc-inulin are inulin or high performance inulin.
The lc-inulin product that is the subject of this GRAS determination is prepared from chicoryderived inulin by Orafti (Belgium) and sold under the trade name BeneoTM HP (formerly known
as Raftiline HP).

C. Intended Use
A combination of GOS and lc-inulin, in a 9:1 ratio of GOS:lc-inulin, is intended to be added to
term infant formulas, toddler formulas and medical foods. The intended uses of the GOS+lcinulin mixture and the typical and maximum concentrations of the mixture are detailed in
Table 1.

Table 1: Intended Uses of GOS+Long-Chain Inulin (9:1) in Term Infant Formulas,

Toddler Formulas and Medical Foods
Product Category
Intended Use
Total GOS+lc-Inulin Content(a)
Term infant formulas and
toddler formulas
Non-exempt and exempt
term infant formulas

Healthy term infants beginning at

birth and healthy term infants with
inborn errors of metabolism,
beginning at birth

8 g/L

Toddler formulas

Healthy children from 12 through 35

months of age

12 g/L

000004

GRAS Exemption Claim

for Long-Chain Inulin (lc-inulin)

-2-

Table 1: Intended Uses of GOS+Long-Chain Inulin (9:1) in Term Infant Formulas,

Toddler Formulas and Medical Foods
Product Category
Intended Use
Total GOS+lc-Inulin Content(a)
Maximum
Typical
Medical foods(c)
Nutritionally incomplete oral
medical foods

The products are intended to

supplement the normal diet and
cannot be used as the sole source of
nutrition. Nutritionally incomplete
products may be either general
formulas or disease-specific
formulas.

25 g /1000 kcal(b)

12.5 g/1000 kcal

Nutritionally complete foods

Oral foods

The products are intended to

supplement the normal diet but can
be used as the sole source of
nutrition when given in the
recommended amount. Nutritionally
complete products may be either
general formulas or disease-specific
formulas.

20 g /1000 kcal

10 g /1000 kcal

Tube feedings

20 g /1000 kcal

10 g /1000 kcal

Enteral nutritional products

Sole source enteral feedings or
12 g/L
for children (pediatric
supplemental nutrition for children
nutritionals)
Notes:
a
The ratio of GOS:lc-inulin is 9:1.
b
For certain disease-specific formulas with low energy densities (e.g., products for the dietary management
of metabolic disease), the maximum use of GOS+lc-inulin is 31.25 g per 1000 kcal.
c
The upper levels of GOS+lc-inulin intake may be well tolerated by some but not necessarily all people. A
specific dose of fiber may be achieved by using a fiber-containing medical food in combination of a fiber
free version of the same medical food.
Abbreviations: g = grams; GOS = galacto-oligosaccharide; kcal = kilocalories; L = liter; lc-inulin = long-chain
inulin

000005

(b) (6)

000006

Expert Panel Report on the

Generally Recognized as Safe Status of the
Proposed Uses of Long-Chain Inulin in
Term Infant Formulas, Toddler Formulas,
and Medical Foods
On behalf of Danone Trading B.V. (Danone), ENVIRON International Corporation (ENVIRON) convened
a panel of experts (Expert Panel), qualified by their scientific training and experience to evaluate the
safety of food ingredients, to determine the safety and the suitability and the Generally Recognized As
Safe (GRAS) status of the proposed uses of long-chain inulin (lc-inulin) prepared from chicory-derived
inulin. The Expert Panel Members included Joseph F. Borzelleca, Ph.D. (Professor Emeritus, Virginia
Commonwealth University School of Medicine; David J.A. Jenkins, M.D., Ph.D., D.Sc. (Professor,
University of Toronto; Director, Risk Factor Modification Centre, St. Michaels Hospital); and Judith
K. Jones, M.D., Ph.D. (The Degge Group, Ltd.).
The Expert Panel, independently and collectively, critically evaluated the available information presented
in documents prepared by ENVIRON (the GRAS dossier) and other materials deemed appropriate or
necessary. This information included a history of use and its regulatory status, the chemical identity of
lc-inulin, product specifications (Danone purchases the lc-inulin from BENEO-Orafti S.A. and certain
manufacturing details remain confidential business information and were not provided), intended
conditions of use and use levels, anticipated exposures, and safety assessment.
Following its independent and collective critical evaluation of the available information, the Expert Panel,
convened on 19 December 2012 with representatives of ENVIRON. Following extensive discussion, the
Expert Panel unanimously agreed to the conclusions described herein. A summary of the basis for these
conclusions follows.

Description of Long-Chain Inulin (lc-inulin), Manufacturing Process, Product

Specifications
The substance in this GRAS determination is long-chain inulin (lc-inulin) derived from inulin extracted
from chicory roots (Cichorium intybus). Inulin is a naturally occurring polysaccharide that belongs to a
class of carbohydrates known as fructans. Inulin-type fructans are linear polydisperse carbohydrates
mainly composed of fructose units joined by a series of 2-1 fructosyl-fructose linkages.
BENEO-Orafti S.A. (Beneo) supplies the lc-inulin product to Danone under the trade name Orafti HP
(formerly known as Raftiline HP or Beneo HP). Greater than 99.5% of Orafti HP long-chain inulin is
carbohydrate while water comprises the remaining < 0.5%. In 1623 batches produced since 2005, the
mean concentration of inulin with a DP 5 was 99.5% of total carbohydrate (SD = 0.5%) with fructose,
glucose, and sucrose combined constituting a mean of 0.1% (SD = 0.2%) of total carbohydrates. The
Chemical Abstracts Service Registry Number (CASRN) for inulin is 9005-80-5; lc-inulin does not have a
distinct CASRN.
The food-grade specifications for lc-inulin to be used by Danone are presented in Table 1. All tested
batches met the established specifications, demonstrating that the product meets appropriate
specifications for food-grade material. Regular compositional analyses by Beneo revealed the following
substances to be below the maximum residue levels set in EU regulation EC149/2008: cadmium and
mercury (< 0.01 milligram (mg)/kilogram (kg)); sulfur dioxide (< 5 mg/kg), nitrate (< 50 mg/kg), nitrite
(< 0.5 mg/kg); aflatoxin B1 (< 1microgram (g)/kg), ochratoxin A (< 0.5 g/kg), zearalenone (< 10 g/kg),
ENVIRON International Corp. 1702 East Highland Avenue, Suite 412, Phoenix, AZ 85016
V +1 602.734.7700 F +1 602.734.7701
environcorp.com

000007

GRAS Status Consensus Statement

Danone Trading B.V.
desoxynivalenol (< 0.1 g/kg); and polychlorinated biphenyls, dioxin-like compounds, and pesticides to be
below the levels of detection. Analyses of six batches demonstrated that phthalates are below limits of
quantitation (< 0.2 mg/kg).
Beneo lc-inulin has a guaranteed shelf of three years when stored in its original packaging.

Proposed Uses and Estimated Daily Intakes

Danone intends to use long-chain inulin in (1) exempt and non-exempt infant formulas for term infants
beginning at birth and toddler formulas intended for use by children from12 through 35 months of age;
and (2) medical foods, including nutritionally incomplete oral products, nutritionally complete oral
products, tube feedings, and pediatric nutritionals. The lc-inulin would be added to infant and toddler
formulas and medical foods in combination with galacto-oligosaccharides (GOS), with GOS accounting
for 90% of the total added prebiotics and lc-inulin accounting for the remaining 10%. The intended use
categories, target populations, and lc-inulin use levels are described below and summarized in Table 2.
The maximum target level of lc-inulin in all infant formulas for term infants (non-exempt and exempt) is
0.8 grams (g) lc-inulin per liter (L) formula as consumed, for a total of 8.0 g GOS/lc-inulin per L. Toddler
formulas are those formulas that are considered suitable for children from 12 through 35 months of age.
The maximum target use of lc-inulin in toddler formulas is 1.2 g lc-inulin per L formula as consumed, for a
total of 12 g GOS/lc-inulin per L.
Table 3 provides estimated intakes of lc-inulin based on the proposed use in infant formula. The proposed
use of up to 8 g GOS+lc-inulin per L in term infant formulas and 12 g GOS+lc-inulin per L in toddler
th
formulas, both in a ratio of 9:1, will result in intakes of up to 9.9 g GOS+lc-inulin per day at the 90
percentile of intake; this value represents the estimated intake from formula by infants 0-5 months of age.
th
Older infants and toddlers are estimated to have 90 percentile intakes of GOS+lc-inulin of 8.9 and
7.0 g/d, respectively. Medical foods are expected to provide up to 25 g GOS+lc-inulin per 1000
kilocalories (kcal) from nutritionally incomplete oral medical foods, 20 g/1000 kcal from nutritionally
complete oral foods and tube feedings, and 12 g/L from enteral nutritional products for children.

Safety Assessment
GRAS notifications 44, 118, 236, 286, 334, and 392 have previously addressed safety and tolerance of
2-1 fructans (inulin, oligofructose or FOS substances).
Absorption, Distribution, Metabolism and Excretion
Inulin, oligofructose and FOS are largely resistant to hydrolysis in the human upper intestine because the
majority of glycosidic bonds in inulin and oligofructose are D-fructofuranosyl 2-1 links. No enzyme able
to split this glycosidic linkage is secreted by salivary glands, the pancreas or in the small intestine of
humans.
Acute, Subchronic, Long-term, Carcinogenicity, Reproductive and Developmental and
Mutagenicity Studies
The prior GRAS notifications provided results for 2-1 fructans feeding studies in rats and mice,
teratogenicity study in rats, a chronic bioassay in rats and in vitro genetic toxicity studies. The animal
studies demonstrated no adverse effects from consumption of the fructans and results from in vitro
genetic toxicity studies demonstrated that commercially available FOS lacks any significant genotoxic
potential.

28-26968C

Page 2 of 7

000008

GRAS Status Consensus Statement

Danone Trading B.V.
Human Studies
Infant formulas containing up to 8 g/L of a combination of 90% GOS and 10% lc-inulin are approved for
use in the European Union and the intended use of the GOS+lc-inulin mixture in infant formulas is
therefore the same as the 8 g per L of this mixture approved for formulas in Europe by the Scientific
1
Committee on Food .
In a 2008 report, Food Standards Australia New Zealand (FSANZ) approved the addition of inulin-derived
2
substance and GOS at a maximum of 8 g/L in infant formulas . FSANZ based this decision on the history
of safe use of these substances in the general food supply in Australia and New Zealand and the
available evidence for infant formulas.
Results from 43 studies of healthy, male and female term infants consuming formula supplemented with
up to 0.8 g lc-inulin and 7.2 g GOS per L for periods ranging from two weeks to 12 months provide
evidence that lc-inulin, in combination with GOS, was well tolerated by infants and no adverse effects
were reported. The supplemented formulas supported normal growth. Additionally, results from these
studies suggest that formulas with added lc-inulin in combination with GOS may influence shifts in infant
gut microflora that result in gut microflora more similar to the gut microflora of breast-fed infants.
Results from 13 clinical trials provide additional evidence that the addition of a GOS/lc-inulin mixture in a
9:1 ratio to infant formula for preterm infants was well tolerated for two to 12 week study periods. These
preterm infants were closely monitored throughout the studies, and no adverse effects were reported on
measures of growth, fluid balance, formula intake, or stool characteristics. The absence of reports of
adverse effects in this highly sensitive and closely monitored study population provides strong evidence
for the safety and suitability of GOS and lc-inulin in a 9:1 ratio in infant formulas.
The results of these clinical studies of 2-1 fructan intake in healthy populations and four studies of
chronically ill populations requiring medical foods demonstrates that the proposed typical uses of lc-inulin,
in combination with GOS are well tolerated, safe, and suitable for the target populations.
Prebiotics such as GOS and lc-inulin are minimally digested in the small intestine (less than 10%) and
exert their putative health effects primarily via fermentation in the colon with the production of short-chain
fatty acids which are absorbed along the length of the colon. Gastrointestinal function is not usually
adversely affected in most infants requiring metabolic formulas; therefore, tolerance of exempt metabolic
infant formulas containing added lc-inulin and GOS ingredients would be expected to be equivalent to
tolerance of comparable doses in standard formulas.
The weight of the available evidence supports the safety of Danones proposed uses of lc-inulin as a food
ingredient in term infant formulas, toddler formulas and medical foods. This GRAS determination of the
proposed uses of this ingredient at the target levels described herein is based upon scientific procedures
as described under 21 C.F.R. 170.30(b), and is corroborated by a history of safe use of chicory.

Scientific Committee on Food (SCF). European Commission: Health and Consumer Protection Directorate-General.
2003. Report of the Scientific Committee on Food on the revision of essential requirements of infant formulae and
follow-on formulae: Adopted April 4, 2003. Available at: http://ec.europa.eu/food/fs/sc/scf/out199_en.pdf. Accessed
February 22, 2012.

Food Standards Australia New Zealand (FSANZ). 2008. Final assessment report: proposal P306, addition of
inulin/FOS & GOS to food. July 16. http://www.foodstandards.gov.au/_srcfiles/P306FOS & GOS FAR FINALv2.pdf

28-26968C

Page 3 of 7

000009

GRAS Status Consensus Statement

Danone Trading B.V.
Table 1. Specifications for LC-Inulin
Parameter
Specification Method
Inulin (% CHO)
>99.5
HPLC
Inulin DP 5 (% CHO)
99.0
HPLC
Average DP
23.0
HPLC
Glucose, fructose, sucrose (%
0.5
HPLC
CHO)
Sulfated ash (% at 525C)
<0.2
ICUMSA GS3/4/7/8-11
Conductivity (S/cm at 28 Brix)
<250
ICUMSA GS2/3-17
Lead (mg/kg)
<0.02
ICP-AES
Arsenic (mg/kg)
<0.03
ICP-AES
Total mesophilic bacteria (cfu/g)
1000
ICUMSA GS2/3-41
Yeasts (cfu/g)
20
ICUMSA GS2/3-47
Molds (cfu/g)
20
ICUMSA GS2/3-47
Enterobacteriaceae (cfu/1 g)
Negative
ISO 21528-2
Staphylococcus aureus
Negative
ISO 6888
(cfu/0.1 g)
Salmonella spp. (cfu/250 g)
Negative
AOAC 2000.07
Abbreviations:
AOAC = Association of Official Analytical Chemists
cfu = colony-forming units
CHO = carbohydrates
HPLC = high performance liquid chromatography
ICUMSA = International Commission for Uniform Methods of Sugar Analysis
ICP-AES = inductively coupled plasma-atomic emission spectrometry
ISO = International Organization for Standardization
(The remainder of this page intentionally left blank.)

28-26968C

Page 4 of 7

000010

GRAS Status Consensus Statement

Danone Trading B.V.
Table 2. Intended Uses of GOS/lc-Inulin in Term Infant
and Toddler Formulas, and Medical Foods
Product Category

Intended Use

Maximum or [Typical]
total GOS/lc-inulin
a
Content

Term infant formulas and toddler

formulas

Non-exempt and exempt term

infant formulas

-Formulas for healthy term infants

beginning at birth and healthy term
infants with inborn errors of
metabolism, beginning at birth.

8 g/L

Toddler formulas

-Formulas for healthy children from 12

through 35 months of age.

12 g/L

Nutritionally incomplete oral

medical foods

The products are intended to

supplement the normal diet and
cannot be used as the sole source of
nutrition. Nutritionally incomplete
products may be either general
formulas or disease-specific formulas.

25 g/1000 kcal
[12.5 g/1000 kcal]

Nutritionally complete foods

Oral foods

-The products are intended to

supplement the normal diet but can
be used as the sole source of nutrition
when given in the recommended
amount. Nutritionally complete
products may be either general
formulas or disease-specific formulas.

20 g/1000 kcal
[10 g/1000 kcal]

Tube feedings

20 g/1000 kcal
[10 g/1000 kcal]

Enteral nutritional products for

children (pediatric nutritionals)

-Children who need sole source

enteral feedings or supplemental
nutrition.

Medical foods

12 g/L

The ratio of GOS:lc-inulin is 9:1.

For certain disease-specific formulas with low energy densities (e.g., products for the dietary management of
metabolic disease), the maximum use of GOS/lc-inulin is 31.25 g per 1000 kcal.
c
The upper levels of GOS/lc-inulin intake may be well tolerated by some but not necessarily all people. A specific
dose of fiber may be achieved by using a fiber-containing medical food in combination of a fiber free version of
the same medical food.
Abbreviations: g = grams; GOS = galacto-oligosaccharide; kcal = kilocalories; L = liter; lc-inulin = long-chain inulin
b

28-26968C

Page 5 of 7

000011

GRAS Status Consensus Statement

Danone Trading B.V.

Table 3. Estimated 2-Day Average Intakes of Long-Chain Inulin from Infant Formula

Population

Infants, 0-5 mo
Infants, 6-11 mo
Toddlers, 12-35 mo

101
168
19

Percent
c
users

100
98.5
2.7

2-Day Average lc-inulin Intakes Per User

d
g lc-inulin/d
g lc-inulin/kg-bw/d
[g GOS/lc-inulin/d]
[g GOS/lc-inulin/kg-bw/d]
90th
90th
Mean
Percentile
Mean
Percentile
0.8

1.0

0.13

0.18

[7.5]

[9.9]

[1.3]

[1.8]

0.6

0.9

0.07

0.11

[6.4]

[8.9]

[0.7]

[1.1]

0.3

0.7

0.03

0.06

[3.3]
[7.0]
[0.3]
[0.6]
Breastfeeding infants and children were excluded from the sample population.
b
Number of people consuming infant formula during the study period.
c
Weighted percent.
d
Analysis of intake in terms of g lc-inulin/kg-bw/d was limited to infants with a measured body weight.
Abbreviations: bw = body weight; d = day; g = grams; GOS = galacto-oligosaccharide; kg = kilogram; lcinulin = long-chain inulin; mo = months.
a

(The remainder of this page intentionally left blank.)

28-26968C

Page 6 of 7

000012

GRAS Status Consensus Statement

Danone Trading B.V.

Conclusions
We, the members of the Expert Panel, have independently and collectively, critically evaluated the
available information on long-chain inulin prepared from chicory-derived inulin (presented in the dossier
prepared by ENVIRON and summarized above) and other information deemed appropriate and we
unanimously conclude that the proposed uses in infant formulas and toddler formulas and in medical
foods of long-chain inulin, manufactured consistent with current Good Manufacturing Practice (cGMP)
and meeting the food grade specifications presented in the dossier, are safe and suitable.
We further unanimously conclude that the proposed uses in infant formulas and toddler formulas and in
medical foods of long-chain inulin, manufactured consistent with current Good Manufacturing Practice
(cGMP) and meeting the food grade specifications presented in the dossier, are Generally Recognized As
Safe (GRAS) based on scientific procedures.
It is our opinion that other experts, qualified by scientific training and experience, and evaluating the same
data and information, would concur with these conclusions.

(b) (6)

Joseph F. Borzelleca, Ph.D.

Chair of the Expert Panel
Professor Emeritus
Pharmacology & Toxicology
School of Medicine,
Virginia Commonwealth University
Richmond, Virginia

Signature:

David J.A. Jenkins, M.D., Ph.D., D.Sc.

