Jurnal Rekristalisasi
Jurnal Rekristalisasi
Jurnal Rekristalisasi
Shailesh P. Zala*, Kena P. Patel, Khyati S. Patel, Jayshri P. Parmar and Dhrubo Jyoti Sen
Department of Pharmaceutical Chemistry, Shri Sarvajanik Pharmacy College,
Gujarat Technological University, Arvind Baug, Mehsana-384001, Gujarat, India
Abstract
Article History:------------------------
Covered in Index Copernicus with IC Value 4.68 for 2010
Shailesh P. Zala
E-mail: shailesh_zala@yahoo.com mobile-phase for checking purity of synthesized
compound, to do isolation properly and recrystallize
it with using suitable solvent. For all this purpose it is
Key words: necessary to remind polarity and solubility chart of
Mucoadhesion, microspheres, bioavailability. different polar and non-polar solvents. It is as
described as below.
How to Cite this Paper:
Shailesh P. Zala*, Kena P. Patel, Khyati S. Selection of Proper Solvent4-6
Patel, Jayshri P. Parmar and Dhrubo Jyoti The choice of solvent is perhaps the most critical step
Sen “Laboratory Techniques of Purification and in the process of recrystallization since the correct
Isolation”, Int. J. Drug Dev. & Res., April-June 2012, solvent must be selected to form a product of high
4(2): 41-55 purity and in good recovery or yield. Consequently a
solvent should satisfy certain criteria for use in
Copyright © 2012 IJDDR, Shailesh P. Zala et recrystallization.
al. This is an open access paper distributed under the The desired compound should be reasonably soluble
copyright agreement with Serials Publication, which in the hot solvent, about 5 g/100 mL (5 mg/100 µL)
permits unrestricted use, distribution, and being satisfactory and insoluble or nearly insoluble in
reproduction in any medium, provided the original the cold solvent. Note that the reference temperature
work is properly cited. for determination of the solubility in "cold" solvent is
often taken to be room temperature. This
combination of solute and solvent will allow Conversely, the impurities should either be insoluble
dissolution to occur in an amount of solvent that is in the solvent at all temperatures or must remain at
not unduly large and will also permit recovery of the least moderately soluble in the cold solvent. In other
purified product in high yield. A solvent having this words, if the impurities are soluble, the temperature
type of solubility properties as a function of coefficient for them must be unfavorable; otherwise
temperature would be said to have a favorable the desired product and the impurities would both
temperature coefficient for the desired solute. crystallize simultaneously from solution.
Covered in Index Copernicus with IC Value 4.68 for 2010
Sr. No. Solvent Formula Boiling point (0C) Melting point (0C) Density (g/mL) Solubility in H2O (g/100g) Relative polarity
1 Cyclohexane C6H12 80.7 6.6 0.779 0.005 0.006
2 Pentane C5H12 36.1 ˗129.7 0.626 0.0039 0.009
3 Hexane C6H14 69 ˗95 0.655 0.0014 0.009
4 Heptanes C7H16 98 ˗90.6 0.684 0.0003 0.012
5 Carbon tetrachloride CCl4 76.7 ˗22.4 1.594 0.08 0.052
6 Carbon disulfide CS2 46.3 ˗111.6 1.263 0.2 0.065
7 p-Xylene C8H10 138.3 13.3 0.861 0.02 0.074
8 Toluene C7H8 110.6 ˗93 0.867 0.05 0.099
9 Benzene C6H6 80.1 5.5 0.879 0.18 0.111
10 Ether C4H10O 34.6 ˗116.3 0.713 7.5 0.117
11 Methyl t-butyl ether C5H12O 55.2 ˗109 0.741 4.8 0.124
12 Diethylamine C4H11N 56.3 ˗48 0.706 M 0.145
13 Dioxane C4H8O2 101.1 11.8 1.033 M 0.164
14 N,N-Dimethylaniline C8H11N 194.2 2.4 0.956 0.14 0.179
15 Chlorobenzene C6H5Cl 132 ˗45.6 1.106 0.05 0.188
16 Anisole C7H8O 153.7 ˗37.5 0.996 0.10 0.198
Review Paper
The desired compound should be reasonably soluble very soluble in a cold, very polar solvent. In this case,
in the hot solvent, about 5 g/100 mL (5 mg/100 µL) a solvent of intermediate polarity may be the choice
being satisfactory and insoluble or nearly insoluble in for a satisfactory recrystallization.
the cold solvent. Note that the reference temperature Occasionally a mixture of solvents is required for
for determination of the solubility in "cold" solvent is satisfactory recrystallization of a solute. The mixture
often taken to be room temperature. This is usually comprised of only two solvents; one of
combination of solute and solvent will allow these dissolves the solute even when cold and the
dissolution to occur in an amount of solvent that is other one does not.
