Vol. 12(31), pp.

4891-4896, 31 July, 2013

DOI: 10.5897/AJB2013.12488
ISSN 1684-5315 ©2013 Academic Journals African Journal of Biotechnology
http://www.academicjournals.org/AJB

Full Length Research Paper

Comparison of antibacterial activity of parent plant of

Tylophora indica Merr. with its in vitro raised plant and
leaf callus
Noor Jahan1,2*, Razia Khatoon1,2, Anwar Shahzad3, Mohammad Shahid1,4 and Siraj Ahmad5
1
Department of Microbiology, Jawaharlal Nehru Medical College and Hospital, Aligarh Muslim University, Aligarh-
202002, India.
2
Department of Microbiology, Era’s Lucknow Medical College and Hospital, Lucknow-226003, India.
3
Plant Biotechnology Laboratory, Department of Botany, Aligarh Muslim University, Aligarh-202002, India.
4
Department of Medical Microbiology, Immunology and Infectious Diseases, College of Medicine and Medical Sciences,
Arabian Gulf University, Manama-26671, Kingdom of Bahrain.
5
Department of Community Medicine, Teerthanker Mahaveer Medical College and Research Centre, Teerthanker
Mahaveer University, Moradabad- 244001, India.
Accepted 25 July, 2013

The antibacterial potential of an endangered medicinal plant Tylophora indica was analyzed by agar
well diffusion method and its activity was compared with that of its in vitro raised plant and callus. The
extracts of parent plant of T. indica showed good antibacterial activity against gram negative bacteria
only; whereas, the extracts from in vitro raised plant and leaf callus showed good activity against both
gram positive and gram negative bacteria. Minimum Inhibitory Concentration (MIC) of the alcoholic leaf
extract of in vitro raised plant was determined by broth microdilution method. MIC against gram
positive bacteria ranged from 3.05 to 12.0 µg/ml and MIC against gram negative bacteria ranged from
1.53 to 24.0 µg/ml. The present study leads to conclusion that extracts of T. indica contains good
antibacterial activity which can be used in the treatment of various infections showing resistance to
treatment by currently used antimicrobial agents. As the in vitro raised plant and callus gave better
results as compared to parent plant, in vitro cultivation of explants may be used to obtain novel
antibacterial compounds. This is the first report on antibacterial activity of T. indica through in vitro
raised plant and its callus.

Key words: Tylophora indica, in vitro raised plant and callus, antibacterial activity.

INTRODUCTION

Infectious diseases account for high proportion of health 2012). This is due to indiscriminate use of commercial
problems and are the leading cause of death worldwide antimicrobial drugs commonly used for the treatment of
(Parekh and Chanda, 2007a). Bacterial and fungal patho- infectious diseases. The global emergence of multidrug
gens have evolved numerous defense mechanisms resistant bacterial strains is increasingly limiting the
against antimicrobial agents and their resistance to old effectiveness of current drugs and significantly causing
and newly produced drugs is on rise (Sangeetha et al., treatment failure of infections (Davies, 1994; Hancock,

*Corresponding author. E-mail: drnoorj@rediffmail.com.