Professor
University of Toronto
Toronto, Ontario, Canada

Signature:

Date:

(b) (6)

Date:

(b) (6)
Judith K. Jones, M.D., Ph.D.
President and CEO
The Degge Group, Ltd.
Arlington, Virginia

28-26968C

Signature:

Date:

Page 7 of 7

~ ENVIRON
000013

Safety Evaluation of
Long-Chain Inulin
for Use in
Term Infant Formulas,
Toddler Formulas,
and Medical Foods

Prepared for:
Danone Trading B.V.
Schiphol, The Netherlands
Prepared by:
ENVIRON International Corporation
Phoenix, Arizona
Date:
April 30, 2013
Project Number:
28-26968B

000014

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Contents
Page
1

Executive Summary

2
2.1
2.1.1
2.1.2
2.1.3
2.1.4
2.1.5
2.2
2.2.1
2.2.2
2.3
2.3.1
2.3.2

Description of the Substance

Identity
Chemical Name and Identity
Common or Trade Names
CAS Registry Number
Physical and Chemical Properties
Chemical Structure of Long-Chain Inulin
Production Process
Source
Manufacturing Process
Product Specifications
Product Specifications and Batch Data
Product Stability

2
2
2
3
3
3
3
4
4
4
6
6
7

3
3.1
3.2
3.2.1
3.2.2
3.2.3
3.3
3.3.1
3.3.2
3.4
3.4.1
3.4.2

History of Use and Intended Uses

Dietary Exposure to Naturally Occurring Sources of Inulin
Dietary Exposure to Inulin Added to Food
Inulin Ingredients
Long-Chain Inulin Added to Infant Formulas
FOS Added to Enteral Formulas
Intended Uses of Long-Chain Inulin
Term Infant Formulas and Toddler Formulas
Medical Foods
Estimated Intakes of lc-Inulin from the Proposed Uses
Term Infant Formulas and Toddler Formulas
Medical Foods

8
8
8
8
10
10
10
11
12
15
15
16

Intended Effect

5
5.1
5.2
5.3
5.4
5.4.1
5.4.2
5.5
5.5.1
5.5.2
5.5.3
5.6
5.6.1
5.6.2
5.7
5.7.1
5.7.2
5.7.3
5.8

Safety Data
Introduction
Prebiotics and Inulin
Resistance of Inulin to Gastric Acidity, Hydrolysis and Absorption
Fermentation of lc-Inulin and Selective Stimulation of Intestinal Bacteria
In Vitro Evidence
In Vivo Evidence
Physiological and Systemic Effects of Inulin
Physiological Effects of Inulin in the Gastrointestinal Tract
Systemic Effects of Inulin
Potential Allergenicity of Inulin
Normal Development and Function of the Intestinal Microflora
Development of Microflora in the Infant
Functionality of Gut Microflora
Human Studies of Inulin and Oligofructose in Infant Formulas
Potential Role of Oligosaccharides in Infant Formula
Long-Chain Inulin and Galacto-Oligosaccharides in Term Infant Formulas and Weaning Foods
Water Balance
Long-Chain Inulin and Galacto-Oligosaccharides in Preterm Infant Formulas

19
19
20
20
22
22
24
26
26
27
28
33
33
34
35
35
36
67
68

Contents

000015

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

5.8.1
5.9
5.9.1
5.9.2

Human Studies of Inulin and Oligofructose in Preterm Infant Formulas

Use of Inulin and Oligofructose in Medical Foods
Potential Role of Fiber in Medical Foods
Tolerance of Inulin and Oligofructose in Medical Foods

68
76
76
76

6
6.1
6.2

Conclusions
Conclusion on the Use of Long-Chain Inulin in Infant Formula
Conclusion on the Use of Long-Chain Inulin in Medical Foods

86
86
86

References

List of Tables
Table 2-1.
Table 2-2.
Table 3-1.
Table 3-2.
Table 3-3.
Table 5-1.
Table 5-2.
Table 5-3.
Table 5-4.
Table 5-5.
Table 5-6.

Physical and Chemical Properties of Orafti HP LC-Inulin

Specifications and Batch Data for the Product: Orafti HP LC-Inulin
Enteral Formulas Containing FOS Available in the U.S.
Intended Uses of GOS/lc-Inulin in Term Infant and Toddler Formulas, and Medical Foods
Estimated 2-Day Average Intakes of Long-Chain Inulin from Infant Formula
In vitro Studies of Fermentation of Long-Chain Inulin
Human Studies of Long-Chain Inulin Effects on Microflora
Human Studies of Tolerance of Long-Chain Inulin
Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Studies of 90% GOS/10% Long-Chain Inulin in Preterm Infant Formula
Studies with FOS/Inulin/Oligofructose in Enteral Formulas

3
6
11
14
16
23
25
33
50
73
82

List of Figures
Figure 1. Chemical Structure of Long-Chain Inulin
Figure 2. Production Process of Orafti HP lc-inulin

4
5

List of Appendices
Appendix A.

Batch Data and Methods of Analysis for Orafti HP

Table A 1. Specifications and Annual Analyses of Product: Orafti HP lc-inulin

Appendix B.

GRAS Notifications for the Use of Inulin, Oligofructose and Fructooligosaccharides

as Food Ingredients

Appendix C.

References

Contents

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1 Executive Summary
Danone Trading B.V. (Danone) intends to add long-chain inulin (lc-inulin) to term infant formulas
(non-exempt and exempt), toddler formulas and medical foods in combination with galactooligosaccharides (GOS) at a ratio of 9:1 (GOS:lc-inulin).
In 2007, ENVIRON prepared a safety assessment and dossier of supporting information for
Nutricias (now Danone) GOS:lc-inulin product, presented it to a panel of food safety experts,
and obtained a generally recognized as safe (GRAS) Consensus Statement for the expert panel
subsequent to the panels deliberations on the GRAS status of Nutricias GOS:lc-inulin
combination.
Danone now wishes to update the lc-inulin safety assessment and dossier in preparation for the
submission of a GRAS notification to the United States Food & Drug Administration (FDA).
To determine that term infant formulas, toddler formulas and medical foods containing lc-inulin
are safe and GRAS at the proposed use levels, Danone engaged ENVIRON International
Corporation (ENVIRON) to prepare an updated safety dossier for lc-inulin for these proposed
uses. ENVIRON collected available characterization and safety data, researched and
assembled the additional publicly available data (in particular literature and information obtained
since the previous 2007 safety dosser was prepared), and conducted a safety review for each of
the principal ingredients. This document presents a safety assessment for the use of lc-inulin
as an ingredient in term infant formulas, toddler formulas and medical foods.
Herein, ENVIRON presents and interprets the currently available chemical characterization and
safety data for the lc-inulin.
ENVIRON concludes that the suitability and safety of the proposed uses is supported by the
appropriate, publicly available, scientifically defensible safety data.
(The remainder of this page intentionally left blank.)

Executive Summary

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2 Description of the Substance

2.1 Identity
2.1.1 Chemical Name and Identity
The substance that is subject of the evaluation for the GRAS determination is long-chain inulin
(lc-inulin) derived from inulin extracted from chicory roots (Cichorium intybus). Inulin is a
naturally occurring polysaccharide that belongs to a class of carbohydrates known as fructans.
Inulin-type fructans are linear polydisperse carbohydrates mainly composed of fructose units
joined by a series of 2-1 fructosyl-fructose linkages (Roberfroid 2005). The chemical formula
of inulin, from which lc-inulin is derived, is C6H11O6(C6H10O5)nOH and the systemic name for all
fructans is -D-glucopyranoside-(1-2)--D-fructofuranosyl-[(1-2)--D-fructofuranosyl]n (notably,
the -D-glucopyranoside-(1-2) is not always present). The n, which represents the number of
fructose units, ranges from 2 to 60 with some longer chains possible in native (or polydispersed)
inulin.
The individual members of the fructan oligo- and polysaccharides may be distinguished by their
source, method of production, or degree of polymerization (DP; the number of fructose or
glucose residues in the chain). All fructan products contain molecules with a range of DPs.
Inulin and related substances are referred to by various terms, including inulin, oligofructose,
fructo-oligosaccharide (FOS), and short-chain (sc) or long-chain (lc) fructo-oligosaccharide. The
nomenclature is not used consistently in the published literature and these terms are used
interchangeably. The nomenclature from GRN 392 is used in this dossier. The following is a
summary of the nomenclature.
Fructooligosaccharide (FOS): A fructan compound with nearly all DPs of less than 10 that is
obtained by the partial enzymatic hydrolysis of inulin or by enzymatic synthesis of sucrose
(transfructosylation). This definition is consistent with that promulgated by the International
Union of Biochemistry-International Union of Pure and Applied Chemistry Joint Commission on
Biochemical Nomenclature and AOAC International (IUB-IUPAC 1982; Niness 1999).
Short-chain Fructooligosaccharide (scFOS): A fructan compound with nearly all DPs 5,
typically with more than 85% of the molecules with a DP equal to three or four and only
negligible occurrence of DP > 5. Short-chain FOS residues are linked in the (2-1)
configuration and are synthesized from sucrose. Nearly all molecules have a glucose endcap.
Oligofructose: A fructan compound with DP < 10, with more than 25% of the molecules having
DP 5 and less than 75% with DP 4, produced by the enzymatic hydrolysis of inulin.
Inulin: A naturally-occurring fructan with DPs ranging from 2 to 60 or slightly higher. Because a
large portion of the molecules in inulin have DPs of 10 or less, oligofructose is a component of
inulin.
Long-chain inulin (lc-inulin): Inulin processed to remove the shorter chain fractions leaving
essentially fructans with DPs from 10 and higher (e.g., OraftiHP long-chain inulin). Because its
composition is primarily of longer-chain fructans, lc-inulin is a polysaccharide.

Description of the Substance

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Oligofructose-enriched inulin: Mixtures of oligofructose and lc-inulin.

The subject of this evaluation is lc-inulin. Lc-inulin is inulin which has been altered by applying
physical separation techniques to remove short chain molecules and leaving long chain fructans
with a DP of 10 and higher with an average DP of 23.

2.1.2 Common or Trade Names

BENEO-Orafti S.A. (Beneo) supplies the lc-inulin product under the trade name Orafti HP
(formerly known as Raftiline HP or Beneo HP). Beneo commonly refers to the product as
long-chain inulin or High Performance Inulin. Because nomenclature is not formally established
in the published literature, some authors refer to lc-inulin as long-chain fructooligosaccharide,
lcFOS or polyfructose.

2.1.3 CAS Registry Number

The Chemical Abstracts Service Registry Number (CASRN) for inulin is 9005-80-5; lc-inulin
does not have a distinct CASRN.

2.1.4 Physical and Chemical Properties

The general physical and chemical properties of Orafti HP lc-inulin are identified in Table 2-1.
OraftiHP consists essentially of carbohydrate and water (<3%). More than 99.5% of the
carbohydrates are lc-inulin and less than 0.5% mono- and disaccharides. In 1623 batches
produced since 2005, the mean concentration of inulin with a DP 5 was 99.5% of total
carbohydrate (SD = 0.5%) with fructose, glucose, and sucrose combined constituting a mean of
0.1% (SD = 0.2%) of total carbohydrates.
Table 4-1. Physical and Chemical Properties of Orafti HP LC-Inulin
Property
Appearance
Appearance in solution
Taste
Conductivity (28 Brix)
pH (30-50 Brix)
Solubility in water
Carbohydrate content
(of dry matter)
Source: Beneo.

Value
Colorless powder
Colorless
Neutral
<250 S/cm
5.0 7.0
<5 g/L
>99.5%

2.1.5 Chemical Structure of Long-Chain Inulin

The chemical structure of lc-inulin is identified in Figure 1. As shown in the figure, the number
of fructan units, n, may vary, but ranges from approximately 10 to 60 or more. Less than
1% of the chains are shorter than a DP of five. The molecular weight varies depending on the
DP, but ranges from approximately one to over 10 kilodaltons (kDa). Figure 1 depicts a terminal
glucose, however, this is occasionally not present. Figure 1 does not depict branched
molecules, which constitute less than two percent of the total molecules of Orafti HP lc-inulin.

Description of the Substance

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2.2

Production Process

Orafti HP lc-inulin is produced under International Food Standard and ISO 9001:2008
standards that includes a HAACP system that is regularly audited by certifying bodies.

Figure 1. Chemical Structure of Long-Chain Inulin

Source: Beneo.

2.2.1 Source
Orafti HP lc-inulin is prepared from chicory roots grown for Beneo by individually subcontracted farmers who receive free seeds from Beneo and grow only approved chicory
varieties using approved agronomic practices including the use of only permitted pesticides.
Chicory inulin has a DP range of two to more than 60 and the distribution is variable and
determined by factors such as climate, soil, harvest season, storage, and processing of the raw
inulin to remove sugars and salts.

2.2.2 Manufacturing Process

The manufacturing process of lc-inulin is depicted in Figure 2. The process involves three
phases. Phase I is extraction of raw inulin wherein the chicory roots are washed and sliced,
then extracted in hot water to yield raw juice. The raw juice contains inulin and other soluble
substances (pulp) as well as non-inulin substances (minerals, proteins, some sugars, and cellwall particles). The pulp is mechanically separated from the raw juice and used as animal feed
and the non-inulin substances are precipitated out using food-grade calcium hydroxide. The
precipicated substances are filtered out and the filtrate is evaporated to produce raw inulin.
In Phase II the raw inulin is crystallized by cooling to precipitate out the lower solubility longchain molecules. The solution is filtered and the dissolved molecules (salts, sugars and shorter
chain fructooligosaccharides) pass through the filter leaving the remaining precipitate. This
precipitate is raw HP.
In Phase III the raw HP is redissolved in water and then subjected to ultra-heat treatment to
reduce the microbial load followed by fine filtration. Adsorption with activated carbon may
optionally be used in addition to provide decolorization. The product is dried by evaporation to
reduce the water content to less than 0.5% as specified.

Description of the Substance

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All the production agents such as precipitation and pressing aids, defoaming agents, antiscaling compounds, flocculants, pH-regulators, biocides, filtration aids, and adsorbents are food
grade or have no food contact. All production aids and additives are approved for use under
regulations listed in parts 173, 182, or 184 of the Code of Federal Regulations except activated
carbon and diatomaceous earth; both are GRAS substances with long histories of safe use in
food processing.

Figure 2. Production Process of Orafti HP lc-inulin

Source: Beneo.

Description of the Substance

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2.3 Product Specifications

2.3.1 Product Specifications and Batch Data
Beneo has established for Orafti HP lc-inulin the product specifications displayed in Table 2-2
to ensure a consistent food-grade product.
Ten non-sequential batches of product were analyzed with regard to the chemical and
microbiological parameters listed in the specifications. As shown in Table A-1 (Appendix A), all
tested batches met the established specifications, demonstrating that the product meets
appropriate specifications for food-grade material and that a consistent product can be and is
produced.
In addition to the parameters included in the product specifications, Beneo regularly analyzes
raw inulin as well as OraftiHP long-chain inulin for possible contaminants, including the heavy
metals cadmium and mercury; sulfur dioxide, nitrate, and nitrite; polychlorinated biphenyls
(PCB) and dioxin-like compounds (DLC); the mycotoxins aflatoxin B1, ochratoxin A,
zearalenone, and desoxynivalenol; and ~450 pesticides for which maximum residue levels are
set in EU regulation EC149/2008. These analyses have found cadmium and mercury levels to
be < 0.01 mg/kg; sulfur dioxide < 5 mg/kg, nitrate < 50 mg/kg, nitrite < 0.5 mg/kg; aflatoxin B1
< 1g/kg, ochratoxin A < 0.5 g/kg, zearalenone < 10 g/kg, desoxynivalenol < 0.1 g/kg; and
PCBs, DLCs, and pesticides below the levels of detection.
Table 4-2. Specifications for Orafti HP LC-Inulin
Parameter
Inulin (% dry matter)
Inulin DP 5 (% dry matter)
Average DP
Glucose, fructose, sucrose
(% dry matter)

Specification
>99.5
99.0
23.0

HPLC
HPLC
HPLC

Method

0.5

Prepared for Danone Trading B.V.

Sensus chicory-derived inulin is two to greater than 60. Based on these proposed uses,
Imperial Sensus estimated that dietary intake of inulin at the 90th percentile level would be
approximately 6 grams per day for infants less than one year of age, approximately 15 grams
per day for infants one year of age, and approximately 20 grams per day for the general
population (i.e., two years of age and older). Based on the information provided by Imperial
Sensus, as well as other information available to FDA, the agency had no questions regarding
Imperial Sensus' conclusion that inulin is GRAS under the intended conditions of use (FDA
GRN 118 response letter 2003, Appendix B).
GTC Nutrition Company (GTC) notified FDA in May, 2000 that its FOS was determined to be
GRAS for use in selected foods as a bulking agent. The FOS produced by GTC Nutrition
Company is manufactured from sucrose syrup; the action of the fungal enzyme fructofuranosidase on the syrup yields fructose chains (primarily 2-4 units) with a terminal
glucose. The proposed uses of FOS in foods included use in baby foods at levels ranging from
0.1 to 3.6%. Based upon the proposed uses, GTC estimated that dietary exposure to FOS from
its intended use as a bulking agent would range from approximately 3.1 to 12.8 grams per
person per day at the 90th percentile consumption level. Based on the information provided by
GTC Nutrition, as well as other information available to FDA, the agency had no questions at
that time regarding GTC Nutrition's conclusion that FOS is GRAS under the intended conditions
of use (FDA GRN 44 response letter 2000, Appendix B).
In January, 2007, GTC notified FDA that GTCs FOS product (DP 2-4) was determined to be
GRAS for foods in general, excluding meat and poultry products and infant formula, at levels up
to 20 grams per day in the general population and at levels of up to 4.2 grams per day for
infants less than one year of age. Examples of typical FOS use levels were provided for a
variety of foods, including infant foods, with typical FOS use levels ranging from 0.4 to 3.6%,
and toddler foods, with typical FOS use levels ranging from 0.8 to 6.7%. FDA informed GTC
Nutrition that the agency had no questions regarding the companys conclusion that FOS is
GRAS under the intended conditions of use (FDA GRN 44 response letter 2007, Appendix B).
In October of 2010, FDA notified Yakult Pharmaceutical Industry Co., Ltd. that FDA had no
questions regarding Yakults use of GOS in term infant formula at 7.2 grams per liter (FDA GRN
334 response letter 2010, Appendix B). Additionally, Pfizer Nutrition and BENEO-Orafti recently
notified FDA that an expert panel concluded that their oligofructose (DP 2-8) was determined to
be GRAS for up to 3 grams per liter in milk-based term infant formula powder and FDA had no
questions (FDA GRN 392 response letter 2012, Appendix B).
In countries other than the U.S., inulin is often legally classified as a food or food ingredient, and
not as an additive (Coussement 1999). Inulin is not listed as an accepted food additive in the
standard positive lists from the European Union (EU) or from Codex Alimentarius. EU
Regulation 1333/2008 explicitly lists inulin as a substance that is not an additive. Describing
inulin as a dietary fiber is consistent with the Codex Alimentarius definition of dietary fiber.
The Codex Alimentarius dietary fibers are:
carbohydrate polymers with ten or more monomeric units, which are not hydrolyzed by the
endogenous enzymes in the small intestine of humans and belong to the following categories:

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Edible carbohydrate polymers naturally occurring in the food as consumed,

carbohydrate polymers, which have been obtained from food raw material by physical,
enzymatic or chemical means and which have been shown to have a physiological effect
of benefit to health as demonstrated by generally accepted scientific evidence to
competent authorities,

synthetic carbohydrate polymers which have been shown to have a physiological effect
of benefit to health as demonstrated by generally accepted scientific evidence to
competent authorities (Codex 2010).

3.2.2 Long-Chain Inulin Added to Infant Formulas

The European Commission, Scientific Committee on Food (SCF) has stated that non-digestible
carbohydrates may be added to infant formulas for technical reasons or if they provide a source
of fermentable substrates for the gut microflora (SCF 2003). The SCF stated that it had no
major concerns with the inclusion of a combination of 90% galacto-oligosaccharides (GOS) and
10% FOS in infant formulas and follow-on formulas, provided that their total content does not
exceed 8 g per L; this maximum concentration is equivalent to 7.2 g GOS and 0.8 g FOS per L
infant formula. The GOS component of GOS-FOS mixtures added to infant formulas in Europe
consists of molecules with relatively short chain lengths (i.e., less than 10 galactose units), while
the FOS component consists of molecules with long chain lengths and corresponds to lc-inulin.
In a 2008 report, Food Standards Australia New Zealand (FSANZ) approved the addition of
inulin-derived substance and GOS at a maximum of 8 g/L in infant formulas (FSANZ 2008).
FSANZ based this decision on the history of safe use of these substances in the general food
supply in Australia and New Zealand and the available evidence for infant formulas. To reach
their decision FSANZ reviewed a number of studies regarding the use of GOS and lc-inulin in
combination (9:1 ratios). These studies are included in this safety evaluation in section 5.7.2.

3.2.3 FOS Added to Enteral Formulas

A variety of enteral formulas containing FOS (DP 2-4) are marketed in the U.S. These products
are summarized in Table 3-1. As shown in the Table, the FOS content in these tube/sole
source nutrition products ranges from 2.5 to 6.7 g per 1000 kcal, while total fiber content ranges
from 2.5 to 21.1 g per 1000 kcal. These oral supplements provide 1 to 3 g FOS per serving,
and 1 to 4 g of total fiber.

3.3

Intended Uses of Long-Chain Inulin

History of Use and Intended Uses

5.0

Perative

1.3

5.0

Pivot 1.5 Cal

1.5

5.0

Suplena

1.8

5.3

8.7

TwoCal

2.5

Vital jr

3.0

3.0
Total Dietary
Fiber per Unit
(g)

Unit Size

Calories

FOS (g) per

Unit

Ensure Fiber

8 fl oz.