not unduly large and will also permit recovery of the
purified product in high yield. A solvent having this Solvent Pairs: Sometimes no single satisfactory
Review Paper
type of solubility properties as a function of solvent can be found, so mixed solvents, or solvent
temperature would be said to have a favorable pairs are used. To use a solvent pair, one dissolves
temperature coefficient for the desired solute. the crystals in the better solvent and adds the poorer
Conversely, the impurities should either be insoluble solvent to the hot solution until it becomes cloudy,
in the solvent at all temperatures or must remain at which means that the solution is saturated with the
least moderately soluble in the cold solvent. In other solute. The two solvents must be miscible with each
words, if the impurities are soluble, the temperature other.
coefficient for them must be unfavorable; otherwise First requirement for the reaction is to choose a
the desired product and the impurities would both solvent, which is not reacting with any starting
crystallize simultaneously from solution. materials and catalyst or which is act as a catalyst
The boiling point of the solvent should be low enough itself or which is act as a reactant. Sometimes there is
so that it can readily be removed from the crystals. no need of solvents and reactants are act as a solvent.
The boiling point of the solvent should generally be Like reaction of chlorination with thionyl chloride is
lower than the melting point of the solid is being used as a solvent as well as reactant for reaction. It
purified. The solvent should not react chemically also depends on the type of reaction like SN1 or SN2,
with the substance being purified. The chemical in both types of reactions different solvents are being
literature is a valuable source of information about used. Like in SN1 reaction ethanol, methanol is used
solvents suitable for recrystallizing known and in reaction of SN2 acetone, dimethyl formamide,
compounds. If the compound has not been prepared etc are being used. Some of the reactions contain
before, it is necessary to resort to trial-and-error property of hydrolysis means they are hydrolyses by
techniques to find an appropriate solvent for using water or moisture, like in reaction of
recrystallization. The process of selection can be chlorination, moisture free atmosphere is necessary.
aided by consideration of some generalizations about In order to obtain satisfactory results in many
solubility characteristics for classes of solutes. Polar syntheses involving air moisture sensitive reactions,
it may be necessary to purify solvents to remove sublimation at normal pressure. Several compounds
reactive impurities such as water. will sublime when heating under reduced pressure.
Protic/acidic materials, or atmospheric contaminants For Example- Sublimation of impure acetanilide,
such as oxygen. A commonly employed purification Iodine, Camphor
method is the solvent still, which involves the Place 50 mg of impure acetanilide (mixed acetanilide
reflux/distillation of an organic solvent in the with a minute amount of carbon black or other
presence of a dehydrating/deoxygenating reagent. substance) in a suction flask.
Covered in Index Copernicus with IC Value 4.68 for 2010
the mother liquor. Because the oil is not a pure cooler water for circulating in the cold finger. Turn
liquid, the solid mass produced from it will be on the heat and keep temperature stable at 135-
impure, as noted earlier. In a case such as this, the 140ºC. Crystals will form on the cold finger. Continue
usual remedy is to reheat the entire mixture to affect heating until sublimation is complete and no more
dissolution, add a few milliliters of additional pure crystals form on the cold finger. Turn off the heat.
solvent and allow the resulting solution to cool. Remove the apparatus from the heat and allow it to
cool at room temperature. Remove the cold finger
Sublimation9-10 from the suction flask gently. Scrape the crystals onto
Technique for purifying organic solid compounds a tare piece of weighing paper and reweigh. Record
with sublimation. Sublimation is a purification the mass of pure acetanilide. Determine its melting
vapor pressure and the impurities must have • Pour reaction mass to water, stir it and extract
significantly lower vapor pressures. By heating, the out product using organic solvent like Ethyl
solid will be vaporized and become solid again when acetate, Dichloromethane.
the vapor contacts with the cold surface. Some solid • It is not useful in case of pyridine, thionyl
compounds, such as iodine, camphor, naphthalene, chloride, DMF, methanol.
acetanilide, benzoic acid, can be purified by
Suction filtration
discarded. You then acidify and extract into a with buffer or solvent to remove further impurities.
new layer of organic solvent. Then, the analyte is eluted with a non-polar solvent
• For basic compounds you use an acid first then or a buffer of the appropriate pH.
base. The procedure can be repeated several
times although you lose quite a lot of product.
(a) (b)
the stationary phase, the analytes in the sample will automated technique that uses common solvents to
interact and retain on the sorbent but the solvent, rapidly extract solid and semisolid samples. ASE
salts and other impurities pass through the cartridge. operates at temperatures above the normal boiling
After the sample is loaded, the cartridge is washed point of most solvents, using pressure to keep the
solvents in liquid form during the extraction process.
Typically, ASE methods are completed in 15–25 min, compound is soluble in water. Alcohols and
while consuming only 15–50 mL of solvent. carboxylic acids, which also contain the -OH group,
are soluble in water, which contains the -OH group.