4892 Afr. J. Biotechnol.

2005). This situation has forced the researchers to search 2008). Through in vitro cultivation, it would be possible to
for new antimicrobial substance from various sources preserve and conserve these endangered plants and
including medicinal plants (Scazzocchio et al., 2001; obtain phytotherapeutic compounds especially at places
Erdogrul, 2002; Bandow et al., 2003; Parekh and where the plant does not grow naturally due to adverse
Chanda, 2008). Plants are a goldmine of novel chemicals atmospheric conditions (Shahid et al., 2009b).
and an impressive number of modern drugs have been Although, T. indica is a versatile medicinal plant, with
developed from them (Reddy, 2010). There are several its use being restricted in localities of Indian sub con-
reports in literature regarding the antimicrobial activity of tinents and parts of Africa; the information on the antimic-
crude extracts prepared from plants (El-Seedi et al., robial activity of Tylophora species is insufficient. Hence,
2002; Rojas et al., 2003; Duraipandiyan et al., 2006; the present study was carried out to evaluate the anti-
Parekh and Chanda, 2007b). Antimicrobials of plant bacterial potential of medicinal plant T. indica Merr. and
origin have proved effective in the treatment of several compare its activity with its in vitro raised plant extract
infectious diseases while simultaneously mitigating many and callus.
of the side effects that are often associated with synthetic
antimicrobials (Samy and Ignacimuthu, 2000). There are
more than 2,70,000 higher plants existing in this planet. MATERIALS AND METHODS
But so far, less than 10% of recorded flora has been
Collection of plant materials
explored phytochemically as well as clinical evaluation for
various biological activities (Reddy, 2010). Fresh leaves were collected from six years old plant of T. indica
Tylophora indica Merr. (Asclepiadaceae) is a medicinal grown in the Botanical garden, Department of Botany, Aligarh
plant indigenous to India. It is also known as ‘Indian Muslim University, Aligarh.
ipecac’ in English, ‘Jangali pikvan’ in Hindi and ‘Anntmool’
and ‘Anthrapachaka’ in Sanskrit, is a dark copper coloured
In vitro culture of explants
delicate creeper found growing wild in the plains of India
and other sub-tropical regions of the world (Bhavan, In vitro shoot regeneration (for in vitro plant extract)
1992). The plant inhabits up to an elevation of 1,260 m in
the sub-Himalayan tract and in the central and peninsular The leaf explants were cultured on Murashige and Skoog’s (MS)
India (Nadkarni, 1976). It is also found in Ceylon, Malay medium (Murashige and Skoog, 1962) containing 5 µM of 6-
island and Borneo (Kirtikar and Basu, 1935). It is a Benzyladenine (BA). The cut ends of the explants started callusing
after 4 weeks of incubation. Shoot bud induction took place in 6
perrineal, small, slender, much branched pubescent twining weeks old culture. Shoot buds transformed into elongated shoots
or climbing herbs or under shrubs. Leaves are ovate- after second subculture passage in the fresh medium of same
oblong to elliptic-oblong. Flowers are minute, 1 to 1.5 cm composition. These microshoots (3 to 5 cm long) were transferred
across. Fruits are up to 7 × 1 cm, ovoid-lanceolate and to root induction medium containing MS + 2.5 µM of Indole 3-
tapering at apex. Flowers and fruits are produced bet- Butyric Acid (IBA). Healthy roots were induced within 2 weeks of
transfer. The rooted plantlets were acclimatized initially in culture
ween August to December (Kirtikar and Basu, 1935; room conditions by transferring in soilrite containing thermocole
Chopra et al., 1956a, 1956b). It is traditionally used as a cups. After one month, these were transferred to green house con-
folk medicine in certain regions of India for the treatment ditions. The plants thus obtained were then used further for
of bronchial asthma (Bielory and Lupoli, 1999), inflam- antimicrobial studies using their leaves.
mation, bronchitis, allergies, rheumatism and dermatitis
(Gupta and Bal, 1956; Dhananjayan et al., 1974; Gore et In vitro induction of leaf callus
al., 1980). The roots and leaves are also reported to be
used in hydrophobia. Dried leaves are emetic diaphoretic The leaf explants were cultivated in callus induction medium com-
prising of MS + 5 µM of 2,4-D (2,4-dichlorophenoxy acetic acid).
and expectorant. It is regarded as one of the best indi- Callusing was initiated from the cut ends of the explants after 25
genous substitute for ipecacuanha, so it was considered days of inoculation. Callus was yellow in colour and friable in
as Indian ipecacuanha in the latter half of the 19th nature. 4 g fresh weight of callus was induced after 5 weeks of
century (Kirtikar and Basu, 1935). culture which was used for evaluation of antimicrobial effect.
The leaves are employed to destroy vermin. The leaf
extracts also act as anti tumour agents (Chitnis et al.,
Plant extracts
1972; Stephen and Vijayammal, 2000). It has reputation
as an alternative blood purifier and has often been used The alcoholic extracts of the plant were tested for antimicrobial
in rheumatism. The roots and leaf powder are used in activity. The extracts were derived according to the method of
diarrhea, dysentery and intermittent fever. It was also Singh and Singh (2000) with some modifications (Shahid et al.,
identified as a good remedy in traditional medicine for 2007, 2009a, 2009b). To prepare alcoholic extracts, fresh leaves
psoriasis, anaphylaxis and leucopenia (Sangeetha et al., (15 g) from both sources (parent plant and in vitro raised plant)
were surface sterilized in 70% ethyl alcohol for 1 min and then
2012). But unfortunately, these plants are disappearing at washed 3 times with sterilized double distilled water (DDW). The
an alarming rate due to indiscriminate deforestation and leaf calli were aseptically removed from the culture tubes and all the
uncontrolled collection of plant materials (Vanila et al., plant materials, including calli, were grounded with sterile pestle
 Jahan et al. 4893