250

1.0

2.8

Ensure Pudding

4 fl oz.

170

1.0

Glucerna Shake

8 fl oz.

220

1.0

2.8

Glucerna Weight Loss Shake

11 fl oz.

290

1.4

4.0

ProSure Shake

8 fl oz.

300

3.0

Product
Oral
Supplement
s

FOS used in all enteral formulas assumed to have DP of 2-4.

Abbrevations: fl oz = fluid ounce; FOS = fructooligosaccharide; g = grams; kcal = kilocalories; mL = milliliter.
Source: Ross Nutrition 2006.

Danone intends to use long-chain inulin in infant formulas for term infants beginning at birth,
including infant formulas specifically designed for infants with metabolic disorders such as
phenylketonuria, tyrosinaemia, maple syrup urine disease, or fatty acid oxidation disorders.
Although the incidence of any particular metabolic disorder is quite rare, approximately 1 in
1,500 individuals has some form of an inborn error of metabolism (Raghuveer et al. 2006).
Formulas specifically formulated for use by infants who have inborn errors of metabolism or low
birth weight, or who otherwise have unusual medical or dietary problems are referred to as
exempt infant formulas (21 CFR 107.3). Metabolic infant formulas and other products for infants
with special nutritional needs are formulated with a selected balance of amino acids that does

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not include one or more amino acids specific to the metabolic disorder. Metabolic infant
formulas are not intended to provide a sole source of nutrition for infants. Rather, infants with
metabolic disorders are fed a combination of standard infant formulas (or human milk) and
exempt infant formulas to meet their nutritional needs. The maximum target use of lc-inulin in
all infant formulas for term infants (non-exempt and exempt) is 0.8 g lc-inulin per L formula as
consumed, for a total of 8.0 g GOS/lc-inulin per L.
Toddler formulas are those formulas that are considered suitable for children from 12 through
35 months of age. The maximum target use of lc-inulin in toddler formulas is 1.2 g lc-inulin per
L formula as consumed, for a total of 12 g GOS/lc-inulin per L.

3.3.2 Medical Foods

Medical foods are used under physician supervision as supplemental or sole sources of nutrition
for individuals who are unable to ingest sufficient nutrition from conventional foods. Danone
intends to add GOS to both nutritionally incomplete and nutritionally complete medical foods,
including pediatric nutritionals.

3.3.2.1 Nutritionally Incomplete Oral Medical Foods

Nutritionally incomplete oral medical foods are products that are intended to supplement the
normal diet. They are not designed to provide all nutrient needs and cannot be used as the sole
source of nutrition. Nutritionally incomplete products may be either general formulas or diseasespecific formulas. General formulas provide high amounts of selected nutrients (often protein)
in a palatable format. The products are used to address malnutrition experienced by patients
suffering from anorexia, cachexia, sarcopenia, burns, hip fracture and other medical conditions,
as well as pre- and post-surgery.
Disease- specific formulas are adapted with respect to macro- and micronutrients to meet the
needs of a specific disease or condition. These products include those for the dietary
management of metabolic disease and also those used to address malnutrition experienced by
renal patients, and patients suffering from pressure ulcers, cystic fibrosis or chronic obstructive
pulmonary disease.
The typical use level of lc-inulin in nutritionally incomplete oral medical foods is approximately
1.25 g lc-inulin per 1000 kcal product. The maximum proposed use level of lc-inulin in these
medical foods is approximately 2.5 g lc-inulin per 1000 kcal product. Typical and maximum use
levels of the GOS/lc-inulin mixture are 12.5 and 25 g GOS/lc-inulin per 1000 kcal, respectively.
For certain disease-specific formulas with low energy densities, for example those used in the
dietary management of metabolic disease, the proposed maximum use level of GOS/lc-inulin
may be up to 25% higher than the use in other medical foods. In these products, the maximum
use of GOS/lc-inulin is 31.25 g per 1000 kcal product, which is equivalent to approximately 28 g
GOS and 3 g lc-inulin per 1000 kcal.

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3.3.2.2 Nutritionally Complete Oral Medical Foods

Nutritionally complete oral medical foods products are intended to supplement the normal diet
but can be used as the sole source of nutrition when given in the recommended amount.
Nutritionally complete products may be either general formulas or disease-specific formulas.
Standard formulas comply with the reference values for macro- and micronutrients for a healthy
population. The products are used to address malnutrition experienced by various patients
such as geriatric patients, patients recovering from illness or patients suffering from neurological
disorders (e.g. Parkinsons, Alzheimers), dysphagia, HIV/AIDS or other diseases affecting food
intake or requiring extra intake of energy and nutrients. Disease-specific formulas are adapted
with respect to macro- and micronutrients to meet the needs of a specific disease or condition.
The products are used to address malnutrition experienced by various patients such as cancer
patients and diabetic patients.
Tube feeds are intended for enteral nutritional support for patients with a functional or partially
functional gastrointestinal tract who are unable or unwilling to eat sufficient quantities of
conventional food to meet their nutritional requirements. The products are nutritionally complete
and intended as the sole source of nutrition for the patients. Tube feeds can either be standard
formulas or disease-specific formulas. Standard formulas comply with the reference values for
macro- and micronutrients for a healthy population. The products are used to supply nutrition to
a variety of patients including pediatric patients, patients with central nervous system disorders,
cerebrovascular disease, multiple sclerosis and respiratory diseases. Disease-specific formulas
are adapted with respect to macro- and micronutrients to meet the needs of a specific disease
or condition. The products are used to supply nutrition to various patients such as cancer
patients and diabetic patients.
Nutritionally complete products are intended to be used when conventional foods cannot be
taken and therefore should supply the same nutrients as conventional foods. The
recommended intake of dietary fiber in the U.S. is 14 g per 1000 kcal, which is equivalent to
21 to 38 g fiber per day for adults (IOM 2002). A diet providing 20-30 g of fiber and 15002500 kcal would provide up to 20 g fiber per 1000 kcal. The typical use level of lc-inulin in
nutritionally complete medical foods is approximately 1.0 g lc-inulin per 1000 kcal product, and
the maximum target use level is approximately 2.0 g lc-inulin per 1000 kcal product. The typical
and maximum use levels of the GOS/lc-inulin mixture in nutritional complete medical foods
therefore are 10 and 20 g GOS/lc-inulin per 1000 kcal, respectively.

3.3.2.3 Pediatric Nutritionals

Danone intends to add long-chain inulin to nutritionals for pediatric populations. These products
are designed for children who need sole source enteral feedings or supplemental nutrition.
Children approximately 1 to 13 years of age are the target population for these nutritional
products. The maximum target use level of lc-inulin in pediatric nutritionals is 1.2 g lc-inulin
per L formula.

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Table 4-2. Intended Uses of GOS/LC-Inulin in

Term Infant and Toddler Formulas, and Medical Foods
Product Category

Intended Use

Maximum or [Typical]
total GOS/lc-inulin
a
Content

Term infant formulas and

toddler formulas
Non-exempt and exempt term
infant formulas

Toddler formulas
Medical foods

-Formulas for healthy term infants

beginning at birth and healthy term
infants with inborn errors of
metabolism, beginning at birth.
-Formulas for healthy children from 12
through 35 months of age.

8 g/L

12 g/L

Nutritionally incomplete oral

medical foods

The products are intended to

supplement the normal diet and
cannot be used as the sole source of
nutrition. Nutritionally incomplete
products may be either general
formulas or disease-specific formulas.

25 g/1000 kcal
[12.5 g/1000 kcal]

Nutritionally complete foods

Oral foods

-The products are intended to

supplement the normal diet but can
be used as the sole source of nutrition
when given in the recommended
amount. Nutritionally complete
products may be either general
formulas or disease-specific formulas.

20 g/1000 kcal
[10 g/1000 kcal]

Tube feedings

20 g/1000 kcal
[10 g/1000 kcal]

Enteral nutritional products for

children (pediatric nutritionals)

-Children who need sole source

enteral feedings or supplemental
nutrition.

12 g/L

The ratio of GOS:lc-inulin is 9:1.

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3.4 Estimated Intakes of lc-Inulin from the Proposed Uses

3.4.1 Term Infant Formulas and Toddler Formulas
Danone proposes to use up to 0.8 g lc-inulin/L in exempt and non-exempt infant formulas for
term infants, and up to 1.2 g lc-inulin/L in toddler formulas. Estimates of potential intakes of lcinulin resulting from these intended uses were calculated using food consumption data reported
in the United States Department of Health and Human Services 2003-2004 National Health and
Nutrition Examination Survey (NHANES). This NHANES data set provides nationally
representative nutrition and health data and prevalence estimates for nutrition and health status
measures in the United States (NCHS 2006).
As part of the examination, trained dietary interviewers collected detailed information on all
foods and beverages consumed by respondents in the previous 24 hour time period (midnight to
midnight). A second dietary recall was administered by telephone 3 to 10 days after the first
dietary interview, but not on the same day of the week as the first interview. A total of
9,043 respondents provided complete dietary intakes for the Day 1 recall, and 8,354 of the
individuals provided a complete Day 2 recall. Using the list of food codes and the NHANES
2003-2004 dietary recall data files from individuals with two complete days of dietary recall,
ENVIRON estimated mean and 90th percentile 2-day average intakes of lc-inulin from infant
and toddler formulas.
The food codes for infant formulas reported by NHANES respondents are identified by brand
name and form (e.g., prepared from powder or ready-to-feed). The majority of infant formulas
specified in the dietary recalls are non-exempt formulas for term infants. Intake of exempt
formulas such as protein hydrolysate formulas and formulas for preterm infants was reported for
some survey participants, though no intake of metabolic formulas was reported. All formulas
were included in the estimates of intake.
Infants with metabolic disorders are fed a combination of standard infant formulas (or human
milk) and exempt infant formulas to meet their nutritional needs. We therefore assumed that the
total amount of infant formula consumed by NHANES participants is representative of the total
amount of formula consumed by infants with metabolic disorders, and that the concentration of
lc-inulin is the same in all formulas. We assumed all formulas for infants contained 0.8 g
lc-inulin per L formula (as consumed), and we assumed toddler formulas contained 1.2 g
lc-inulin per L formula. The 2-day average intakes represent the total estimated intakes of
lc-inulin during the two days of recall divided by two (i.e., (IntakeDay 1 + IntakeDay 2)/2).
Intakes were calculated for three subpopulations of infants (0-5 months, 6-11 months, and
12-35 months). The estimates were generated using survey sample weights to adjust for
differences in representation of subpopulations; results therefore are representative of the
U.S. population.
Estimates of lc-inulin intake from infant formula based on the proposed uses of the ingredient
are shown in Table 3-3. The mean estimated intake of lc-inulin by infants 0-5 months old is
0.8 g lc-inulin/day and the mean lc-inulin intake by infants 6-11 months old is 0.6 g lc-inulin/day.
The estimated 90th percentile 2-day average intake of lc-inulin by infants 0-5 months old is 1.0 g
lc-inulin/kg-bw/day, and the estimated 90th percentile intake by infants 6-11 months of age is
0.9 g lc-inulin/kg-bw/day. A small number of infants consume formulas after the first year of life,

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and formula intakes are lower than intakes by infants 0-5 or 6-11 months of age. The estimated
mean and 90th percentile 2-day average intakes of lc-inulin by toddlers, the target consumers of
toddler formulas, are 0.3 and 0.7 g/day, respectively.
Table 4-3. Estimated 2-Day Average Intakes of Long-Chain Inulin from Infant Formula

Population

Infants, 0-5 mo
Infants, 6-11 mo
Toddlers, 12-35 mo

101
168
19

2-Day Average lc-inulin Intakes Per User

d
g lc-inulin/d
g lc-inulin/kg-bw/d
[g GOS/lc-inulin/d]
[g GOS/lc-inulin/kg-bw/d]
90th
90th
Mean
Percentile
Mean
Percentile

Percent
c
users

100.0
98.5
2.7

0.8

1.0

0.13

0.18

[7.5]

[9.9]

[1.3]

[1.8]

0.6

0.9

0.07

0.11

[6.4]

[8.9]

[0.7]

[1.1]

0.3

0.7

0.03

0.06

[3.3]

[7.0]

[0.3]

[0.6]

Breastfeeding infants and children were excluded from the sample population.
Number of people consuming infant formula during the study period.
c
Weighted percent.
d
Analysis of intake in terms of g lc-inulin/kg-bw/d was limited to infants with a measured body weight.
b

Abbreviations: bw = body weight; d = day; g = grams; GOS = galacto-oligosaccharide; kg = kilogram; lc-inulin = longchain inulin; mo = months.

The estimates of potential lc-inulin intake by infants were based on the survey population of
non-breastfeeding infants. The youngest infants (i.e., prior to introduction of weaning foods) in
the sample population therefore are presumably consuming only infant formula. Some infants,
however, consume a combination of human milk and infant formula. The intakes of lc-inulin by
infants consuming a combination of human milk and formula would likely be lower than the
estimates of lc-inulin intake based on the population of exclusively formula fed infants.

3.4.2 Medical Foods

5 Safety Data
5.1

Introduction

Inulin, oligofructose, and FOS are characterized by chains of fructose units connected by the
characteristic 2-1 linkage usually connected to a single glucose molecule. The safety of inulin
and related 2-1 fructans has been reviewed and several ingredients have been determined to
be Generally Recognized As Safe for use in foods (Appendix B).
Imperial Sensus notified the U.S. Food and Drug Administration of the GRAS status of chicoryderived inulin in 2002 (GRAS Notice No. GRN 000118). Based on the proposed uses, Imperial
Sensus estimated that dietary intake of inulin at the 90th percentile level would be
approximately 6 grams per day for infants less than one year of age, approximately 15 grams
per day for infants one year of age, and approximately 20 grams per day for the general
population (i.e., two years of age and older). The GRAS uses of chicory-derived inulin were
later revised to include addition of up to 1 g inulin per serving of baby food, excluding infant
formula. The DP in this chicory-derived inulin is from two to greater than 60. Beneo previously
determined that the use of inulin and oligofructose as an ingredient in selected foods is GRAS,
though FDA was not notified of this determination.
FOS (DP 2-4) manufactured from sucrose syrup (GTC Nutrition) was determined to be GRAS
for foods in general, excluding meat and poultry products and infant formula, at levels up to
20 grams per day in the general population and at levels of up to 4.2 grams per day for infants
less than one year of age. FDA was notified in 2000 of the GRAS status of FOS when used in
selected foods (GRAS Notice No. GRN 000044), and in 2007 GTC notified FDA of the GRAS
status of the expanded list of uses. Yakult Pharmaceutical Industry Co., Ltd. notified FDA that
the use of GOS in term infant formula at 7.2 grams per liter was GRAS (GRN 000334) and in
October of 2010, FDA responded that it had no questions regarding Yakults GRAS
determination. Pfizer Nutrition and Beneo recently notified FDA that an expert panel concluded
that their oligofructose (DP 2-8) was GRAS and FDA had no questions (GRN000392; May
2012).
Information on the gastrointestinal fate, animal toxicity, and human tolerance of inulin and
related 2-1 fructans was considered in making the safety evaluations of both chicory-derived
inulin and FOS. As detailed in the GRAS determinations and summarized by FDA
(Appendix B), the 2-1 linkage is largely resistant to digestion in the small intestine and arrives
intact in the colon where it is subject to fermentation. Fermentation by the microflora produces
gases and short chain fatty acids. The short-chain fatty acids are used locally as an energy
source by the resident flora, taken up systemically via the colonocytes and transported to the
liver for caloric utilization by the host, or excreted in the feces.
The GRAS determinations state that results from published animal studies including acute
studies in rats and mice, 6-week feeding studies in rats, a teratogenicity study in rats, and a
chronic bioassay in rats, support the safety determination. Results from in vitro genetic toxicity
studies demonstrated that commercially available FOS lacks any significant genotoxic potential.

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Based on a review of studies of human tolerance of inulin and related 2-1 fructans, it was
concluded that adults could regularly consume 40 to 70 grams of inulin per day (preferably in
multiple doses throughout the day) without experiencing any significant adverse effects. The
estimated 90th percentile intake of chicory-derived inulin is approximately 20 g/day for the
general population, which is no more than half the amount shown to be well-tolerated
(Appendix B; FDA 2003).
In the evaluation of the safety of ingestion of long-chain inulin (lc-inulin) under its intended
conditions of use along with GOS, ENVIRON reviewed available information on the digestion,
fermentation and tolerance of this particular subset of 2-1 fructans. The safety review begins
with a description of prebiotics and the general criteria by which inulin has been identified as a
prebiotic. The introduction is followed by a discussion of the digestion of 2-1 fructans and a
review of in vitro and in vivo data regarding the fermentation of lc-inulin. Information on other
gastrointestinal and systemic effects of inulin consumption is then summarized, and data on the
tolerance of lc-inulin are presented. We then review the normal development and function of
the colonic microflora. Studies of infants consuming the proposed combination of 90% GOS
and 10% lc-inulin in infant formulas and weaning foods are reviewed and summarized, as are
studies of individuals consuming medical foods containing a mixture of 90% GOS and 10%
lc-inulin or medical foods containing 2-1 fructans. ENVIRON conducted searches for
information on these topics using Medline and the Toxicology and Medicine categories within
the DIALOG database. The terms used in the searches included inulin, fructooligosaccharide, oligofructose, and HP inulin. Information published from the year 2000 (the
year of the first GRAS Notification for inulin) to 2007 (the time of this safety review) was
included in the search. ENVIRON conducted additional searches using PubMed and Google
Scholar with the same terms to include information published up to mid-2012. Potentially
relevant articles were identified and reviewed; the findings are described below. To maintain
consistent terminology, ENVIRON used lc-inulin to describe lcFOS in the studies where test
substance information was adequately descriptive. If the test substance was inadequately
described to make a determination, then ENVIRON used the same terms as the study.

5.2

Prebiotics and Inulin

Inulin is considered a prebiotic based on its (1) resistance to gastric acidity, hydrolysis by
mammalian enzymes, and gastrointestinal absorption; (2) fermentation by intestinal microflora;
and (3) selective stimulation of the growth and/or activity of those intestinal bacteria that
contribute to health and well-being (Roberfroid 2007). Based on these criteria, inulin has been
determined to be a prebiotic. Galacto-oligosaccharides (GOS) also have been determined to be
a prebiotic.

5.3

Resistance of Inulin to Gastric Acidity, Hydrolysis and Absorption

The majority of glycosidic bonds in inulin and oligofructose are D-fructofuranosyl 2-1 links.
No enzyme able to split this glycosidic linkage is secreted by salivary glands, the pancreas or in
the gastric and small intestinal epithelium brush border in mammals. Inulin, oligofructose and
FOS therefore are largely resistant to hydrolysis in the human upper intestine (Cherbut 2002).
No studies in which the digestibility of lc-inulin was tested were identified; results from studies of
inulin, oligofructose and FOS, however, provide evidence about the digestibility of the polymer.