and mortar in 150 ml of absolute alcohol. The homogenized tissues The lowest concentration that did not show any visible growth
were centrifuged at 5000 rpm for 15 min, and the supernatant was was considered the MIC of that extract for the tested bacterial
filtered and taken as the alcoholic extract. The extracts were species. All the MIC experimentations were performed in duplicate.
immediately used for experimentation.
Statistical analysis
Microorganisms tested
All the experiments of antimicrobial susceptibility testing were
The clinical bacterial strains included in our study were Staphy- performed in triplicate. The results were expressed as the mean ±
lococcus aureus, Staphylococcus epidermidis, Streptococcus standard error (SE). Data were statistically analyzed by one way
pyogenes, Enterococcus faecalis, Bacillus subtilis, Escherichia coli, analysis of variance (ANOVA) followed by Tukey’s multiple analysis
Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus test using SPSS Software, Chicago, III, version 10. P values were
vulgaris, Salmonella typhi, Salmonella typhimurium, Shigella calculated by one-sample T-test and P < 0.05 was taken as statis-
dysenteriae type 1 and Vibrio cholerae isolated from clinical tically significant.
specimens in the Department of Microbiology, Jawaharlal Nehru
Medical College and Hospital, Aligarh Muslim University, Aligarh,
India. The control strains tested were S. aureus (ATCC 25923), E. RESULTS AND DISCUSSION
coli (ATCC 25922) and P. aeruginosa (ATCC 27853) obtained from
National Institute for Communicable Diseases (NICD), New Delhi, Antimicrobial activity of alcoholic extracts of leaves of
India. These strains were grown on Blood agar or MacConkey agar parent plant as well as its in vitro raised plant and leaf
plates at 37°C and maintained on nutrient and blood agar slants.
callus against the tested bacterial species is shown in
Tables 1 and 2. Negative control (ethanol) showed the
Antibacterial susceptibility testing zone of inhibition in the range of 7.33 ± 0.33 to 8.67 ±
Antibacterial activity was determined using agar well diffusion
0.33 mm. Positive control (gentamicin) showed the zone
method (Vanden-Berghe and Vlietinck, 1991; Akinpelu, 2001), with of inhibition in the range of 9.67 ± 0.33 to 13.00 ± 0.58
some modifications (Shahid et al., 2007). Antibacterial tests were mm. All the extracts showed good antibacterial activity.
performed as per Clinical and Laboratory Standards Institute, for- The alcoholic leaf extract of parent plant showed good
merly National Committee for Clinical Laboratory Standards (2000) activity against tested gram negative bacteria only and no
using Mueller-Hinton Agar (M 173; HiMedia, India). For fastidious activity against tested gram positive bacteria (Tables 1
organisms such as Streptococci, 5% sheep blood agar was used.
An inoculum containing 106 cfu/ml of bacteria was used for and 2). It showed significant (P<0.05) activity against E.
inoculating the susceptibility plates. The plates were lawn cultured coli (P = 0.024), K. pneumoniae (P = 0.020) and P.
with the bacterial suspensions with the help of sterile swabs and aeruginosa (P = 0.038). Various studies have been
wells of 5 mm diameter were made in each plate using a sterile undertaken previously by different researchers to analyze
borer. Plant extracts (20 µl) were poured in the wells using micro- the antibacterial potential of parent plant of T. indica. A
pipette. 20 µl of 95% ethanol was used to serve as negative control,
whereas, antibacterial agent gentamicin (500 µg/20 µl) was used as
study done by Parekh and Chanda (2008) also showed
positive control. The plates were kept upright for 5 to 10 min until no activity of alcoholic leaf extract of parent plant of T.
the solution diffused into the medium and then incubated aero- indica against S. aureus and S. epidermidis, which sup-
bically at 37°C for 24 h. Later, the zone of inhibition was measured ports our present research findings. On the other hand,
and recorded. All experiments were performed in triplicate. study done by Reddy (2010) showed significant activity of
this plant against S. aureus, K. pneumoniae, E. coli, S.
typhi, P. aeruginosa and P. vulgaris. Another study done
Determination of minimum inhibitory concentrations (MIC)
by Sangeetha et al. (2012) showed significant activity of
MIC was determined by broth micro-dilution method, performed leaf extract of parent plant of T. indica against S. aureus
according to Clinical and Laboratory Standards Institute (formerly only and no activity against E. coli and P. aeruginosa.
National Committee for Clinical Laboratory Standards, 2000), with These findings are in contrast with our study. This could
minor modifications (Shahid et al., 2007). Doubling dilutions of the be due to different concentrations of extracts used in their
extracts were prepared using RPMI-1640 broth (HiMedia, India)
supplemented with 0.3 g/L L-glutamine (HiMedia, India), 0.165
study as well as variation in active metabolites present in
mol/L of 3-[N-morpholino] propanesulfonic acid (MOPS) buffer plant extracts derived from different places.
(HiMedia, India) and 0.01% of dimethyl sulphoxide (DMSO) The alcoholic leaf extract of in vitro raised plant of T.
(Qualigens Fine Chemicals, India). The extracts were dissolved in indica showed good antibacterial activity against most of
DMSO, and further diluted 1:50 in RPMI-1640 medium, and each the tested gram positive bacteria, except S. pyogenes
resulting solution was used for a doubling dilution series. Microtitre and E. faecalis (Table 1). It showed significant (P<0.05)
plates were prepared containing 100 µl of undiluted extracts in the
first well, followed by doubling dilutions of extracts from second activity against S. aureus (P = 0.005), S. epidermidis (P =
well. The standardized inoculum of each bacterial species was 0.003) and B. subtilis (P = 0.044), with its MIC ranging
added to the respective dilution wells including the first well. The from 3.05 to 12.0 µg/ml (Figure 1). The alcoholic leaf
final concentrations of the extracts ranged from 25 × 103 to 48 × 10- extract of in vitro raised plant showed good activity
3
µg/ml. For each test, there was a sterility control well containing against most of the tested gram negative bacteria (Table
alcoholic extract in RPMI-1640 broth plus DMSO and a growth
control well containing bacterial suspension without alcoholic
2). It showed significant activity (P<0.05) against E. coli
extract. The microtitre plates were incubated at 35 ± 2°C for 24 h (P = 0.003), K. pneumoniae (P = 0.012), P. aeruginosa (P
with their upper surface covered by sterile sealers. = 0.010), S. dysenteriae type 1 (P = 0.012) and S. typhi
4894 Afr. J. Biotechnol.