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Results from studies in ileostomy patients provide evidence of the resistance of inulin and
oligofructose to hydrolysis. In a study of 10 patients with conventional ileostomies due to
ulcerative colitis, 88% (95% Confidence Interval, 76-100%) of a 17 g dose of pure inulin (derived
from chicory root with an average DP of 10) was recovered in the ileostomy effluent. Eight of
the original 10 patients were tested again and 89% (95% Confidence Interval, 64-114%) of a
17 g dose of oligofructose (average DP 4) was recovered in the ileostomy effluent (Ellegard et
al. 1997). In another study of ileostomy patients, patients were fed inulin (DP >10 for over 30%
of the chains) extracted from Jersusalem artichoke and recovery was measured in the ileal
effluent (Knudsen and Hessov 1995). Recovery of inulin in the effluent was not different
following ingestion of two separate doses; recovery was 86.4 4.4% and 87 3.3% from
10 and 30 g inulin doses, respectively. The fructose:glucose ratio of the ingested inulin was 4.1.
The fructose:glucose ratio of the recovered inulin in ileal effluent was 4.7 1.1% at an intake of
10 g inulin and 4.5 0.1% at an intake of 30 g inulin. The higher ratio of fructose:glucose
recovered in the effluent as compared to the ratio in the ingested inulin suggests that lowmolecular-weight components of inulin are more sensitive to hydrolysis than the higher
molecular weight components. The investigators speculated that the small loss of inulin during
passage through the small intestine was due to hydrolysis by either acids or enzymes or to
microbial degradation by the microflora colonizing the distal small intestine.
The passage of FOS (DP 2-4) into the colon in healthy adults was investigated using the direct
method of measuring the flow rate in the distal ileum (Molis et al. 1996). After an equilibration
phase, the six healthy subjects consumed 20.1 g FOS per day, given in three postprandial
doses. Results showed that an average of 89 8.3% of the ingested FOS was delivered to the
colon, and analysis of the constituent oligosaccharides in the ileal contents showed that most
unabsorbed FOS was in an intact unhydrolyzed form. Approximately 0.12% (24 12 mg) of the
ingested FOS dose was recovered in 24-hour urine samples, and approximately 1% of the FOS
that disappeared from the small intestine was recovered in urine. It was assumed that excretion
of FOS in the urine resulted from absorption of intact FOS in the small intestine, and that a
fraction of the ingested FOS was hydrolyzed during passage through the gastrointestinal tract
and then absorbed as glucose and fructose. Stool samples contained no FOS, indicating that
FOS that was not hydrolyzed in the upper gastrointestinal tract was completely fermented in the
colon.
Results from breath hydrogen studies in humans following ingestion of FOS or inulin provide
further evidence that the carbohydrates are not absorbed. Stone-Dorshow and Levitt (1987)
measured breath hydrogen excretion in 10 subjects consuming 5 g FOS (Neosugar, Meiji Seika
Co., Japan; DP 2-4) three times daily for a period of 12 days. Following the 12-day period of
FOS ingestion, breath hydrogen excretion after a 10 g dose of FOS was approximately 50%
higher as compared to the baseline measurement; the increase was not statistically significant.
Breath hydrogen excretion after the 10 g FOS dose was similar to excretion after ingestion of
10 g lactulose, which suggests near total malabsorption of FOS. Geboes et al (2003)
conducted breath hydrogen tests using oligofructose (Raftilose), inulin (Raftiline ST), or lc-inulin
(Raftiline HP). Following ingestion of 5 g lc-inulin, an increase in hydrogen excretion of 10 ppm
above baseline was seen in five of six study volunteers in one trial, and in 15 of 17 volunteers in
another trial; lc-inulin was determined to be a suitable substrate for measurement of orocaecal
transit time via the hydrogen breath test. In another study (Schneider et al. 2007), ingestion of

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5 g lc-inulin was found to cause a rise in breath hydrogen excretion in 27 of 29 volunteers, and
again was determined to be an appropriate substrate for measurement of small bowel transit
times.
In summary, results from studies in humans provide evidence that inulin, oligofructose and FOS
are largely undigested in the upper gastrointestinal tract and are not absorbed. Nearly all of the
ingested oligomers are delivered to the colon. Because inulin and its related 2-1 fructans are
largely non-digestible, they can be regarded as dietary fibers. In vivo data on the digestibility of
lc-inulin were not identified, though given that lc-inulin contains the same characteristic
glycosidic linkage as inulin with shorter chain lengths, it is reasonable to assume that lc-inulin
also is largely undigested and functions as a dietary fiber.

5.4

Fermentation of lc-Inulin and Selective Stimulation of Intestinal Bacteria

More than 90% of ingested inulin is estimated to arrive in the colon (Van Loo et al. 1999).
Non-digested carbohydrates that reach the colon intact contribute to the fermentable load.
Fermentation of non-digested carbohydrates in the colon produces short-chain fatty acids such
as acetate, propionate and butyrate; gases including hydrogen, carbon dioxide, sulfur and
methane; and organic acids such as lactate, succinate and pyruvate (Roberfroid et al. 1998).
The increased concentration of short-chain fatty acids and organic acids in turn lowers the
colonic pH.

5.4.1 In Vitro Evidence

In vitro studies provide evidence of the fermentation of inulin, oligofructose or FOS and selective
stimulation of bifidobacteria. These studies have been conducted using pure culture
fermentation or using batch or continuous culture systems of human feces. Roberfroid and
colleagues (1998) reviewed the fermentation of 2-1 fructans and determined that all are well
fermented. Studies of the in vitro fermentation of lc-inulin are summarized in Table 5-1 and
below.
Based on in vitro studies in minimal nutrition media, Biedrzycka and Bielecka (2004) reported
that highly purified inulin with a higher degree of polymerization (22 or 23) was less able to
support the growth of Bifidobacterium longum and B. animalis as compared to FOS,
oligofructose and inulin.
Corradini and colleagues (2004) demonstrated the degradation and fermentation of FOS
(DP 3-10) and lc-inulin (DP 25) using Bifidobacterium and fecal cultures. In two different
cultures, one of B. cuniculi MB280 and one of B. adolescentis ATCC151703, both FOS and lcinulin presented degradation patterns that were closely correlated with bifidobacterial growth of
both strains. In order to evaluate the consumption of the fructans, anaerobic fermentation
vessels were inoculated with fecal samples. Analysis suggested that the fermentation of FOS
and lc-inulin resulted in the complete consumption of these carbohydrates. Mean concentration
of bifidobacteria increased from a baseline level of 1.9 x 105 to 3.7 x 107 and 1.2 x 107 for FOS
and lc-inulin samples, respectively.
Using fecal batch cultures, Langlands and colleagues (2004) found a mixture of oligofructose
and lc-inulin (mixture DP 10) to significantly increase mean ( SEM) surface counts of

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bifidobacteria compared to a control culture (7.3 0.2 log10 CFU/slide vs. 6.6 0.2 log10
CFU/slide), respectively. Mean concentrations of acetate and butyrate were also significantly
higher compared to the control (200 13.9 log10 CFU/slide vs. 90 4.1 log10 CFU/slide for
acetate and 89 5.3 log10 CFU/slide vs. 42 2.0 log10 CFU/slide for butyrate), respectively.
Other bacteria and SCFA concentrations evaluated in the study were not significantly altered as
a result of FOS fermentation.
Perrin and colleagues (2002) compared the fermentation of FOS (92% DP 2-20), native FOS
(60% DP 2-20) and lc-inulin (70% DP > 20) and observed that the lc-inulin had a longer rate of
consumption whereas FOS (92% DP 2-20) was consumed most rapidly.
Rossi and colleagues tested FOS (DP 3-10), lc-inulin (DP 25) and a combination of the two
prebiotics in both pure and fecal batch cultures. The substrates were analyzed in 55 different
Bifidobacterium strains, representing 11 species. Whereas FOS was fermented by most
strains, only eight grew in the lc-inulin substrate. The investigators also observed a straindependent capability to degrade fructans of different lengths.
Overall, results from these in vitro studies of lc-inulin provide evidence that the polymer is
fermented by bifidobacteria and that the rate of fermentation of lc-inulin is slower as compared
to fermentation of shorter chain fructans.
Table 5-1. In vitro Studies of Fermentation of Long-Chain Inulin
Reference

Substrate
FOS (DP 2-4)
Oligofructose (DP 2-8)
Inulin (DP 9)
Inulin-EXL (DP 22)
Inulin HP (DP 23)
Raftilose Synergy:
Raftiline HP (DP 25)
Raftilose P95 (DP 3-10)
Chicory scFOS
(70% DP 2-10)
Chicory lc-inulin
(50% DP 10)
Raftiline HP &
oligofructose
(mixture DP 10)

Study Design
Evaluated growth of
three strains of
Bifidobacterium
longum and ten strains
of B. animalis
Bifidobacterium and
fecal culture
fermentation
Metabolism/breakdown
of substrates by three
strains of
Bifidobacterium

Perrin et al.
2002

scFOS (5% DP >20)

Native chain FOS
(30% DP >20)
lc-inulin (70% DP >70)

Metabolism/breakdown
of substrates by three
strains of
Bifidobacterium

Rossi et al. 2005

FOS (DP 3-10)

Made of: Raftilose
Synergy & Raftilose P95
Raftiline HP (DP 25)

Pure and fecal batch

cultures using 55
Bifidobacterium strains

Biedrzycka and
Bielecka 2004
Corradini et al.
2004

Durieux et al.
2001
Langlands et al.
2004

Batch culture
fermentation

Results
Highly polymerized and highly
purified inulins were not as well
fermented as FOS, oligofructose,
and inulin.
Increase in bifidobacteria
concentration and decrease in pH.
FOS substrates fully consumed by
all test strains.
Substrates increased
bifidobacteria and concentrations
of butyrate and acetate.
Substrates with lower DP
consumed first. Increased DP
resulted in increased residual
FOS and longer rate of
consumption.
Almost all strains grew on FOS
while only eight grew on inulin.
Butyrate predominantly produced
by inulin; acetate and lactate
predominantly produced by FOS.

Abbreviations:
DP = Degree of Polymerization; FOS = fructooligosaccharide; lc-inulin = long-chain inulin

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5.4.2 In Vivo Evidence

Studies in humans provide additional evidence of the fermentation of inulin, oligofructose and
FOS in the colon. Studies of the fermentation of inulin and related fructans have been reviewed
and results indicate that ingestion of the substances is associated with increased numbers of
potentially health promoting colonic bacteria such as bifidobacteria (Roberfroid 2007). The
effects of lc-inulin or combinations of lc-inulin and oligofructose on microbial populations are
summarized below and in Table 5-2. The effects of lc-inulin fermentation on laxation are
summarized later in the document along with tolerance of lc-inulin.
Tuohy and colleagues (2001) evaluated the effects of Raftiline HP (lc-inulin) ingestion on fecal
microflora composition using in situ hybridization (FISH) enumeration techniques. Ten healthy
volunteers (five male, five female, 20 55 years old) consumed 8 g maltodextrin per day for
14 days. Over the following 14 days, the volunteers consumed 8 g lc-inulin/d. Fecal counts
measured after the period of maltodextrin consumption were compared to counts after 7 and
14 days of lc-inulin consumption. Nine of the ten volunteers completed the trial (reason for
dropout not available). Mean fecal populations of Bifidobacterium spp. increased significantly
after seven days of lc-inulin consumption compared to the maltodextrin period (8.9 0.8 vs
8.8 0.9 log10 cells/g, mean standard deviation) and levels remained elevated following
another week of lc-inulin consumption (9.0 0.6 log10 cells/g; not significantly different from
placebo or week one levels). Individuals with lower initial fecal bifidobacterial counts were
observed to have an increased response to lc-inulin ingestion. Mean fecal populations of
Bacteroides spp. and lactobacilli/enterococci did not change during the lc-inulin feeding period,
though total bacteria increased from 10.2 0.2 to 10.4 0.3 log10 cells/g after one week of
lc-inulin intake. A statistically significant increase in fecal Clostridium spp. was measured after
14 days of lc-inulin consumption as compared to levels following maltodextrin consumption
(7.6 0.5 vs 7.2 0.7 log10 cells/g); the investigators determined the increase to be of limited
clinical value given the small magnitude of change. The investigators speculated that lc-inulin is
not completely fermented in the proximal region of the colon, and therefore more of the
nondigestible carbohydrate may be available for fermentation in the distal region.
Fecal bifidobacteria counts in adults consuming 10 g lc-inulin daily for one week were not
different from counts in the placebo group (Bouhnik et al. 2004) as assessed by microbiological
plating techniques. The lc-inulin ingestion had no effect on counts of total anaerobes,
lactobacilli, Bacteroides or enterobacteria.
In the study by Langlands et al. (2004), biopsy samples from the cecum and descending colon
were used to analyze mucosal gut microflora in response to diet. In the study, 14 adults were
administered a total of 15 g lc-inulin and oligofructose (7.5 g of each) daily for two weeks and
other group of 15 adults served as the control group. Total anaerobe counts were unaffected by
the addition of the prebiotic mixture to the diet. Changes in total Bacteroides numbers or
species distribution were also not observed. There was a significant increase in eubacteria and
lactobacilli in both the proximal and distal colon for the prebiotic group compared to the control
at the end of the study. Bifidobacteria concentrations significantly increased compared to the
control, but only in the distal colon.

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In summary, results from these studies indicate that daily intake of 8 g lc-inulin, 7.5 g lc-inulin
plus 7.5 g oligofructose or 9 g of a 9:1 combination of GOS/lc-inulin is associated with increased
populations of bifidobacteria in feces and the mucosa. No change in fecal bifidobacteria counts
was seen after one week of daily intake of 10 g lc-inulin. Ingestion of lc-inulin had little impact
on other microbial populations including bacteroides, lactobacilli/enterococci, coliforms and
clostridia.
Table 5-2. Human Studies of Long-Chain Inulin Effects on Microflora
Reference

Bouhnik et al.
2004

Langlands et al.
2004

Treatment
Population
n=8 per
treatment; mean
30 y

10 g lc-inulin/d
consumed in two
equal portions

Healthy
volunteers

Source: Raftiline
HP, Orafti

n=14; mean 59 y

7.5 g lc-inulin & 7.5

g oligofructose
consumed in three
equal portions

Subjects on
colonoscopy
waiting list

n=17; 23-48 7
Shadid et al 2007

Pregnant women,
third trimester of
pregnancy

n=9; 20-55 y
Tuohy et al. 2001
Healthy
volunteers

Dose

Treatment
Duration

1 wk

2 wk

Source: Raftiline HP
and Raftilose P95;
Orafti
0.9 g lc-inulin and
8.1 g GOS/d
consumed in three
equal portions
8 g lc-inulin/d
consumed in two
equal portions
Source: Raftiline
HP, Orafti

Approx 15
wk

14 d

Effects
-No change in fecal
bifidobacteria in lc-inulin
group.

-Higher levels of
eubacteria in proximal and
distal colon.
-Increase in bifidobacteria
in distal colon.
-No changes in total
anaerobes clostridia,
Bacteroides or coliforms.
-No changes in
proliferation indices.
-Increased percentage of
bifidobacteria in maternal
fecal samples.
-No effect on fecal
lactobacilli.
-Increase in fecal
bifidobacteria and total
bacteria counts after 7 d;
increase in fecal clostridia
after 14 d.
- No effect on fecal
bacteroides or
lactobacilli/enterococci.

Abbreviations:
d = day; g = grams; GOS = galacto-oligosaccharide; lc-inulin = long-chain inulin; y = year.

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mechanism. The authors speculated that inulin may have activated Toll-like receptors and the
innate immune system. The authors concluded that hypersensitivity to inulin is rare, mostly
benign, and of an unknown mechanism.

5.5.3.2 Review of Allergenicity Findings

In response to the case reported by Gay-Crosier et al. (2000), Orafti reported that further testing
was conducted in an attempt to better understand the reported reaction of the male patient.
Orafti reported that in the same patient, no allergic reactions to foods containing oligofructose
were observed. In addition, evaluations of Raftiline products by SDS-PAGE or Coomassie Plus
Protein Assay did not detect any proteins, which are known to be associated with inducing
allergic responses, as opposed to polysaccharides.
Since 2003, Danone has maintained a registry of adverse events attributed to the use of their
products in European markets. Over a nearly three-year period beginning in November 2005
(the oldest available records), approximately 200 consumer complaints were registered under
the keyword allergic reactions. Danone modified its registry program though information from
2007 to 2012 is consistent with the earlier information. The majority of the complaints are in
regard to infant formulas or exempt infant formulas (powder products), a few are for liquid
products (toddler formulas), and none are concerned with medical foods. The complaints
regarding infant formula (non-exempt) primarily refer to disturbances in gastrointestinal function
while one reports heavier breathing and some report an allergic reaction or an allergic skin
reaction without additional details. The complaints regarding exempt infant formula include
reports of skin rash and wheezing, often in infants who had recently started using hydrolyzed
formula containing GOS/lc-inulin. Hydrolyzed formulas are for infants who are, or suspected to
be, intolerant to cows milk protein. For infant formulas based on hydrolysates claiming a
reduction of risk to allergy to milk proteins, European legislation requires the product label to
state that the product is not for infants allergic to the intact proteins from which it is
manufactured unless generally accepted clinical tests demonstrate tolerance in more than 90%
of infants (confidence interval 95%) hypersensitive to proteins from which the hydrolysate is
manufactured 1. In other words, up to 10% of infants starting on infant formula based on protein
hydrolysates may experience intolerance problems to the milk protein component of the
product. These infants are generally recommended to move to a semi-elemental product or, in
rare cases where problems persist, an amino acid based formula. Danone uses GOS/lc-inulin
in hydrolyzed formulas (both protein hydrolysates and semi-elemental products) but not in
amino acid based formulas since these formulas are free from ingredients of dairy origin.
Although it is not possible to detect whey protein in GOS, the manufacturing process of GOS
does not exclude the possibility that trace amounts of native whey protein may remain in the
ingredient. In hydrolyzed formulas, GOS is therefore subjected to the same enzymatic
hydrolysis step as the native cows milk protein used in the product.
It is important to note that the registry of adverse events are self-reported incidents with little
detail about the actual products ingested or other potential sources of allergens that the
individual may have been exposed to. In addition, no adverse effects suggestive of an allergic
1

Commission Directive 2006/141/EC; Official Journal of the European Union L 401/1, 30.12.2006; See Annex IV;
http://eur-lex.europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2006:401:0001:0001:EN:PDF

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reaction were observed in a double-blind, placebo controlled clinical study of 249 infants
suffering from atopic dermatitis (AD) (Moro et al., 2006). In fact, there were fewer reports of AD
infants in the group that received formula containing 0.8 g GOS/FOS per 100 ml (10/102 (9.8%)
than the control group that received maltodextrin-containing formula (24/104, 23%).
In summary, inulin-containing foods have been in the food supply for more than a decade.
During this time, two case reports of allergic reactions to inulin in foods in adults with artichoke
allergies have appeared in the publicly available literature. The available evidence indicates
that it is reasonable to conclude that the risk of allergic reactions to inulin in foods, including
infant formula and medical foods, is extremely small.

5.5.3.3 Human Tolerance of lc-Inulin and Effects on Laxation

Previous reviews of clinical studies of inulin and oligofructose ingestion have indicated that
humans can comfortably consume 20 g of the substances or more daily (Carabin and Flamm
1999; ENVIRON 2002). Potential adverse effects resulting from excessive intake are typically
gastrointestinal disturbances including bloating, flatulence, and abdominal distention. The
effects are generally mild in nature and reduced when the oligosaccharide is consumed in
divided doses throughout the day. Continued exposure to the oligosaccharide also tends to
result in reduced gastrointestinal symptoms as a consequence of adaptation. Longer chain
fructans are more slowly fermented than shorter chains (e.g., scFOS) and therefore tend to be
better tolerated. Fermentation of inulin and oligofructose in the colon also has been shown to
increase stool bulk, fecal water content, and frequency of defecation. Studies in which the
tolerance of lc-inulin and effects on laxation were assessed are summarized below and in Table
5-3.
Hond et al. (2000) studied the effects of the administration of lc-inulin (Raftiline HP) on healthy,
constipated adults in a double-blind, placebo controlled cross-over study. Six participants, one
male and five females, with a mean age of 28.5 years were enrolled in the study. None of the
participants had undergone abdominal surgery or had taken medication that could influence
intestinal motility or flora in the three months prior to study initiation. All participants went
through a washout period of one week at the initiation of the study. During week 2, subjects
were randomly assigned to consume lc-inulin (n=3) or sucrose, the placebo, (n=3). Week
3 consisted of another washout period. In week 4, participants switched treatments. Each
participant was administered 5 g lc-inulin or placebo, three times per day, for a total daily intake
of 15 g/d. By the end of the study, each participant had consumed lc-inulin for a period of one
week. Participants were instructed to consume a standard diet with the exclusion of any pre- or
probiotics. During the study weeks (lc-inulin and placebo administration), subjects maintained a
diary on fecal habits and gastrointestinal complaints. The mean number of stools significantly
increased after lc-inulin supplementation compared to baseline measurements (from 4.0 per
week at baseline to 6.5 per week after lc-inulin supplementation). For each individual, an
increase of at least one stool per week was observed. No significant increases in mean stool
weight, gastric emptying rate, orocecal transit time or total transit time were observed. During
the lc-inulin period, average complaints were made 0.8 and 1.2 times per week for flatulence
and abdominal cramps, respectively. No reports of flatulence or abdominal cramps were
reported for participants during the placebo period. This difference did not reach statistical
significance and researchers noted a large inter-individual variation for these gastrointestinal

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parameters. Intestinal permeability, expressed as the 6-hour urinary percent dose excretion of
[51Cr]EDTA also did not change after ingestion of lc-inulin.
Bouhnik and colleagues (2004) monitored the digestive symptoms in healthy volunteers
consuming one of seven types of nondigestible carbohydrates or a placebo (n = 8 per group).
The nondigestible carbohydrates provided to study participants included lc-inulin (Raftiline HP)
and FOS (53% DP 3). Participants consumed 10 g of each test substance daily in two divided
doses (5 g after lunch and dinner) for one week. Significant increases in excess flatus, bloating,
borborygmi and abdominal pain were reported by participants in each group, including the
placebo group, though there were no differences in gastrointestinal symptoms among all study
groups (Bouhnik et al. 2004). No significant change was observed in the number of daily stools
and diarrhea was not reported in any of the groups.
In a study by Langlands and colleagues (2004), 14 adult subjects consumed a mixture of 7.5 g
lc-inulin (Raftiline HP) and 7.5 g oligofructose (Raftilose P95) daily for two weeks; the mean
DP of the mixture was 10. Compliance was reportedly high, 97 percent in the treatment group,
and the majority of gastrointestinal complaints including flatulence and bloating were mild to
moderate in nature.
Results from these studies indicate that a total intake of 10 to 15 g of lc-inulin (or a combination
of lc-inulin and oligofructose) is well tolerated by adults. The noted side effects included
flatulence, bloating or abdominal pain. Daily intake of 15 g lc-inulin was associated with
increased stool frequency.
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Table 5-3. Human Studies of Tolerance of Long-Chain Inulin

Reference

Bouhnik et
al. 2004

Treatment
Population

n=8; mean 30 y
Healthy volunteers

n=6; mean 28.5 y

Hond et al.
2000

Healthy,
constipated adults

n=14; mean 59 y
Langlands
et al. 2004

Subjects on
colonoscopy
waiting list

n=17; 23-48 y
Shadid et
al. 2007

Pregnant women,
third trimester of
pregnancy

Treatment
Duration

Dose

10 g lc-inulin/d
consumed in two equal
portions

1 wk

Source: Raftiline HP
(Orafti)
15 g lc-inulin/d
consumed in three
equal portions

1 wk

Source: Raftiline HP;

Orafti
7.5 g lc-inulin & 7.5 g
oligofructose
consumed in three
equal portions

2 wk

Source: Raftiline HP
and Raftilose P95;
Orafti
0.9 g lc-inulin and 8.1
g GOS/d
consumed in three
equal portions

Approx 15
wk

Prepared for Danone Trading B.V.