Table 1. Antibacterial activity of alcoholic extracts of parent plant of Tylophora indica and its in vitro raised plant and leaf callus against
pathogenic gram-positive bacteria.

Zone of inhibition (mm) ± SE

Bacteria tested Alcoholic leaf Alcoholic leaf Alcoholic Ethanol† Gentamicin£
extract of extract of in vitro extract of leaf (negative (positive
parent plant∆ raised plant∆ callus∆ control) control)
Staphylococcus aureus 0.00±0.00a 14.67±0.33ab 13.00±0.58b 7.33±0.33d 12.67±0.33b
Staphylococcus epidermidis 0.00±0.00a 15.33±0.67a 12.67±0.33c 8.33±0.33b 12.33±0.33c
0.00±0.00a d d
Streptococcus pyogenes 0.00±0.00 0.00±0.00 8.67±0.33a 13.00±0.58a
0.00±0.00a d d
Enterococcus faecalis 0.00±0.00 0.00±0.00 7.67±0.33c 9.67±0.33d
0.00±0.00a c d
Bacillus subtilis 13.00±0.58 0.00±0.00 7.33±0.33d 12.33±0.33c
0.00±0.00a b a
S. aureus ATCC 25923 14.33±0.33 13.67±0.33 8.33±0.33b 13.00±0.58a
† = 20 µl of 95% ethanol used as negative control; ∆ = concentration of extracts used in the test that is, 2 mg / 20 µl; £ = concentration of
gentamicin used in test that is, 500 µg / 20 µl. Diameter of zone of inhibition is a mean of triplicates ± SE (mm). Differences were assessed
statistically using one way ANOVA followed by Tukey’s test. P<0.05 was considered as significant. The mean represented by same letter is not
significantly different within the column.

Table 2. Antibacterial activity of alcoholic extracts of parent plant of Tylophora indica and its in vitro raised plant and leaf callus
against pathogenic gram-negative bacteria.

Zone of inhibition (mm) ± SE

Bacteria tested Alcoholic leaf Alcoholic leaf Alcoholic Ethanol† Gentamicin£
extract of extract of in vitro extract of leaf (negative (positive
parent plant∆ raised plant∆ callus∆ control) control)
Escherichia coli 12.67±0.33b 14.33±0.33ab 11.33±0.33c 7.67±0.33c 11.67±0.33b
Klebsiella pneumoniae 11.67±0.33c 13.00±0.58b 11.67±0.33b 7.33±0.33d 10.67±0.33d
0.00±0.00e e f
Proteus vulgaris 0.00±0.00 0.00±0.00 8.33±0.33b 11.67±0.33b
Pseudomonas aeruginosa 11.33±0.33d 12.33±0.33c 11.33±0.33c 7.67±0.33c 10.67±0.33d
0.00±0.00e d d
Salmonella typhi 11.33±0.33 10.67±0.33 7.33±0.33d 10.33±0.33e
0.00±0.00e e f
Salmonella typhimurium 0.00±0.00 0.00±0.00 7.67±0.33c 10.67±0.33d
0.00±0.00e cd e
Shigella dysenteriae type 1 11.67±0.33 10.33±0.33 8.67±0.33a 9.67±0.33f
0.00±0.00e e f
Vibrio cholerae 0.00±0.00 0.00±0.00 7.33±0.33d 10.33±0.33e
13.00±0.58a a a
E. coli ATCC 25922 14.67±0.33 12.33±0.33 8.33±0.33b 12.67±0.33a
11.67±0.33c bc b
P. aeruginosa ATCC 27853 12.67±0.33 11.67±0.33 7.67±0.33c 11.33±0.33c
† = 20 µl of 95% ethanol used as negative control; ∆ = concentration of extracts used in the test that is, 2 mg / 20 µl; £ = concentration of
gentamicin used in test that is, 500 µg / 20 µl. Diameter of zone of inhibition is a mean of triplicates ± SE (mm). Differences were assessed
statistically using one way ANOVA followed by Tukey’s test. P<0.05 was considered as significant. The mean represented by same letter is
not significantly different within the column.

(P = 0.038), with its MIC ranging from 1.53 to 24.0 µg/ml this plant; therefore, our findings could not be compared.
(Figure 2). It is interesting to note that these bacteria
which are found to be susceptible to the extracts of in Conclusion
vitro raised plant of T. indica are important human
pathogens responsible for wound infection, enteric fever, The alcoholic extract of leaves from in vitro raised plant of
dysentery, urinary tract infection, pneumonia and diarrhea. T. indica and its in vitro cultivated leaf callus showed
The in vitro cultivated leaf callus also showed good anti- better antibacterial activity as compared to parent plant
bacterial activity which was comparable to the activity extract. These extracts showed wide range of antibacte-
shown by in vitro raised plant (Tables 1 and 2). Its alco- rial activity against various gram negative bacteria as well
holic extract showed significant activity (P < 0.05) against as against gram positive bacteria like S. aureus and S.
S. aureus and S. epidermidis (Table 1) and most of the epidermidis; hence, they could be used in the treatment
tested gram negative bacteria (Table 2). To the best of of infectious diseases caused by these organisms, which
our knowledge, this is the first study analyzing the anti- otherwise pose problem of resistance to the commonly
bacterial potential of in vitro raised plant and leaf callus of used antimicrobial agents. Thus, it leads to a conclusion
 Jahan et al. 4895

Figure 1. MIC determination of alcoholic leaf extract of in vitro raised plant of Tylophora indica
against tested gram positive bacteria.

Figure 2. MIC determination of alcoholic leaf extract of in vitro raised plant of Tylophora indica against
tested gram negative bacteria.

that high antibacterial activity of in vitro raised plant and prospect of these extracts which can be used as novel
callus may be due to enhancement of the bioactive antibacterial agents. Also, in vitro callus induction may be
compounds responsible for antibacterial effects by used to obtain phytotherapeutic compounds, especially at
nutritional and hormonal manipulations in the cultivation places where this plant does not grow naturally because
medium as depicted in our study. This shows the future of adverse atmospheric conditions.
4896 Afr. J. Biotechnol.

REFERENCES Reddy BU (2010). Enumeration of antibacterial activity of few medicinal