L. debueckii. At the end of the 6-week study, the investigators also noted that bifidobacteria and
lactobacilli accounted for 80% of fecal microflora in the breast-fed and GOS/lc-inulin groups
while lactobacilli and bifidobacteria accounted for 50% of fecal microflora in infants fed the
standard formula.
Moro et al. (2002) studied the effects of consumption of a formula supplemented with GOS and
lc-inulin on infant microflora. Ninety term infants between 6.3 to 7.2 days old were randomly
assigned to one of three formula groups: standard formula (n=33), formula supplemented with
0.4 g lc-inulin per L and 3.6 g GOS per L (n=30), or formula with 0.8 g lc-inulin per L and 7.2 g
GOS per L (n=33). Infants consumed the assigned formula for four weeks. A dose-dependent
increase in fecal bifidobacteria was observed in the GOS/ lc-inulin groups, with baseline levels
of 8.5 CFU/g and 7.7 CFU /g, respectively, and levels of 9.3 CFU /g and 9.7 CFU /g,
respectively by the end of the study period. Fecal lactobacilli increased in the GOS/ lc-inulin
groups as compared to the control group, but there was not a difference between the two GOS
groups. A dose-dependent decrease in stool consistency was found; while stool frequency
increased and fecal pH decreased only in infants consuming formula supplemented with 0.8 g
lc-inulin per L and 7.2 g GOS per L. Weight gain and length increment were similar among
groups, and the diets did not influence the incidence of crying, regurgitation or vomiting, and no
infant had diarrhea during the study period. Fecal samples from infants in the high-dose group
(0.8 g lc-inulin per L and 7.2 g GOS per L) and the control groups were examined for the
presence of GOS and lc-inulin (Moro et al. 2005). The oligosaccharides were detected in stool
samples from all infants consuming the GOS/ lc-inulin -supplemented formula and none of the
infants in the control group. The amounts of GOS and lc-inulin detected in the feces were not
quantified. The investigators noted that human milk oligosaccharides also can be detected in
feces of infants consuming human milk.
Schmelzle and colleagues (2003) studied the effects of a formula containing partially hydrolyzed
protein, a high level of -palmitic acid and nondigestible oligosaccharides (GOS/lc-inulin)
versus a standard formula in a population of healthy, term infants. Infants consumed the
assigned formula from or before age 2 weeks until the age of 12 weeks. There were no
differences between groups in the number of dropouts or reasons for dropping out. Infants
consuming the new formula with added GOS/lc-inulin were found to have increased fecal
bifidobacteria compared to baseline levels, a higher percentage of fecal bifidobacteria as
compared to infants in the control group, and a higher proportion of softer stools. Formula
intake and energy intake per kg bodyweight were lower in infants consuming the supplemented
formula versus the control formula, though there were no differences at 12 weeks between
groups in weight gain.
In another study involving a formula containing hydrolyzed protein, -palmitate and GOS/lcinulin, 14 infants consumed the test formula from birth to 2 months of age (Rigo et al. 2001).
Infants consuming breast milk or standard formulas in previous studies were used as reference
controls. Growth and anthropomorphic development of the infants followed a standard healthy
infant growth curved throughout the study. Infants consuming the new formula had a higher
percentage of endogenous bifidobacteria compared to baseline levels when measured at 25,
45, and 68 days of age. One dropout was reported in the study due to gastro-esophageal
reflux.

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Bakker-Zierikzee et al. (2005) studied the effects of a GOS/lc-inulin-supplemented infant

formula versus a probiotic formula containing Bifidobacterium animalis on the microflora of
healthy infants over the first 16 weeks of life. Within three days post-partum, 19 infants were
enrolled in each of the three formula groups (0.6 g lc-inulin per L and 5.4 g GOS per L, 6 x 1010
CFU B animalis per L, or control). Sixty-three breast-fed infants were followed as a reference
group. Infants consuming the GOS/lc-inulin supplemented formula were found to have a lower
fecal pH, higher percentages of fecal acetate and lower percentages of propionate, butyrate and
isobutyrate, isovalerate and valerate compared with infants consuming the control formula. No
differences among groups were found in the percentage of fecal bifidobacteria. During the
intervention, fecal sIgA levels were higher in infants consuming the GOS/lc-inulin supplemented
formula as compared to the control formula (Bakker-Zierikzee et al. 2006). All formulas were
reportedly well accepted and tolerated.
Savino et al. (2003, 2006) conducted two studies on older infants experiencing minor
gastrointestinal problems including colic. The first study was an observational study in which
604 infants with an average age of 1.35 months consumed a partially hydrolyzed formula
containing GOS/lc-inulin (0.8 g lc-inulin per L and 7.2 g GOS per L), palmitic acid in the
position and low levels of lactose for a period of two weeks (Savino et al. 2003). At the end of
the feeding period, infants experienced fewer episodes of colic, fewer regurgitation problems,
and less constipation. Reductions in the incidence of crying in infants with colic also were
observed in a randomized, controlled trial of infants consuming either a formula with partially
hydrolyzed whey proteins, GOS/lc-inulin (0.8 g lc-inulin per L and 7.2 g GOS per L) and a high
-palmitic acid level or a standard formula and simethicone (Savino et al. 2006).
Rinne et al. (2005) conducted a trial that evaluated microflora of infants at 6 months of age with
a family history of atopic eczema, allergic rhinitis or asthma. Inclusion criteria required infants to
have been on their respective formulas since at least 2 months of age. Of the 32 infants in the
study, eight consumed the GOS/lc-inulin formula and the remaining infants consumed a
standard formula, breast-milk or a probiotic-supplemented formula. Infants consuming the
GOS/lc-inulin-supplemented formula had increased fecal bifidobacteria after four months, and a
lower occurrence of atopic eczema compared to the standard formula group at 10 months.
Weight and length of infants were comparable among all formula groups.
Moro and colleagues (2006) also tested GOS/lc-inulin formula on infants at high risk of
developing atopy. Inclusion criteria required infants to have started formula feeding within the
first two weeks of life. Study visits were scheduled for each infant at 3 and 6 months of age. Of
the 206 infants completing the study, 102 were in the GOS/lc-inulin group with the remaining in
a maltodextrin placebo group. In a subset of 98 of the infants, 50 coming from the GOS/lc-inulin
group, fecal samples were analyzed. Infants consuming the GOS/lc-inulin formula were found
to have increased fecal bifidobacteria levels at 3 and 6 months of age as well as an increase in
stool frequency and softer stool consistency. Infants in the treatment group also had a
significantly lower incidence of dermatitis. Reasons for dropouts were similar between groups,
with reports of regurgitation and crying significantly lower in the GOS/lc-inulin group.
Alliet and colleagues (2007) studied the impact of a GOS/lc-inulin formula on serum cholesterol
and triacylglycerol levels in 8-week old infants. The treatment group included 75 infants

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followed until 26 weeks of age, for a total treatment period of 18 weeks. Since infants were not
restricted from breastfeeding, a sub-group of infants that consumed formula for 97.5% of their
total number of feedings (the number of infants in this sub-group was 22) was also followed.
Infants in the treatment group consumed 5.4 g/L GOS and 0.6 g/L lc-inulin. They were
compared to a control formula group (that also had a sub-group of high-compliance infants) and
an exclusively breast-fed group. For analytical purposes, both treatment formula and standard
formula groups were analyzed both as an entire group and with just the high-compliance infants
with the breast-fed group. Lipid levels were measured at 8 and 26 weeks of age and compared
between groups. The investigators observed no difference in median HDL or triacylglycerol
levels among groups at either of the time points measured in each. Median cholesterol and LDL
levels were significantly lower for both the entire and sub-group of infants in both formula groups
compared to the breast-fed infants. The median cholesterol levels for the entire GOS/lc-inulin
group and the entire control group at 26 weeks of age were and 130.00 and 121.50 mg/dL,
respectively while breast-fed infants had a median cholesterol level of 146.00 mg/dL at the
same time point. Median LDL levels for the entire treatment and control groups were 70.00 and
66.00 mg/dL compared to 86.00 for the breast-fed group. Of the 215 infants recruited,
187 completed the study.
One infant study was identified that involved the administration of GOS/lc-inulin as a solid
weaning food (Scholtens et al. 2006b). This randomized, double-blind, placebo-controlled trial
involved 35 fully formula-fed infants with an average age of 16 weeks. Two treatments were
administered: standard baby food with maltodextrin as the placebo and baby food with a
GOS/lc-inulin formulation. Nineteen infants were in the GOS/lc-inulin-supplemented group,
11 of whom were included in the per-protocol analysis (which specified compliance of 60%
consumption during the final two weeks of the study). Over the course of the 6-week study
period, infants consumed a total of 0.45 g lc-inulin and 4.05 g GOS per day in three equivalent
doses. Infants consuming the GOS/lc-inulin-supplemented food had a significant increase in
fecal bifidobacteria (43% at week 0 vs. 57% at week 6) above baseline levels, indicating the
ability of supplemental GOS/lc-inulin to alter fecal microflora in the weaning period.
Costalos and colleagues (2007) studied the impact on infant growth, tolerance and fecal
microflora of a GOS/lc-inulin formula in 70 healthy term infants 14 days of age. Infants
consumed 0.4 g lc-inulin per L and 3.6 g GOS per L for 12 weeks. Anthropometric
measurements were conducted at study entry and repeated at age 6 and 12 weeks. Stool
samples were taken at 6 weeks from 32 randomly selected infants. Mothers kept a diary and
questionnaire in which intake of formula and stool characteristics were recorded. Results were
compared to a control group of 70 infants. In total, 140 of the 160 infants originally enrolled
completed the study. Ten infants from the GOS/lc-inulin-group and ten from the control group
dropped out of the study. No clinically relevant adverse events related to the study formula were
reported. Nine infants (5 in the GOS/lc-inulin group and 4 in the control group) dropped out due
to change in formula and seven (4 in the GOS/lc-inulin and 3 in the control group) dropped out
due to minor gastrointestinal complaints. Four infants dropped out of the study for unknown
reasons. Measurements of weight gain, length gain and head circumference gain did not differ
between groups at 6 or 12 weeks of age. The GOS/lc-inulin infants experienced an increase in
stool frequency and consistency. No difference between groups with regard to infant
satisfaction, tolerance or occurrence of possetting was reported. No changes in the median

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of different doses of prebiotic supplemented formulas as compared to standard formula and

breast fed infants. The authors evaluated the results of parental diaries that recorded formula
intake, stool frequency and consistency, crying behavior, occurrence of regurgitation or
vomiting. The authors also evaluated anthropometric data (length, weight, and head
circumference) and the fecal bacteria in stool samples collected on days 3, 14 and 28 (flora
analysis).
Veereman-Wauters et al. (2011) found no significant differences between groups for
anthropometric parameters, stool frequency, crying behavior, regurgitation or vomiting. The
authors found that prebiotic supplement at 8 g/L of formula contributed to softer stools with a
consistency resembling that of the breast fed infants. The authors also found infants receiving
the 8 g/L prebiotic supplemented formulas also had comparable levels of bifidobacteria as the
breast fed group. Veereman-Wauters et al. (2011) reported that no serious adverse events
occurred during the study. Non-serious issues included hunger, regurgitation, and symptoms of
gastroesophageal reflux. The number of dropouts was evenly distributed across groups (6 8
per group) including controls leading the authors to conclude that the intervention was not a
contributing factor.
In summary, results from 43 studies of healthy, term infants consuming formula supplemented
with up to 0.8 g lc-inulin and 7.2 g GOS per L for periods ranging from two weeks to up to twelve
months provide evidence that lc-inulin, in combination with GOS, is well tolerated by infants and
produces no adverse effects. The supplemented formulas supported normal growth.
Additionally, results from these studies suggest that formulas with added lc-inulin in combination
with GOS may influence shifts in infant gut microflora that result in gut microflora more similar to
the gut microflora of breast-fed infants.

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Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Alliet et al.
2007

To study the effects

of a 9:1 GOS/lc-inulin
mixture on
cholesterol and
triacylglycerol levels
in infants.

Treatment
Population a

n = 75;
age = 8 wks

Arslanoglu
et al. 2007

Arslanoglu
et al. 2008

Randomized, doubleblind, placebocontrolled.

To investigate the
sumulative incidence
of allergic
manifestations,
infectious episodes,
and growth.

Formula with 5.4 g

GOS/L & 0.6 g lcinulin/L

Treatment
Duration

18 weeks

Source: Numico (now

Danone)

Randomized, doubleblind, controlled.

To investigate the
effects of a prebioticsupplemented
formula on infections
in infants with a
history of atopy.

Test Substance and

Dose

n = 206;
age = 2 wks

8 g/L of scGOS/lcinulin; 9:1 ratio

6 months

Source: Numico

Randomized, doubleblind, placebocontrolled.

Safety Data

n = 134;
age = up to 2 yrs

8 g/L of scGOS/lcinulin; 9:1 ratio for

first 6 months of life
Source: Numico

6 months

Results of GOS/lc-inulin Ingestion

Growth: Only baseline anthropometric data given.
Tolerance: Not measured.
Dropouts: 86 infants entered GOS/lc-inulin treatment
group and 75 completed the study; no explanation of
dropouts provided.
Other Endpoints:
-No difference in levels of total cholesterol and LDL
cholesterol between GOS/lc-inulin and standard
formula-fed infants; total cholesterol and LDL cholesterol
for both formula groups lower than breast-fed infants.
-No difference in HDL cholesterol or triacylglycerols
among groups.
Growth: No difference in growth.
Tolerance: Not stated.
Dropouts: same as in placebo group (5); flatulence,
hard stools, relocation, or no reason.
Other Endpoints:
-Fewer infection episodes.
-Lower cumulative incidence of recurring infection.
-Lower cumulative incidence of recurring respiratory
infection.
-Increased bifidobacteria.
Growth: No difference in growth.
Tolerance: Not stated.
Dropouts: See Arslanoglu et al. 2007.
Other Endpoints:
-Lower incidences of recurrent wheezing, atopic
dermatitis, and allergic urticaria.
-Fewer episodes of infections and fever.
-Fewer antibiotic prescriptions.

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Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

BakkerZierikzee et
al. 2005,
2006

Bisceglia et
al. 2009

Study the effects of

GOS/lc-inulin on fecal
microflora and SIgA
levels.

Treatment
Population a

n = 19;
age = 3 days
postpartum

Randomized, doubleblind.

To evaluate ability of
prebiotics to
modulate jaundice in
otherwise healthy
term newborns.
Prospective,
randomized, doubleblind, placebocontrolled.

Test Substance and

Dose

Treatment
Duration

Formula with 5.4 g

GOS/L & 0.6 g
lc-inulin/L
16 weeks
Sources: Friesland
Foods Domo (GOS);
Orafti (FOS)

Formula
supplemented with
0.8g/dL of a 9:1
scGOS/lc-inulin or
maltodextrin placebo.

n = 76

Formula was
provided when breast
milk was not
available; therefore,
the precise doses for
each newborn are
unknown

28 days

Results of GOS/lc-inulin Ingestion

Growth:
-Data not measured/reported.
Tolerance:
-No effect on fecal consistency or frequency.
Dropouts:
-5 infants dropped out of the GOS/lc-inulin group; 4
dropped out of the control group; and 4 dropped out of
the Bb-12 group. Reasons for dropping out included
colic, suspicion of cows milk allergy, constipation, and
practical problems. Reasons by group were not
specified.
Other Endpoints:
-No change in % fecal bifidobacteria.
-Decreased fecal pH.
-No change in total fecal SCFA; increased % fecal
acetate, decreased % fecal propionate, butyrate,
isobutyrate, isovalerate, valerate.
-No change in fecal lactate.
-Increase in median fecal SIgA concentration at week
16.

Growth: Gain in weight, length and head circumference

did not differ.
Tolerance: The authors reported no adverse effects in
either group.
Dropouts: 10 due to denied consent; 8 for unknown
reasons.
Other Endpoints:
-Beginning at 72 hours and at every point thereafter the
prebiotic-supplemented formula group had lower
bilirubin levels.
-Higher stool frequency.

Source: Numico

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Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Reference

Bruzzese
et al. 2009

Study Design &

Objective
To evaluate the
effects of prebiotics
on the incidence of
intestinal and
respiratory tract
infections and
anthropometric
measures.

Treatment
Population a

n = 201;
age = 15 120 d

Test Substance and

Dose

4 g/L GOS/lc-inulin
9:1 ratio
Source: Not reported.

Treatment
Duration

First 12
months of
life.

Growth: Temporary increase in body weight of

prebiotic-fed infants.
Tolerance: Softer stools in with prebiotics.
Dropouts: 56 due to lack of follow-up, 80 due to
protocol violations, 5 due to intolerance of cows milk
protein (342 started, 201 completed).
Other Endpoints:
-Lower incidence of gastroenteritis.
-Fewer infants had multiple antibiotic courses per year.

12 weeks

Growth:
-Gain in weight, length and head circumference did not
differ.
Tolerance:
-Increase in stool frequency and number of stool per
day.
-No difference in formula satisfaction, tolerance or
occurrence of possetting.
Dropouts:
-10 infants dropped out of both groups; no adverse
events related to study formula reported.
Other Endpoints:
-At 6 wk, median bifidobacteria, clostridia and E. coli
fecal proportions as a percentage of the total
microorganism count did not differ.

Prospective,
randomized, placebocontrolled, open.

Costalos et
al. 2007

To study the effects

of GOS/lc-inulin on
infant growth,
acceptability and
fecal microflora.
Randomized, doubleblind, placebocontrolled

Safety Data

n = 70;
age = mean 4.7
days

Formula with 3.6 g

GOS/L & 0.4 g lcinulin/L

Source: Numico

Results of GOS/lc-inulin Ingestion

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Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Fanaro, et
al. 2005

Firmansya
h et al.
2011

To study tolerance
and the effects of
formula with AOS or
GOS+FOS/AOS on
intestinal flora and
stool characteristics
in term infants.

To determine if
Bifidobacterium
longum, Lactobacillus
rhamonosus,
prebiotics, and longchain
polyunsaturated fatty
acids (LCPUFA)
increased growth
versus a control milk.

Treatment
Population a

n = 46

n = 393

Test Substance and

Dose

3 treatments:
Standard formula,
formula with 5.4 g
GOS/L & 0.6 g lcinulin/L + AOS,
formula with AOS

Mixture of probiotics
(Bifidobacterium
longum, Lactobacillus
rhamnosus),
prebiotics (30% inulin
and 70% FOS),
LCPUFA or a control
milk.

Treatment
Duration

6 weeks

12 months

400 mL/day.
Randomized, doubleblind, placebocontrolled.