plants by bioassay method. E. J. Chem. 7: 1449-1453.
Akinpelu DA (2001). Antimicrobial activity of Anacardium occidentale Rojas R, Bustamante B, Bauer J, Fernandez I, Alban J, Lock O (2003).
bark. Fitoterapia. 72: 286-287. Antimicrobial activity of selected Peruvian medicinal plants. J.
Bandow JE, Brotz H, Leichert LIO, Labischinski H, Hecker M (2003). Ethnopharmacol . 88: 199-204.
Proteomic approach to understanding antibiotic action. Antimicrob. Samy RP, Ignacimuthu S (2000). Antibacterial activity of some folklore
Agents. Chemother. 47:948-955. medicinal plants used by tribals in western ghats of India. J.
Bielory L, Lupoli K (1999). Herbal interventions in asthma and allergy. J. Ethnopharmacol. 69: 63-71.
Asthma, 36: 1-65.Bhavan BV (1992). Selected Medicinal Plants of Sangeetha S, Shankar MEU, Mythili S, Sathiavelu A (2012).
India. Bombay, India: Tata Press. p. 333-6. Antimicrobial activity of Cassia auriculata and Tylophora indica. Inter.
Chitnis MP, Khandalekar DD, Adwamkar MK, Sahasrabudhe MB J. Univ. Pharm. Life. Sci. 2: 8-15.
(1972). Anti-tumor activity of the extracts of stem and leaf of Scazzocchio F, Comets MF, Tomassini L, Palmery M (2001).
Tylophora indica. Ind. J. Med. Res. 60: 359-362. Antibacterial activity of Hydrastis Canadensis extract and it’s major
Chopra RN, Nayar SL, Chopra IC (1956a). Glossary of Indian Medicinal isolated alkaloids. Planta. Med. 67: 561-563.
Plants,. New Delhi: CSIR Publication. 1: 32. Shahid M, Shahzad A, Malik A, Anis M (2007). Antibacterial activity of
Chopra RN, Nayara SL, Chopra IC (1956b). Glossary of Indian aerial parts as well as in vitro raised calli of the medicinal plant
medicinal plants; Council of Scientific and Industrial Research: New Saraca asoca (Roxb.) de Wilde. Can. J. Microbiol. 53: 1-7.
Delhi. p. 168-169. Shahid M, Rahim T, Shahzad A et al. (2009a). Ethnobotanical studies
Davies J (1994). Inactivation of antibiotic and the dissemination of on Berberis aristata DC. root extracts. Afr. J. Biotech. 8: 556-563.
resistance genes. Sci. 264: 375-382. Shahid M, Shahzad A, Anis M (2009b). Antibacterial potential of the
Dhananjayan R, Gopalakrishnan C, Kameswaran L (1974). extracts derived from leaves and in vitro raised calli of medicinal
Pharmacological action of Tylophora indica. Ind. J. Pharm. 36: 167. plants Pterocarpus marsupium Roxb., Clitoria ternatea L., and
Duraipandiyan V, Ayyanar M, Ignacimuthu S (2006). Antimicrobial Sanseveiria cylindrical Bojer ex Hook. Orien. Pharm. Exp. Med. 9:
activity of some ethnomedicinal plants used by Paliyar tribe from 174-181.
Tamil Nadu, India. BMC Comp. Alt. Med. 6: 35-41. Singh I, Singh VP (2000). Antifungal properties of aqueous and organic
El-Seedi HR, Ohara T, Sata N, Nishiyama S (2002). Antimicrobial extracts of seed plants against Aspergillus flavus and A. niger.
terpenoids from Eupatorium glutinosum (Asteraceae). J. Phytomorphology. 50: 151-157.
Ethnopharmacol . 81: 293-296. Stephen J, Vijayammal PL (2000). Antitumor activity of Tylophora
Erdogrul OT (2002). Antibacterial activities of some plant extracts used asthmatica. Anc. Sci. Life. 20: 88-91.
in folk medicine. Pharma. Biol. 40: 269-273. Vanden-Berghe DA, Vlietinck AJ (1991). Screening methods for
Gore KV, Rao K, Guruswamy MN (1980). Physiological studies with antibacterial and antiviral agents from higher plants. In Methods in
Tylophora asthmatica in bronchial asthma. Ind. J. Med. Res. 71: 144- plant biochemistry. Edited by PM Dey, JB Harborne, and
148. Kohostettmann), Academic Press, London. p. 47-70.
Gupta B, Bal SN (1956). Pharmacognostic studies of Tylophora indica Vanila D, Ghanthikumar S, Manickam VS (2008). Ethnomedicinal Uses
(Burm.f.) Merr. J. Sci. Industrial. Res. 15: 111. of Plants in the Plains Area of the Tirunelveli-District, Tamilnadu,
Hancock EW (2005). Mechanisms of action of newer antibiotics for India. Ethnobot. Leaf. 12: 1198-1205.
Gram-Positive pathogens. Lancet. Infect. Dis. 5:209-218.
Kirtikar KR, Basu BD (1935). Indian Medicinal Plants, Second edition
(Published by Lalit Mohan Basu, Allahabad, India). Vol. 2, p. 1492.
Murashige T, Skoog F (1962). A revised medium for rapid growth and
bioassays with tobacco tissue cultures. Physiol Plant. 15: 473-497.
Nadkarni K (1976). Indian Materia Medica, Vol 1. Bombay, India:
Popular Prakashan. p. 1252.
National Committee for Clinical Laboratory Standards (2000). Methods
for dilution antimicrobial susceptibility tests for bacteria that grow
th
aerobically. Approved Standard 5 ed. M7-A5. NCCLS, Villanova,
PA, USA.
Parekh J, Chanda S (2007a). In vitro screening of antibacterial activity
of aqueous and alcoholic extracts of various Indian plant species
against selected pathogens from Enterobacteriaceae. Afri. J.
Microbiol. Res. 1: 92-99.
Parekh J, Chanda S (2007b). In vitro antibacterial activity of ethanol
extract of Woodfordia fruticosa kurz. Flower (Lythraceae).Braz. J.
Micro.38: 204-207.
Parekh J, Chanda S (2008). Antibacterial Activity of Aqueous and
Alcoholic Extracts of 34 Indian Medicinal Plants against Some
Staphylococcus Species. Turk J Biol. 32: 63-71.