Grber et
al. 2010

To assess the effect

of prebiotics on
occurrence of atopic
dermatitis in infants.
Randomized, doubleblind, placebocontrolled.

Safety Data

Source: Not reported.

n = 1130;
age < 8 wks

6.8 g/L of scGOS/lcinulin (9:1 ratio) and

1.2 g/L pAOS in a
non-hydrolyzed cows
milk-based formula
Source: Nutricia

1 year

Results of GOS/lc-inulin Ingestion

Growth:
- No difference in growth patterns.
Tolerance:
-Softer stool consistency compared to control group.
-No difference in crying, vomiting or regurgitation
patterns.
Dropouts:
-None reported.
Other Endpoints:
-Increased fecal bifidobacteria compared to control and
AOS groups.
-Decreased pH compared to control and AOS groups.
Growth: pro-/prebiotics w/LCPUFA had higher weight
gain; length, head circumference did not differ.
Tolerance: No study related adverse events.
Dropouts: 38 dropped out of experimental milk group,
41 dropped out of control milk group.
Other Endpoints:
- There were no significant differences between groups
in anti-vaccine immunoglobulin G, mental and motor
development measures, cognitive and adaptive behavior
scores, stool characteristics, body mass index,
gastrointestinal tolerance.
Growth: No difference (see Piemontese et al. 2011).
Tolerance: No difference between groups.
Dropouts: 129 total; similar across groups; due to
withdrawal, occurrence of disease, protocol violations,
intolerance of formula, change of formula.
Other Endpoints:
-Lower incidence of atopic dermatitis than the placeboformula group, but not than the breast-fed group.
-No difference in severity of atopic dermatitis.

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lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Kim et al.
2007

Cross-over study to
investigate the effect
of native inulin in
formula-fed babies.

Safety Data

Treatment
Population a

n = 14
age = 12.6 6.4
wks

Test Substance and

Dose
0.25g/kg/day, 3
weeks inulin; followed
by 3 weeks without,
or vice versa.
Average intake of
inulin was 1.5( 0.3
g/d) milk-based
formula of 1.5 g OS
(raffinose, GOS &
inulin) or control of
same formula without
OS supplement for 3
weeks. After 3
weeks the group
switched formulas.

Treatment
Duration

Results of GOS/lc-inulin Ingestion

3 weeks

Growth: No differences in growth.

Tolerance: No study-related adverse effects.
Dropouts: None reported.
Other Endpoints:
-No significant difference in total # of anaerobic bacteria
or Bacteroides spp between treatment groups.
-Increase in Bifidobacterium spp cfu/g and Lactobacillus
spp in oligosaccharide treatment group.
-Consumption of inulin increased the content of
Bifidobacterium and Lactobacillus in the feces of
formula-fed babies, without affecting the number of
Bacteroides or the total anaerobic count.

000070

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Treatment
Population a

Test Substance and

Dose

Treatment
Duration

Results of GOS/lc-inulin Ingestion

Growth: Data not measured/reported.
Tolerance: Increase in median stool frequency. Mild
flatulence in all groups.
Dropouts: Eight dropouts; 3 from test group, 3 from
control group and 2 from reference group.

Knol et al.
2005a;
Haarman
and Knol
2005, 2006

Determine effects of
GOS/lc-inulin on
microflora in infants
exclusively formulafed since birth.

Randomized, doubleblind, placebocontrolled

intervention.

Safety Data

n = 15-19;
age = mean 7.7
wks

Formula with 7.2 g

GOS/L & 0.8 g
lc-inulin/L
6 weeks
Source: Aptamil,
Milupa

Reasons included rotavirus-infection (n=1), medical

problems of regurgitation (n=1) and formula-related
complaints (n=4; 2 from test and 2 from control).
Unknown reasons for drop outs in reference group.
Other Endpoints:
-Increased % fecal bifidobacteria.
-B. infantis, B. breve and B. longum remained
predominant species in GOS and HM groups.
-Decreased % B. adolescentis from baseline, decreased
%. B catenulatum from control.
-Increased % fecal lactobacilli.
-Increased L. acidophilus, L. paracasei and L. casei and
decreased L delbrueckii.
-At study completion fecal bifidobacteria and lactobacilli
accounted for 80% of fecal bacteria.
-Fecal bacteria more closely resembled HM-fed infants.
-Increased fecal acetate and decreased butyrate and
propionate.
-Decreased fecal pH.
-Increased fecal lactate and % D-lactate.

000071

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Lugonja et
al. 2010

Magne et
al. 2008

To study the
bifidogenic effect of
an infant formula
supplemented with
inulin and FOS
clinically and in vitro.

To test the safety and

effect of formula with
GOS, lc-inulin & AOS
on fecal microbiota in
infants at the age of
partial formula
feeding.
Double-blind,
placebo-controlled,
randomized
intervention.

Safety Data

Treatment
Population a

n = 21

Test Substance and

Dose

Formula
supplemented with
chicory-derived inulin
and FOS at 4.0 g/L.

Treatment
Duration

28 days

Growth: Gain in weight and length did not differ.

Tolerance: No significant adverse effects.
Dropouts: None reported.
Other Endpoints:
-Bifidobacteria levels in the stools from the formula-fed
infants were significantly higher than in the HM-fed
infants.
-The number of lactobacilli increased in both the
formula-fed and HM-fed infants.
-Increase in bifidobacteria and lactobacilli populations,
decrease in the total aerobes, anaerobes and yeasts
and fungi levels

2 months
partial
formulafeeding
period,
followed by
two month
formula
only
feeding
period

Growth: Weight gain did not differ.

Tolerance: No significant adverse effects.
Dropouts: 10 (did not discontinue breast feeding).
Other Endpoints:
-Bacteria group populations differed significantly after
formula-only feeding.
-There was an increase in bifidobacteria, decrease in
Bacteroides and C. coccoides in both the scGOS/lcinulin and scGOS/lc-inulin/AOS treatment groups, the
scGOS/lc-inulin/AOS had a significant increase over the
scGOS/lc-inulin group.

Source: Not reported.

n = 82

Formula with scGOS

and lc-inulin or
formula with scGOS,
lc-inulin and partially
unsaturated and
methylated AOS
Source: Numico

Results of GOS/lc-inulin Ingestion

000072

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Evaluate effects of
GOS/lc-inulin on
bifidobacteria.

Moro et al.
2002; 2005

Treatment
Population a

Test Substance and

Dose

Treatment
Duration

Formula with 3.6 g

GOS/L & 0.4 g lcinulin/L
n = 27-33; 6.3-7.2 d
postpartum

Randomized,
placebo-controlled

4 wk

7.2 g GOS/L & 0.8 g

lc-inulin/L
Source: Numico

Moro et al.
2006;
Arslanoglu
et al. 2007

Study the effect of

GOS/lc-inulin on the
incidence of atopic
dermatitis and
infections in high risk
infants.
Prospective, doubleblind, randomized,
placebo controlled

Safety Data

n = 102

Formula with 7.2 g

GOS/L & 0.8 g lcinulin/L
Source: Numico

< 6 mo

Results of GOS/lc-inulin Ingestion

Growth:
-No effect on growth.
Tolerance:
-Dose-dependent decrease in stool hardness, and
increase in stool frequency in high-dose group.
-No effect on incidence of crying, regurgitation, or
vomiting.
-No diarrhea occurred during study.
Dropouts:
-None reported.
Other Endpoints:
-Dose-dependent increase in fecal bifidobacteria, and
increase in fecal lactobacilli (not dose-dependent).
-Dose-dependent decrease in fecal pH.
-GOS and FOS detected in feces.
Growth: Data not measured/reported.
Tolerance:
-Increase in stool frequency and softer stool consistency
in treatment group.
-Lower reports of regurgitation and crying in treatment
group; incidence of vomiting same between groups.
Dropouts:
-53; 26 control (21 started breastfeeding, 5 other
reasons); 27 treatment (22 started breastfeeding, 5
other reasons).
Reasons for dropout not significant between groups.
Other Endpoints:
-Increase in bifidobacteria count in treatment group.
-Decrease in incidence of dermatitis in treatment group
compared to control group.
-Severity of dermatitis unaffected by diet.
-Fewer episodes of all types of infections, and lower
incidence of recurring infections.

000073

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Reference

Panigrahi
et al. 2008

Study Design &

Objective
To evaluate the longterm colonization
potential
Lactobacillus
plantarum in neonatal
guts.

Treatment
Population a

n = 31;
1-3 days post birth;
all cesarean
section.

Double-blind,
placebo-controlled,
randomized.

Piemontes
e et al.
2011

Puccio et
al. 2007

To assess the
tolerance and safety
of the prebiotics in
infants.

Prospective,
randomized, doubleblinded, referencecontrolled, parallelgroup clinical trial.

Safety Data

Receive either an
oral preparation of L.
plantarum and FOS
or dextrose saline
placebo for 7 days.

Treatment
Duration

6 months

Source: Not reported.

n = 1130;
age 8 wks

Double-blind,
placebo-controlled,
randomized.
To assess the safety
of an experimental
probiotic formula
containing B. longum
culture and
GOS:FOS.

Test Substance and

Dose

6.8 g/L of scGOS/lcinulin (9:1 ratio) and

1.2 g/L pAOS in a
non-hydrolyzed cows
milk-based formula

n = 138

Source: Nestl
Nutrition

Growth: No difference in % increase in weight.

Tolerance: No significant adverse effects reported.
Dropouts: 2 withdrawals; 1 protocol violation.
Other Endpoints:
-The number of bacteria species was significantly
higher.

1 year

Growth: No difference in growth between groups.

Tolerance: No difference in adverse events between
formula groups.
Dropouts: 129 total; similar across groups; due to
withdrawal, occurrence of disease, intolerance of
formula, change of formula.
Other Endpoints:
-No difference in mean weight, length, head
circumference, skin fold thickness, arm circumference
gains, & stool frequency between groups.

7 months

Growth: No difference in weight gain, length and head

circumference.
Tolerance: No study formula-related adverse events.
Dropouts: 41 (no statistical difference between control
and treatment groups).
Other Endpoints:
-Reduced incidence of constipation.
-Increased stool frequency.

Source: Nutricia

Experimental
probiotic formula
containing B. longum
culture (2 x 107 cfus)
and GOS/FOS (4 g/L)

Results of GOS/lc-inulin Ingestion

000074

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Raes et al.
2010

To evaluate the effect

of scGOS/lc-inulin on
basal immune
parameters.

Treatment
Population a

n = 215

Randomized, doubleblind, placebocontrolled.

Rao et al.
2009

Systematic review of
randomized control
studies.

Rinne et al.
2005

Assess the
quantitative and
qualitative differences
of microflora in
infants following
GOS/lc-inulin intake
of formula.

Safety Data

Test Substance and

Dose
Formula with 0.6
g/100 mL scGOC/lcinulin;
unsupplemented
formula or HM ad
libitum throughout the
study period.

Treatment
Duration

26 weeks

Results of GOS/lc-inulin Ingestion

Growth: Not reported.
Tolerance: No negative side effects from the prebiotics.
Dropouts: 28; no explanation reported.
Other Endpoints:
-No difference in supplemented groups in C-reactive
protein, total IgA, IgG, IgM & IgE.
-Breast fed group had higher white blood cell counts.

Source: Danone

Varied by study.
Not applicable.

n = 8; >8 wk
Infants had at least
one close relative
afflicted with atopic
eczema, allergic
rhinitis or asthma.

Sources: Varied by
study.

Not
applicable.

Formula with GOS/lcinulin

Dose not reported
Source:
Omneo, Nutricia,
Cuijk

4+ mo

Growth: Not evaluated.

Tolerance: All studies reviewed reported that the
substances were well tolerated.
Dropouts: Not applicable.
Other Endpoints:
-6 of 9 studies reported increased bifidobacteria counts.
-of 3 studies, all 3 reported higher stool frequency
(similar to breast fed, not an adverse outcome).
-Inadequate evidence regarding the prevention of atopic
diseases.
Growth:
-No differences in length or weight gain.
Tolerance:
-Clinical characteristics comparable among groups at 6
and 12 mo of age.
Dropouts:
-None reported.
Other Endpoints:
-Increase in fecal bifidobacteria after 4 mo; no effects on
fecal Clostridia, Lactobacilli/enterococci or Bacteroides.
-Lower occurrence of atopic eczema in breastfed and
pre/pro-biotic groups at 10 months.

000075

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Rigo et al.
2001

Salvini et
al. 2011

Study the effects of a

hydrolyzed protein, palmitic acid formula
with GOS/lc-inulin on
growth, miceral
accretion and
intestinal flora
development.

To determine if the
prebiotic
supplementation
could result in longterm colonization of
bifidobacteria.
Double-blind,
randomized, placebocontrolled.

Safety Data

Treatment
Population a

n = 14; at birth

Test Substance and

Dose

Formula with 3.6 g

GOS/L & 0.4 g lcinulin/L

Treatment
Duration

2 mo

Source: Numico

n = 20;
infants who were
not breast fed and
born to hepatitis C
virus- infected
mothers.

8 g/L scGOS/lc-inulin
supplemented
formula.
Source: Danone

6 months

Results of GOS/lc-inulin Ingestion

Growth:
-Growth parameters for study infants followed standard
infant growth curve.
Tolerance:
-Investigators reported feeding tolerance as being
excellent.
-Adequate volume formula intake observed throughout
study, although milk intakes were lower than those
previously observed in infants consuming a standard
formula.
Dropouts:
-One drop-out due to gastro-esphagial reflux.
Other Endpoints:
-Increase in the percentage of endogenous
bifidobacteria observed through 68 days of age (when
final measurement was taken).
Growth: Not evaluated.
Tolerance: Well tolerated; no adverse events.
Dropouts: 2; 1 control (non-compliance); 1 treatment
(unknown reason).
Other Endpoints:
-No change in fecal pH, serum IgE and IgG levels.
-Higher counts of bifidobacteria and lactobacilli.

000076

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Savino et
al. 2003

Study effect of a
partially hydrolyzed
formula with GOS/lcinulin, palmitic acid &
low levels of lactose
on formula-fed infants
with G.I. problems.

Treatment
Population a

n=604; mean 1.35

Test Substance and

Dose

Formula with 7.2 g

GOS/L & 0.8 g lcinulin/L

Treatment
Duration

2 weeks

Source: Not indicated

Observational study.

Savino et
al. 2006

Study effects of a
formula containing
GOS/lc-inulin in
infants with colic.
Prospective
randomized
controlled study

Safety Data

n = 96; < 16 wk

Formula with 7.2 g

GOS/L & 0.8 g lcinulin/L and 41%
palmitic acid
Source: GOS Friesland Foods
Domo; Orafti

2 weeks

Results of GOS/lc-inulin Ingestion

Growth:
-Data not reported/measured.
Tolerance:
-Fewer regurgitation problems
-Increase in daily number of stools
-Fewer daily colic episodes
Dropouts:
-None reported.
Other Endpoints:
-91% positive subjective rating from parents.
-95% positive subjective rating from pediatricians.
Growth:
-Data not measured/reported.
Tolerance:
-Reduction in crying episodes after 7 and 14 days.
Dropouts:
-None reported; exclusion from analysis occurred with
infants from both groups due to not meeting inclusion
criteria and loss to follow-up.
Other Endpoints:
-Not relevant.

000077

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Study Design &
Objective

Reference

Schmelzle
et al. 2003

Study the nutritional

efficacy and
bifidogenic
characteristics of a
formula containing
partially hydrolyzed
whey protein, high palmitic acid content,
starch and
GOS+FOS vs. a
standard formula.

Treatment
Population a

n = 49; < 2 wk
postpartum

Test Substance and

Dose

Two treatments:
Standard formula,
formula with 7.2 g
GOS/L & 0.8 g lcinulin/L

Treatment
Duration

10 weeks

Source:
Omneo/Comformil,
Numico

Randomized, doubleblind study

Scholtens
et al.
2006b

Study effects of solid

foods with GOS+FOS
on microflora in term
infants.
Randomized, double
blind, placebocontrolled
intervention trial

Safety Data

Intent-to-treat:
n = 19; 16 wk
Per protocol
analysis:
n = 11; 16 wk (data
presented in paper)

baby food with 4.05 g

GOS/d & 0.45 g lcinulin/d; divided into 3
equal servings
6 weeks
Sources:
GOS - Borculo
Domo;
FOS - Orafti

Results of GOS/lc-inulin Ingestion

Growth:
-At 6 wk, girls had larger weight gains and at 12 wk girls
had larger head circumference gains.
-Boys had larger total skinfold thickness gains.
-Weight and length gains were similar at 12 wk and
groups were in-line with published growth curves.
Tolerance:
-Softer stools.
-Formulas were well tolerated.
Dropouts:
-None reported.
Other Endpoints:
-Decreased energy intake from formula.
-Increased fecal bifidobacteria compared to baseline,
and higher % bifidobacteria vs control group at 6 wk.
-No effects on serum total protein, albumin or urea; no
clinically significant differences in prealbumin and amino
acid values.
Growth:
-Data not measured/reported.
Tolerance:
-Non-significant decrease in stool consistency and
increase in frequency compared to control for baseline
measurements of 12 infants from per-protocol group.
Dropouts:
-11 infants withdrawn from final evaluation due to noncompliance; 1 infant withdrawn due to age > study
protocol.
Other Endpoints:
-Increase in % fecal bifidobacteria; no effect on fecal
bifidobacteria count (no effect in intent-to-treat analysis).
-No effect on change in fecal pH, total SCFA, or %
acetate, propionate, or sum of valerate, isovalerate and
isobutyrate; decrease in % butyrate.

000078

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Reference

Scholtens
et al. 2008

Schouten
et al. 2011

Study Design &

Objective
To evaluate the effect
of prebiotics on
intestinal microbiota
composition and fecal
sIgA.
Randomized, doubleblind, placebocontrolled.
To measure the
plasma kappa and
lambda Ig-fLC
concentrations in
infants at risk of
developing allergic
disease.

Treatment
Population a

n = 215

Stam et al.
2011

Double-blind,
placebo-controlled,
randomized.

Safety Data

6 g/L scGOS/lc-inulin
(9:1 ratio)

Treatment
Duration

Results of GOS/lc-inulin Ingestion

First 26
wks of life.

Growth: Not reported.

Tolerance: similar percentage of adverse events
between groups (42 47%); descriptions of adverse
events not reported.
Dropouts: 28; no reasons provided.
Other Endpoints:
-Prebiotic group had higher percentage of bifidobacteria,
higher concentration of sIgA.

6 months

Growth: Data not measured/reported.

Tolerance: See Moro et al. (2006).
Dropouts: See Moro et al. (2006).
Other Endpoints:
-Plasma of infants suffering from atopic dermatitis had
higher total kappa and lamba Ig-fLC than infants without
atopic dermatitis.
-Infants receiving the scGOS/lc-inulin supplemented
formula had significantly lower Ig-fLC levels (p < 0.001).

1 year

Growth: No difference in growth (see Piemontese et al.

2011).
Tolerance: See Piemontese et al. 2011.
Dropouts: See Piemontese et al. 2011.
Other Endpoints:
-No difference in Haemophilus influenza type b antibody
levels.
-No difference in tetanus immunization response (IgG
levels).

Source: Numico

n = 74
(see Moro et al.
2006)

Hypoallergenic whey
formula with 8 g/L
scGOS/lc-inulin
(9:1 ratio)
Source: Numico

Prospective, doubleblind, randomized,

placebo controlled.
To evaluate the effect
of the prebiotics on Ig
responses to
vaccinations.

Test Substance and

Dose

n = 164;
(subset of Grber
et al. 2010 and
Piemontese et al.
2011)

6.8 g/L of scGOS/lcinulin (9:1 ratio) and

1.2 g/L pAOS in a
non-hydrolyzed cows
milk-based formula
Source: Nutricia

000079

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Reference

Vaisman et
al. 2010

Study Design &

Objective
To determine if
prebiotics modulate
inflammatory
processes of acute
diarrhea.
Double-blind,
placebo-controlled,
randomized,
prospective.

van der Aa
et al. 2010

To evaluate the effect

of synbiotics on the
severity of atopic
dermatitis in infants.
Double-blind,
placebo-controlled,
randomized.

van der Aa
et al. 2011

To investigate the
effect of synbiotics on
the prevalence of
asthma-like
symptoms in infants
with atopic dermatitis.
Double-blind,
placebo-controlled,
randomized.

Safety Data

Treatment
Population a

n = 104;
age = 9 24
months; suffering
from acute diarrhea

n = 90;
age < 7 mo; with
atopic dermatitis

n = 75;
mean age = 17
mos; with atopic
dermatitis

Test Substance and

Dose

6 g/day of 80%
GOS/lc-inulin (9:1
ratio), 20% pAOS

scGOS/lc-inulin (9:1
ratio) mixture in a
hydrolyzed wheybased formula that
included
Bifidobacterium breve
9
M-16V (1.3x10
CFU/100mL)
Source: Nutricia
scGOS/lc-inulin (9:1
ratio) mixture in a
hydrolyzed wheybased formula that
included
Bifidobacterium breve
9
M-16V (1.3x10
CFU/100mL)

Treatment
Duration

Results of GOS/lc-inulin Ingestion

12 days

Growth: Data not measured/reported.

Tolerance: Data not measured/reported.
Dropouts: 15; failed to receive intervention.
Other Endpoints:
-No difference in stool characteristics.
-No difference in cytokine levels, except TNF- (but not
clinically relevant).

12 weeks

Growth: Not evaluated.

Tolerance: Well tolerated no adverse events.
Dropouts: 8; none related to treatment.
Other Endpoints:
-No effect on severity of atopic dermatitis or serum IgE.
-Increased bifidobacteria, reduced pathogenic bacteria.
-Lowered fecal pH, increased fecal lactate, lowered
SCFAs.
-Fewer infants with dry stools, less constipation.

12 weeks

Growth: Not evaluated.

Tolerance: Well tolerated no adverse events.
Dropouts: 8; none related to treatment.
Other Endpoints:
-Lower prevalence of frequent wheezing & wheezing
and/or noisy breathing apart from colds.
-No difference in IgE levels.

Source: Nutricia

000080

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Reference

van der Aa
et al. 2012

Study Design &

Objective
To determine the
effects of synbiotics
on atopic markers,
cytokine production,
and circulating T cells
in infants with atopic
dermatitis.
Double-blind,
placebo-controlled,
randomized.
To measure the
modulation of
response to cows
milk protein and
vaccinations by
prebiotics.

van Hoffen
et al. 2009

van
Stuijvenber
g et al.
2011

Prospective, doubleblind, randomized,

placebo-controlled.
To assess the effect
of prebiotics on the
number of fever
episodes in infants.
Randomized, doubleblind, placebocontrolled.

Safety Data

Treatment
Population a

n = 90;
age < 7 mos; with
atopic dermatitis

Test Substance and

Dose
scGOS/lc-inulin (9:1
ratio) mixture in a
hydrolyzed wheybased formula that
included
Bifidobacterium breve
9
M-16V (1.3x10
CFU/100mL)

Treatment
Duration

12 weeks

Results of GOS/lc-inulin Ingestion

Growth: Not evaluated.

Tolerance: Well tolerated no adverse events.
Dropouts: 8; none related to treatment.
Other Endpoints:
-No difference in ex vivo cytokine production by
peripheral blood mononuclear cells.
-No difference in circulating regulatory T cell
percentages.

Source: Nutricia

n = 84; (subset of
Moro et al. 2006)

Hypoallergenic whey
formula with 8 g/L
scGOS/lc-inulin
(9:1 ratio)
Source: Numico

n = 830
(subset of Grber
et al. 2010 and
Piemontese et al.
2011)

6.8 g/L of scGOS/lcinulin (9:1 ratio) and

1.2 g/L pAOS in a
non-hydrolyzed cows
milk-based formula
Source: Nutricia

3 months
postvaccination
(6 months
of age)

Growth: Data not measured/reported.

Tolerance: See Moro et al. (2006).
Dropouts: See Moro et al. (2006).
Other Endpoints:
-Significant reduction in plasma IgE, IgG1, IgG2, IgG3
and cows milk protein-specific IgG1.
-No difference in IgG4 and diphtheria, tetanus, & poliospecific Ig.

1 year

Growth: No difference in growth (see Piemontese et al.

2011).
Tolerance: See Piemontese et al. 2011.
Dropouts: See Piemontese et al. 2011.
Other Endpoints:
-No difference in the number of fever episodes between
groups.

000081

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Table 5-4. Studies of 90% GOS/10% Long-Chain Inulin in Term Infant Formulas and Weaning Foods
Reference

VeeremanWauters et
al. 2011

Vivatvakin
et al. 2010

Study Design &

Objective
To assess safety,
tolerance, and
bifidogenic effects of
different doses of
prebioticsupplemented
formula
Prospective,
randomized, doubleblind, controlled.
To investigate the
effects of prebiotics
on gastrointestinal
comfort in infants.
Prospective,
randomized, doubleblind, placebocontrolled.

Treatment
Population a

n = 110

Test Substance and

Dose
Formula with 0.4 g/dL
of a 50:50 mixture of
lc-inulin and FOS;
formula with 0.8 g/dL
lc-inulin/FOS; or
formula plus 0.8 g/L
GOS/FOS (9:1 ratio).

Treatment
Duration

28 days

Source: Beneo

n = 144
(completed);
age = 30 days

Whey-based formula
containing LCPUFAs
and 4 g/L of a
GOS/FOS mixture
(9:1 ratio).

3 months

Results of GOS/lc-inulin Ingestion

Growth: No differences between groups.
Tolerance: No serious adverse events;
gastroesophageal reflux, regurgitation, hunger; similar
across groups.
Dropouts: 33; similar across groups; not attributed to
prebiotic use.
Other Endpoints:
-0.8 g/L supplementation had bifidobacteria counts
comparable to breast fed group.
-0.8 g/L supplementation had softer stools, consistency
comparable to breast fed group.
Growth: No difference in anthropometric
measurements.
Tolerance: No difference in adverse events between
groups.
Dropouts: 55; no difference across groups.
Other Endpoints:
-Fewer hard stools, more soft stools.
-No difference in bacterial composition, gastric emptying
time.

AOS = acidic oligosaccharides; cfu = colony forming units; d = day; f = female; FOS = fructooligosaccharides; g = grams; GOS = galacto-oligosaccharides; HDL = highdensity lipoprotein; HM = human milk; Ig-fLC = immunoglobulin free light-chain; lc-inulin = long-chain inulin; LDL = low-density lipoprotein; m = male; pAOS = pectin-derived
acidic oligosaccharides (identical ingredient to AOS); sc = short-chain; SCFA = short-chain fatty acids; SCORAD = SCORing Atopic Dermatitis; sIga = secretory
immunoglobulin A; wk = week.
a
Age at enrollment into the study.

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Safety Data

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5.7.3 Water Balance

In 2003, the EU Scientific Committee on Food assessed the safety of adding GOS/FOS to infant
formula in Report of the Scientific Committee on Food on the Revision of Essential
Requirements of Infant Formulae and Follow-on Formulae (SCF 2003). The evaluation noted
the potential adverse effects on infant water balance for hypernatremia due to a high dietary
renal solute load. The Committee found that based on the information available at the time the
addition of up to 0.8 g/100mL 9:1 GOS:FOS did not demonstrate adverse effects, particularly to
growth or markers of water balance. EFSA considered the issue and in a 2004 opinion based
on five studies available at the time did not find the information sufficient to eliminate the
possibility of a dehydration risk.
Notably, EFSA considered loose stools an adverse event. In many of the studies in sections
5.7 and 5.8, researchers reported that infants fed combinations of GOS and FOS had softer or
looser stools (Boehm et al. 2003; Veereman-Wauters et al. 2011; Westerbeek et al. 2010) but
these were not considered adverse events. Other studies noted less constipation in the
prebiotic-fed groups and constipation could be considered an adverse event (Mihatsch et al.
2006; Puccio et al. 2007; Savino et al. 2006; Van der Aa et al. 2010). Still other researchers
reported less or no diarrhea (an adverse event) in the prebiotic-fed group (Mihatsch et al. 2006;
Moro et al. 2002; Vivatvakin et al. 2010) while several studies found no difference in stool
frequency or consistency (Modi et al. 2010; Piemontese et al. 2011; Scholtens et al. 2006b).
Food Standards Australia New Zealand (FSANZ) addressed the EFSA opinion directly in the
following excerpt from the Final Assessment Report Proposal P306 Addition of Inulin/FOS &
GOS to Food:
In regard to the use of other ratios of oligosaccharides, or inulin-derived substances
alone in infant formula, FSANZ has considered a concern expressed by EFSA relating to
water balance. The available evidence that oligofructose and inulin are fermented by
colonic microflora in formula fed infants as described above (presence of SCFA in stool,
in vitro stool studies, and evidence from adult toleration studies), reduces the concern
that water balance could be adversely affected by an increase in osmotic potential due
to undigested inulin-derived substances in the colon. In addition, a 12-week study in
term infants which indicated oligofructose at 3 g/L had no significant effect on growth,
blood chemistry, or reported adverse events, supports the safety of inulin-derived
substances in infant formula products. It has been shown that 8 g/L of GOS:long-chain
inulin is safe, levels as high as 25 g HMOs [human milk oligosaccharides]/L in breast
milk is safe, and inulin-derived substances are likely to be fermented to a similar degree
to GOS and HMOs, therefore, FSANZ concludes up to 8 g/L added inulin derived
substances will be safe for young infants. This is supported by a recent 28-day study on
up to 8 g/L of a 1:1 oligofructose and long-chain inulin combination. This conclusion
applies equally to the use of GOS alone or any ratio of GOS:inulin-derived substances
so long as the total level of oligosaccharides is no greater than 8 g/L (FSANZ 2008,
p160).
These issues is addressed in greater detail in FSANZ (2008) and in GRN 392, but to summarize
here, studies and opinions published since the EFSA opinion in 2004 support the safety of the

Safety Data

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prebiotic-supplementation in infant formula and do not suggest a risk of hypernatremic

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oligosaccharides. Danone Research provided the scGOS/lc-inulin/AOS and maltodextrin

powders. The authors reported that the baseline nutritional and patient characteristics did not
differ between groups.
In Westerbeek et al. (2010), the authors objective was to determine the effect of the prebiotic
mixture on serious infectious morbidity, feeding tolerance, and short-term outcome in preterm
infants. The authors evaluated incidence of serious infectious morbidity, time to full enteral
feeding, age when parenteral feeding was discontinued, occurrence of necrotizing enterocolitis,
growth, need for mechanical ventilation, presence of periventricular-intraventricular hemorrhage,
patent ductus ateriosus treated with ibuprofen, indomethacin and/or surgical ligation,
bronchopulmonary dysplasia, retinopathy of prematurity, age at discharge from neonatal
intensive care, age at discharge from the hospital, and death. Westerbeek et al. (2010) found
no statistically significant difference between the prebiotic group and the placebo group in any of
these outcomes except one. The authors found that the group receiving the prebiotic mixture
had a lower incidence of bronchopulmonary dysplasia (0% vs. 23%, p = 0.003). Westerbeek et
al. (2010) concluded that the prebiotic supplementation by scGOS/lc-inulin/AOS did not
significantly reduce the risk of serious infectious morbidity in preterm infants though the authors
did observe a trend toward decreased serious infectious morbidity that should be investigated in
a larger study.
In Westerbeek et al. (2011a), the authors objective was to determine the effect of enteral
supplementation of the prebiotic mixture on fecal interleukin-8 (f-IL-8) and fecal calprotectin
(f-calprotectin) in preterm infants and to determine host- and treatment-related factors
associated with f-IL-8 and f-calprotectin in these preterm infants. The authors measured IL-8
and calprotectin in fecal samples taken from the diaper at 48 h after birth, postnatal days 7,
14 and 30. The authors observed that f-IL-8 and f-calprotectin were strong correlated at all time
points (p < 0.001) but they did not observe a difference between the prebiotic supplemented
group and the placebo group. Westerbeek et al. (2011a) concluded that the enteral
supplementation of the prebiotic mixture did not affect f-IL-8 and f-calprotectin.
In Westerbeek et al. (2011b), the authors objective was to determine the effect of the prebiotic
mixture on intestinal permeability in the first week of life in preterm infants and to determine
host-and treatment-related factors associated with intestinal permeability in these preterm
infants. The authors measured intestinal permeability by a sugar absorption test preformed at
approximately 36 15 h after birth and days 4 and 7 post-birth. Westerbeek et al. (2011b)
observed that while decreased intestinal permeability was correlated with increased body weight
(p = 0.002), the difference between the prebiotic supplemented group and the placebo group
was not significantly different. Westerbeek et al. (2011b) concluded that the enteral
supplementation of the prebiotic mixture did not enhance the postnatal decrease in intestinal
permeability in preterm infants in the first week of life.
In Westerbeek et al. (2011c), the authors objective was to determine the effect of the prebiotic
mixture on stool viscosity, stool frequency and stool pH in preterm infants. The authors
observed lower stool viscosity (p = 0.03) and lower stool pH (p = 0.009) at day 30 in the
prebiotics group but no significant difference in stool frequency.

Safety Data

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Results from these thirteen clinical trials provide additional evidence that the addition of a
GOS/lc-inulin mixture in a 9:1 ratio to infant formula for preterm infants is well tolerated over the
2 to 12 week study periods. These preterm infants were closely monitored throughout the
studies, and no adverse effects were observed on measures of growth, fluid balance, formula
intake, or stool characteristics. The absence of reports of adverse effects in this highly sensitive
and closely monitored study population provides strong evidence for the suitability of GOS and
lc-inulin in a 9:1 ratio in infant formulas.

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Safety Data

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Table 5-5. Studies of 90% GOS/10% Long-Chain Inulin in Preterm Infant Formula
Reference

Boehm et al.
2002 and
2003; Lidestri
et al. 2003;
Knol et al.
2005b

Indrio et al.
2009a/b

Study Objective &

Design

Investigate effects
of preterm formula
containing GOS/lcinulin on fecal flora
and stool
characteristics of
infants born at 32
wk gestational age.

Study effect of
GOS/lc-inulin added
to an enteral milk
formula on the
maturation of
gastrointestinal
motility and
permeability in
preterm infants.

Treatment
Population

n = 15;
age = 7.9 - 8.3
d

Test Substance
and Dose

Preterm formula
with 9 g GOS/L &
1 g lc-inulin/L

Treatment
Duration

Results of GOS/lc-inulin ingestion*

28 d

Growth:
-No effect on weight or length gains.
Tolerance:
-Softer stool consistency and increased stool frequency.
-No effect on incidence of crying, regurgitation, vomiting.
Dropouts:
-None reported.
Other Endpoints:
-Increase in fecal bifidobacteria; no effects on lactobacilli, E. Coli.,
Clostridium spp. Bacteroides, Enterobacter, Citrobacter, Proteus,
Klebsiella and Candida.
-No effects on plasma concentrations of calcium and phosphorus or
plasma activity of alkaline phosphatase; trend toward higher calcium
concentrations in urine (p=0.055).
-Decrease in fecal counts and % of total selected pathogens (sum of
Staphylococcus aureus, S. epidermidis, S. haemolyticus,
Pseudomonas aeruginosa, Enterobacter, Klebsiella, Proteus,
Streptococcus group B, Clostridium difficile, Bacillus subtilis,
Acinetobacter).

Source: Numico
(now Danone)

n = 49;
GA = ~34
weeks

Preterm formula
with 7.2 g GOS/L
& 0.8 g lc-inulin/L
30 days
Source: Numico

Growth: No difference in daily increase of body weight.

Tolerance: No adverse events reported.
Dropouts: None reported.
Other Endpoints:
-Faster gastric emptying time.

Double-blind,
randomized,
controlled, parallel.

Safety Data

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Table 5-5. Studies of 90% GOS/10% Long-Chain Inulin in Preterm Infant Formula
Reference

Mihatsch et al.
2006

Study Objective &

Design
Evaluate feeding
tolerance in preterm
infants given
formula containing
GOS/lc-inulin

Treatment
Population

Modi et al.
2010

Double-blind,
randomized,
controlled, parallel

Safety Data

Treatment
Duration

preterm formula
with 9 g GOS/L &
1 g lc-inulin/L
n = 10;
age = mean 42
d postpartum

Double-blind,
controlled
Study enteral
tolerance of
GOS/lc-inulin
formula in preterm
infants whose
mothers milk is in
shortage or
unavailable.

Test Substance
and Dose

n = 160;
age < 1 d

14 days
Source: Milupa
GmbH (fully
owned subsidiary
of Danone)
Nutriprem
formula with 7.2 g
GOS/L & 0.8 g lcinulin/L
~9 wk

Source: Numico

Results of GOS/lc-inulin ingestion*

Growth:
-No effects on daily formula intake or daily weight gains.
Tolerance:
-Decrease in stool viscosity, decreased change in gastrointestinal
transit time (decrease of 6 d vs. increase of 9 d in control group).
Dropouts:
-No adverse effects observed.
Other Endpoints:
-Lower fecal pH.
Growth: No difference between groups in mean weight gain,
increase in body length and head circumference.
Tolerance: Intake levels did not differ between study groups.
Dropouts: 10;
-6 infants withdrawn due to parental request (2 in control, 4 in
treatment group).
-5 infant deaths. 2 in control group, 3 in GOS/inulin group.
-For all adverse events, possible or presumable relationship to
treatment was excluded by clinician.
Other Endpoints:
-No difference in fecal flora, gastrointestinal signs or indices of fluid
balance, or in daily number of stools and stool characteristics.

000090

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Table 5-5. Studies of 90% GOS/10% Long-Chain Inulin in Preterm Infant Formula
Reference

Underwood et
al. 2009

Study Objective &

Design

To compare the
effect of prebiotics
& probiotics on
weight gain, stool, &
stool SCFAs in
premature infants.

Treatment
Population

n = 90

Randomized,
blinded, placebocontrolled trial

Westerbeek et
al. 2010;
2011a/b/c

To evaluate the
effect of prebiotics
on infections,
intestinal
inflammation,
intestinal
permeability, stool
viscosity, stool
frequency, stool pH.
Randomized,
double-blind,
controlled trial.

Test Substance
and Dose
Supplement
containing 2
lactobacillus
species plus FOS
(Culturelle); or a
supplement
containing
several species
of lactobacilli and
bifidobacteria
plus FOS
(ProBioPlus);
twice daily.

Treatment
Duration

Results of GOS/lc-inulin ingestion*

28 days or
discharge

Growth: No significant effect on growth.

Tolerance: No adverse reactions were noted.
Dropouts: None reported.
Other Endpoints:
-64% of infants receiving ProBioPlus became colonized with
bifidobacteria, compared to 18% of infants receiving Culturelle and
27% of infants receiving placebo.
-No differences were noted between groups in colonization rates for
lactobacilli, Gram-negative enteric bacteria or staphylococci.
-Bifidobacteria content in the stools of the infants receiving
ProBioPlus was higher than in the infants receiving Culturelle or
placebo.
-No significant differences in stool short chain fatty acid content were
detected between groups.

28 days

Growth: No significant effect on growth.

Tolerance: No adverse reactions were noted. No effect on feeding
tolerance.
Dropouts: None reported.
Other Endpoints:
-No reduction in risk of serious infectious morbidity, NEC, or other
complications.
-No effect on fecal interleukin-8 or fecal calprotectin.
-No effect on postnatal decrease in intestinal permeability.
-Decreased stool viscosity and decreased stool pH.
-No effect on stool frequency.

Source:
Culturelle ConAgra;
ProBioPlus UASLabs

n = 113;
GA < 32 weeks

enteral
supplementation
of 80%
scGOS/lc-inulin
and 20% AOS
(1.5 g/kg/d)
Source: Danone

AOS = acidic oligosaccharides; cfu = colony forming units; d = days; f = female; FOS = fructooligosaccharides; g = grams; GA = gestational age; GOS = galactooligosaccharides; HDL = high-density lipoprotein; HM = human milk; lc-inulin = long-chain inulin; LDL = low-density lipoprotein; m = male; NEC = necrotizing enterocolitis;
sc = short-chain; SCFA = short-chain fatty acids; wk = weeks; y = years.

Safety Data

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Prepared for Danone Trading B.V.

5.9 Use of Inulin and Oligofructose in Medical Foods

5.9.1 Potential Role of Fiber in Medical Foods
Medical foods are administered to many chronically ill individuals in order to improve nutritional
parameters (Cockram et al. 1998). Until recently, most enteral foods were intentionally
produced without added fiber, in part due to concerns about tube obstruction and in part to
provide rest for the intestinal tract (Lochs et al. 2006). However, individuals receiving fiber-free
medical foods were observed to have significantly prolonged gut transit times (Silk 2001) and
significant decreases in anaerobic bacteria (Schneider et al. 2000), leading to diarrhea,
constipation, and bloating. It is now recognized that consumption of fiber-containing enteral
formulae and tube feeds by patients improves stool consistency, gut function, and control of
glycemic and lipid parameters. The addition of fiber to enteral formulas therefore may have
important clinical benefits (Elia et al. 2007).
Soy polysaccharides were the first fiber source added to medical foods (Green 2001). Types of
fiber currently added to medical foods include soy polysaccharides, cellulose, resistant starch,
guar gum, acacia fiber, pectin, lignin, inulin, FOS, and other products. Tolerance to enteral
formulas containing soy polysaccharides has been established in a variety of patient
populations (Green 2001). However, data on tolerance to enteral formulas containing lc-inulin
or related 2-1 fructan products are more limited, and most of the available data on tolerance
have been obtained in studies testing a combination of fiber products (Table 5-6).

Prepared for Danone Trading B.V.

Table 5-6. Studies with FOS/Inulin/Oligofructose in Enteral Formulas

Reference

Treatment
Population
n = 20;
age = mean 76.2 y

Bunout et
al. 2002

Healthy, elderly
adults administered
influenza &
pneumococcal
vaccines

n = 26;
age = mean 46 y
Cockram
et al. 1998

Free-living adults
with end-stage
renal disease

n = 18;
Garleb et
al. 1996

Safety Data

Healthy, male
university students

Dose

Treatment
Duration

6 g/d
oligofructose & inulin
Source:
70% Raftilose, 30%
Raftiline, Orafti

28 wk

15.6 g/d (wk 1)

18.5 g/d (wk 2) FOS
Source:
reformulated Nepro;
Ross Products.
(DP=2-4)

2 wk

15 g/d or 31 g/d
FOS
Source: Golden
Technologies Co,
Inc.; Colorado.
(DP=2-4)

2 wk

Effects
Safety/Tolerance:
-No difference in episodes of infections, including gastrointestinal.
Dropouts:
-3 withdrawn from study due to adverse events; not treatment-related.
-9 subjects withdrawn for non-compliance; 5 in treatment group.
Microbiological:
-Not measured in this study.
Other Endpoints:
-No effect on serum proteins, immunoglobulins, C-reactive protein, secretory IgA,
antibodies, or IL-4 and interferon- secretion by cultured monocytes.
Safety/Tolerance:
-No changes in number or severity of GI symptoms including nausea, diarrhea,
flatulence, cramping, distention; no change in stool frequency or consistency.
-No change in urea kinetics; no difference in mean energy or protein intakes.
Dropouts:
-5 withdrawn due to GI symptoms; 1 from treatment group.
Microbiological:
-Not measured in this study.
Other Endpoints:
-Improved serum phosphorus and calcium phosphorus product at 15 and 22 d.
Safety/Tolerance:
-No change in body weights.
-No effect on serum chemistry profiles.
-No difference in complaints of nausea, cramping, distension, or vomiting.
-Comparable incidence of diarrhea; slightly more complaints in high-dose
treatment group of diarrhea, distension and flatus.
Dropouts:
-1 withdrawn from high-dose group after 1 d due to intolerance of the liquid
product; subject replaced with an alternate.
Microbiological:
-Increase in fecal bifidobacteria levels for both treatment groups.
-Increase in SCFA concentration by d 14.
-No difference in fecal pH.

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Table 5-6. Studies with FOS/Inulin/Oligofructose in Enteral Formulas

Reference

Treatment
Population

n = 40; >18 y
20 per treatment
group
Gori et al.
2011

Non-symptomatic,
antiretroviral-nave
HIV seropositive
adults

n = 27;

Guimber
et al. 2010

age = 11.9 3.9 y;

Long-term enteral
nutrition patients.

Sobotka et
al.
1997

Safety Data

n = 9;
age = mean 45 y
Adult patients
requiring enteral
nutrition.

Dose

Treatment
Duration

13.5 g GOS/d & 1.5

lc-inulin g/d
or
6.75 g GOS/d &
0.75 g lc-inulin/d
dissolved in water or
juice or mixed with
yogurt

12 wk

Sources: scGOS
Borculo Domo; lcinulin Orafti; AOS Sudzucker
Nutritionally
complete, multi-fiber
product containing
10.3% FOS & 22.2%
inulin

3 months
with 1
month runin, 1 month
washout

Source: Nutricia

30-37.5 g/d inulin

Source: Raffinerie,
Belgium

1 wk

Effects
Safety:
-Increase in overall G.I. symptoms in high-dose group at wk 4 and 12 compared
to low-dose group and control.; decrease in complaints observed at wk 12
compared to wk 4.
-Increase in abdominal distension and flatulence at wk 12.
-Complaints overall were mild and not clinically relevant.
-No unexpected clinical adverse events, or serious adverse events observed.
Dropouts: 10; 4 due to adverse events; 2 withdrew consent; 4 lost to follow-up.
Microbiological:
-Increased bifidobacteria.
-decreased Clostridium coccoides/Eubacterium rectale cluster.
-Decreased C. lituseburnense/C. histolyticum groups levels.
Other Effects:
-Reduction of soluble CD14.
+
+
-Activated CD4 /CD25 T cells.
-Increased natural killer cell activity.
Safety: Well tolerated; no changes in stool frequency or consistency, no change
in blood micronutrient status.
Dropouts: 7; not related to the study treatments.
Microbiological:
-Increase in stool bifidobacteria.
Other Effects:
-Reduction in stool pH.
Safety:
-Increase in flatulence.
-No change in stool frequency or consistency.
-No change in intestinal permeability.
Dropouts:
-No dropouts; all patients completed study.
Microbiological:
-No change in fecal SCFA concentration.

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Table 5-6. Studies with FOS/Inulin/Oligofructose in Enteral Formulas

Reference

Treatment
Population

Dose

Treatment
Duration

9.5 g/d FOS

n = 10; 21-34 y

soy polysaccharides

Whelan et
al. 2005

2 wk
Healthy adults
consuming enteral
formula

n = 11; mean 28 y
Whelan et
al. 2006

Safety Data

Healthy adults
consuming enteral
formula

Source: Nestle
(DP=2-4)

9.8 g/d FOS

Pea fiber
Source: Nestle
(DP=2-4)

2 wk

Effects
Safety/Tolerance:
-No difference in weight loss.
-Intake compliance 83% for treatment group; lower than for control.
-Increase in stool frequency; decrease in daily fecal weight.
-No difference in incidence or severity of gastrointestinal complaints.
Dropouts:
-2 subjects unable to consume enteral formula as sole source of nutrition; 1
subject dropped out for personal reasons unrelated to study; 1 subject not
included in analysis due to a positive Giardia lamblia test.
Microbiological:
-Increase in bifidobacteria compared with baseline and control levels.
-Decrease in clostridia compared to baseline.
-Increase in total SCFA, acetate and propionate concentrations; decrease in
butyrate.
Safety/Tolerance:
-No difference in macronutrient intakes.
-No difference in mean body weight.
Dropouts:
-2 participants dropped out due to dislike of formula; 1 dropped out for personal
reasons.
-No treatment-related dropouts.
Microbiological:
-Not measured in study.
Other Endpoints:
-Increase in mean fullness, minimum fullness and satiety.

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Table 5-6. Studies with FOS/Inulin/Oligofructose in Enteral Formulas

Reference

Treatment
Population

n = 32; mean 7.5 y

Zheng et
al. 2006

Pediatric cancer
patients consuming
enteral formula

Dose

Treatment
Duration

1.22 g/d
oligofructose & inulin
60 mg-kg-bw/d
Source: 70%
Raftilose, 30%
Raftiline

13-30 d

Effects
Safety/Tolerance:
-No effect on biochemical parameters (total bilirubin, alanine aminotransferase,
alkaline phosphatase, blood urea nitrogen, creatinine, glucose, etc.).
-Increase in hemoglobin and hematocrit; no effect on other hematological
parameters.
-Normal growth rates.
-1 report of rectal discomfort, 3 reports of mild flatulence, 1 report of mild
diarrhea; reports of flatulence included two occasions of abdominal pain.
-Twelve patients in control and 11 in treatment complained of nausea.
Dropouts:
-6 patients in the control group and 3 patients in treatment group voluntarily
discontinued enteral product between d 13 and 30 (Continuation was optional).
-1 patient developed diarrhea and was withdrawn from study at 3 d; nontreatment related.
Microbiological:
-No effect on bifidobacteria.
Other Endpoints:
-Normal immunological parameters: TNF-, IL-6, IL-2, LI(CD4/CD8).
-AGP levels different at 13 d of treatment; no difference in proportion of patients
with AGP increases above 1.4 g/L.
-Decrease in Prognostic Inflammatory and Nutritional Index at 30 d of treatment.

Abbreviations:
AOS = acidic oligosaccharides; cfu = colony forming units; d = day; DP = degree of polymerization; f = female; FOS = fructooligosaccharides; g = grams; GA =
gestational age; GI = gastrointestinal; GOS = galacto-oligosaccharides; HDL = high-density lipoprotein; HM = human milk; IL = interleukin; L = liter; lc-inulin = longchain inulin; LDL = low-density lipoprotein; m = male; NEC = necrotizing enterocolitis; sc = short-chain; SCFA = short-chain fatty acids; TNF = tumor necrosis factor;
wk = week; y = years.

Safety Data

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6 Summary and Conclusions

6.1

Summary and Conclusion on the Use of Long-Chain Inulin in Infant Formula

Numerous large, randomized, double-blind, placebo-controlled trials have been conducted to

assess the impact of formulas supplemented with a combination of GOS and lc-inulin on infant
growth, tolerance, and fecal microflora. Results from these studies consistently indicate that
formulas containing up to 8 g of the GOS/lc-inulin mixture support normal growth, are well
tolerated, and tend to produce fecal microflora and stool patterns more characteristic of infants
fed human milk as compared to infants fed standard formula. Results from studies of highly
sensitive and closely monitored preterm infants provide additional evidence that the GOS/lcinulin mixture is suitable for use in infant formulas.
None of the study populations in the clinical trials of GOS/lc-inulin supplementation included
infants with metabolic disorders. Given the rare occurrence of most inborn errors of
metabolism, it is difficult to conduct a clinical trial in this population. Trials in the general
population of infants, however, are appropriate to test the efficacy and safety of some novel
ingredients added to infant formula. Prebiotics such as GOS and lc-inulin are not digested and
exert their putative health effects primarily via fermentation in the colon. Gastrointestinal
function is not adversely affected in most infants requiring metabolic formula, therefore
tolerance of exempt metabolic infant formulas containing added lc-inulin and GOS ingredients
would be expected to be equivalent to tolerance of comparable doses in standard formulas.
A critical evaluation of the available evidence indicates that infant formulas containing up to 0.8
g lc-inulin, in combination with 7.2 g GOS, are well tolerated, safe, and suitable for term infants
from birth and that toddler formulas containing up to 1.2 g lc-inulin and 10.8 g GOS are well
tolerated and suitable for children 12 to 35 months of age.

6.2

Summary and Conclusion on the Use of Long-Chain Inulin in Medical Foods

The lc-inulin and related 2-1 fructans, in amounts of up to 37.5 g/day, are well-tolerated by
most individuals as demonstrated in studies conducted in a variety of populations, including
both healthy and chronically ill individuals. Initial mild discomfort tends to lessen with the
individuals adjustment to the 2-1 fructan intake. Some individuals, however, may not tolerate
higher levels of non-digestible carbohydrates. These medical foods will be administered under
medical supervision. The content of the GOS/lc-inulin mixture in the product will be clearly
labeled, which will enable health care professionals to administer the product in such a way that
ingestion of the product is well tolerated. Medical foods will be available in forms that do not
contain added lc-inulin and GOS, so it will be possible to customize fiber intakes as needed by
administering a combination of fiber-containing and fiber-free products.
A critical evaluation of the results of clinical studies of 2-1 fructan intake in populations
requiring medical foods demonstrates that the proposed typical uses of lc-inulin, in combination
with GOS are well tolerated, safe, and suitable for the tolerated by the target population. A
critical evaluation of the available evidence indicates that medical foods containing up to 2.5 g
lc-inulin per day, with a total GOS/lc-inulin intake of approximately 25 g per day, are well
tolerated, safe, and suitable for the general population under medical supervision.

Conclusions

000102

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

6.3

References

000111

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Prepared for Danone Trading B.V.

Safety Evaluation of
Long-Chain Inulin
for Use in
Term Infant Formulas,
Toddler Formulas,
and Medical Foods

Appendix A
Batch Data and Methods of Analysis for Orafti HP

Prepared for:
Danone Trading B.V.
Schiphol, The Netherlands
Prepared by:
ENVIRON International Corporation
Phoenix, Arizona
Date:
April 30, 2013
Project Number:
28-26968B

Appendix A

000116

Appendix A: Lc-Inulin Safety Evaluation

Prepared for Danone

Table A-1. Annual Analyses of Product: Orafti HP lc-inulin

Batches and Production Date
Parameter
Physical/Chemical
Inulin (% CHO)
Inulin DP 5 (% CHO)
Average DP
Glucose, fructose, sucrose (% CHO)
Sulfated ash (% at 525C)
Conductivity (S/cm at 28 Brix)
Contaminant
Lead (mg/kg)
Arsenic (mg/kg)
Microbiological
Total mesophilic bacteria (cfu/g)
Yeasts (cfu/g)
Molds (cfu/g)
Enterobacteriaceae (cfu/1 g)
Staphylococcus aureus (cfu/0.1 g)
Salmonella spp. (cfu/250 g)

28-26968B

HPBNN
6DNN6
10/27/06

HGQEO
7CEO7
04/19/07

HGQEQ
7CEQ7
04/21/07

HPHOT
7DOT7
11/26/07

HGQEC
8CEC8
04/08/08

HGQEE
8CEE8
04/10/08

HPQER
9CER9
04/22/09

HPQEX
9OEX9
04/27/09

HPBNK
9DNK9
10/24/09

HGBLT
0HLV0
09/15/10

99.8
99.3
30
0.2
0.18
182

99.9
99.9
31
0.1
0.05
54

99.9
99.8
32
0.1
0.07
77

99.8
99.4
27
0.2
0.14
242

99.9
99.8
33
0.1
0.13
133

99.9
99.8
34
0.1
0.11
137

99.7
99.2
34
0.3
0.05
84

99.8
99.4
35
0.2
0.10
150

99.9
99.6
30
0.1
0.07
136

99.7
99.3
27
0.3
0.12
156

<0.01
<0.03

0
0
0
negative
negative
negative

000117

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Safety Evaluation of
Long-Chain Inulin
for Use in
Term Infant Formulas,
Toddler Formulas,
and Medical Foods

Appendix B
GRAS Notifications for the Use of Inulin, Oligofructose and
Fructooligosaccharides as Food Ingredients

Prepared for:
Danone Trading B.V.
Schiphol, The Netherlands
Prepared by:
ENVIRON International Corporation
Phoenix, Arizona
Date:
April 30, 2013
Project Number:
28-26968B

Appendix B

000118

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

List of GRAS Notifications Contained on CD

GRN044 FOS (GTC Nutrition) FDA response letter 2000
GRN044 FOS (GTC Nutrition) FDA response letter 2007
GRN044 FOS (GTC Nutrition)
GRN118 Inulin (Sensus) FDA response letter 2003
GRN118 Inulin (Sensus) FDA response letter 2008
GRN118 Inulin (Sensus)
GRN236 GOS (Friesland Foods) FDA response letter 2008
GRN236 GOS (Friesland Foods)
GRN285 GOS (GTC Nutrition)
GRN286 GOS (GTC Nutrition) FDA response letter 2009
GRN286 GOS (GTC Nutrition)
GRN334 GOS (Yakult) FDA response letter 2010
GRN334 GOS (Yakult)
GRN392 GOS Beneo-Orafti FDA response letter 2012
GRN392 GOS Beneo-Orafti

Appendix B

000119

lc-Inulin Safety Evaluation

Prepared for Danone Trading B.V.

Safety Evaluation of
Long-Chain Inulin
for Use in
Term Infant Formulas,
Toddler Formulas,
and Medical Foods

Appendix C
References

Silk et al. 2001.
Sobotka et al. 1997.
Stam et al. 2011.
Stark and Lee 1982
Stone-Dorshow and Levitt 1987.
Ten Bruggencate et al. 2003.
Ten Bruggencate et al. 2004.
Ten Bruggencate et al. 2005.
Ten Bruggencate et al. 2006.
Tuohy et al. 2001
Underwood et al. 2009.
Vaisman et al. 2010.
van der Aa et al. 2010.
van der Aa et al. 2011.
van der Aa et al. 2012.
van Hoffen et al. 2009.
Van Loo et al. 1995.
Van Loo et al. 1999.
Van Loo 2004.
van Stuijvenberg et al. 2011.
Vandenplas et al. 2011.
Veereman 2007.
Veereman-Wauters et al.. 2011.
Vivatvakin et al. 2010.
Ward et al. 2006.
Watzl et al. 2005.
Weaver. 2005.
Westerbeek et al. 2010.
Westerbeek et al. 2011a.
Westerbeek et al. 2011b.
Westerbeek et al. 2011c.
Whelan et al. 2005.
Whelan et al. 2006.
Zheng et al. 2006.

Appendix C

000124

Pages 000125-001465 containing GRAS notices 44, 118, 236, 285, 286, 334, and 392
have been removed. They can be found in the "GRAS Notice Inventory" at
http://www.fda.gov/Food/IngredientsPackagingLabeling/GRAS/NoticeInventory/default.h
tm

Pages 001466-005452 have been removed in accordance with copyright

laws. Please see the bibliography list of the references that have
been removed from this request on pages 000104-000115.

SUBMISSION END

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Ucm370435 PDF

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GRAS Notice (GRN) No.

GRAS Notification for Long-chain Inulin (lc-inulin)

Dear Ms. Gaynor:

GRAS Exemption Claim for

A. Name and Address of Notifier

B. Common or Usual Name of GRAS Substance

Table 1: Intended Uses of GOS+Long-Chain Inulin (9:1) in Term Infant Formulas,

Healthy term infants beginning at

Healthy children from 12 through 35

GRAS Exemption Claim

Table 1: Intended Uses of GOS+Long-Chain Inulin (9:1) in Term Infant Formulas,

The products are intended to

12.5 g/1000 kcal

Nutritionally complete foods

The products are intended to

Tube feeds are intended for enteral

Enteral nutritional products

Expert Panel Report on the

Description of Long-Chain Inulin (lc-inulin), Manufacturing Process, Product

GRAS Status Consensus Statement

Proposed Uses and Estimated Daily Intakes

GRAS Status Consensus Statement

GRAS Status Consensus Statement

GRAS Status Consensus Statement

Term infant formulas and toddler

Non-exempt and exempt term

-Formulas for healthy term infants

-Formulas for healthy children from 12

Nutritionally incomplete oral

The products are intended to

Nutritionally complete foods

-The products are intended to

-Tube feeds are intended for enteral

Enteral nutritional products for

-Children who need sole source

The ratio of GOS:lc-inulin is 9:1.

GRAS Status Consensus Statement

2-Day Average lc-inulin Intakes Per User

(The remainder of this page intentionally left blank.)

GRAS Status Consensus Statement

Joseph F. Borzelleca, Ph.D.

David J.A. Jenkins, M.D., Ph.D., D.Sc.

lc-Inulin Safety Evaluation

Description of the Substance

History of Use and Intended Uses

lc-Inulin Safety Evaluation

Human Studies of Inulin and Oligofructose in Preterm Infant Formulas

Physical and Chemical Properties of Orafti HP LC-Inulin

Batch Data and Methods of Analysis for Orafti HP

GRAS Notifications for the Use of Inulin, Oligofructose and Fructooligosaccharides

lc-Inulin Safety Evaluation

lc-Inulin Safety Evaluation

2 Description of the Substance

Description of the Substance

lc-Inulin Safety Evaluation

Oligofructose-enriched inulin: Mixtures of oligofructose and lc-inulin.

2.1.2 Common or Trade Names

2.1.3 CAS Registry Number

2.1.4 Physical and Chemical Properties

2.1.5 Chemical Structure of Long-Chain Inulin

Description of the Substance

lc-Inulin Safety Evaluation

Figure 1. Chemical Structure of Long-Chain Inulin

2.2.2 Manufacturing Process

Description of the Substance

lc-Inulin Safety Evaluation

Figure 2. Production Process of Orafti HP lc-inulin

Description of the Substance

lc-Inulin Safety Evaluation

2.3 Product Specifications

Sulfated ash (% dry matter)

Description of the Substance

lc-Inulin Safety Evaluation

2.3.2 Product Stability

Description of the Substance

lc-Inulin Safety Evaluation