Purification techniques Benzene is not water-soluble but will dissolve other
• Recrystallization (if impurities are less than 5 %) hydrocarbons like hexane. Grease is soluble in
• Trituration (used for less than 500mgs) gasoline because both are hydrocarbons. It is
• Chromatography desirable to have a compound dissolve upon heating
Covered in Index Copernicus with IC Value 4.68 for 2010
• In the laboratory, most compounds are not are two techniques that may be used to attempt
method is the use of another piece of hot equipment. containing the solid. Stir each mixture and determine
A short-stemmed funnel is definitely better than a the solubility of the unknown in each solvent at room
long-stemmed one for filtration. temperature. Use the definitions of soluble, slightly
• When a recrystallization has been completed, soluble, or insoluble given earlier.
the crystals are collected on a Buchner (or Hurch) • If the unknown sample is insoluble in a
funnel. particular solvent, warm the test tube in the hot-
water. Stir or swirl the contents of the tube and note
Covered in Index Copernicus with IC Value 4.68 for 2010
Solvent Selection for recrystallization whether the unknown is soluble in hot solvent. If the
• Although different criteria are used for defining solid is soluble in the hot solvent but only slightly
solubility, plan to use the following definitions in this soluble or insoluble at room temperature, allow the
experiment: (a) soluble-20 mg of solute will dissolve hot solution to cool to room temperature slowly. If
in 0.5 mL of solvent; (b) slightly soluble-some but crystals form in the cool solution, compare their
not all of the 20 mg of solute will dissolve in 0.5 mL quantity, size, color, and form with the original solid
of solvent; (c) insoluble-none of the solute appears to material and with those obtained from other
dissolve in 0.5 mL of solvent. solvents.
• For known compounds, place about 20 mg (a • It is a good idea to test the solubility of a solute
spatula-tip full) of the finely crushed solid in a test in a variety of solvents. Even though nice crystals
Review Paper
tube and add about 0.5 mL of water using a may form in the first solvent you try, another one
calibrated Pasteur pipette. Stir the mixture with a might prove better if it provides either better
glass rod or spatula to determine whether the solid is recovery or higher-quality crystals. To assist in
soluble in water at room temperature. If the solid is determining the best solvent to use in recrystallizing
not completely soluble at room temperature, warm an unknown, you should construct a table containing
the test tube in the hot-water or steam bath and stir the solubility data you gather by the systematic
or swirl its contents to determine whether the solid is approach described above.
soluble in hot water. • If these solubility tests produce no clear choice
• Repeat the solubility test for the solutes using for the solvent, mixed solvents might be considered.
95% ethanol and then petroleum ether (b.p. 60-80̊C, Before trying any combinations of solvent pairs take
760 Torr). After completing these additional tests, about 0.2 mL of each pure solvent being considered
record which of the three solvents you consider best and mix them to ensure that they are miscible in one
suited for recrystallization of each of the solutes. another. If they are not, that particular combination
• For unknown compounds, a systematic cannot be used.
approach is important for determining their
solubility, and the following protocol accomplishes
this. The following solvents may be tried: water,
ethanol, diethyl ether and hexanes.
• After selecting the solvents, obtain enough
clean, dry test tubes so that there is one for each
solvent to be tested. Place about 20 mg (a spatula-tip
full) of the finely crushed unknown in each test tube
and add about 0.5 mL of a solvent to a tube Figure 8: Trituration
Trituration19
• Some compounds contain stickiness and non- Chromatography
polar impurities. • Chromatography is an effective and very useful
• To remove stickiness from the compound method for separation and purification of organic
trituration technique is being used. compounds. Chromatography separates components
• Generally non-polar solvents like n-hexane, n- of a mixture based upon the principle that how well
pentane, diethyl ether are used as a trituration they are adsorbed on the stationary phase, versus
Covered in Index Copernicus with IC Value 4.68 for 2010
solvent to remove non-polar impurities. how well they dissolve in the mobile phase. The
• Oils may persist on cooling with no evidence of components with greater affinity for the mobile
crystallization. These may often be induced to phase will move faster than those components with
crystallize by scratching the oil against the side of the greater affinity for the stationary phase, causing the
flask with a glass rod at the interface of the oil and components to separate. There are many
the solution. If this fails, several small seed crystals of chromatographic methods characterized by the
the original solid may be added to the oil, and the nature of the stationary and mobile phases. Among
mixture allowed to stand for a period of time. Failure these methods, column chromatography, thin-layer
of these alternatives may necessitate separation of chromatography and paper chromatography are
the oil from the solution and crystallization of it from more common ones.
Review Paper
another solvent.
Figure 9: Chromatography
Thin-layer chromatography20-23 • Obtain a TLC chamber and place solvent, a 5%
• Generally first of all confirm the purity of ethyl acetate in dichloromethane to 0.5 cm height.
compound by taking TLC. Place a piece of filter paper around the inside surface
• Prepare 1 TLC plate (4 × 7 cm dimension). of the container and extend into the solvent.
• Handle it only on the edges, as fingerprints • A glass jar with a lid or a beaker with a watch
contain UV-active materials. Using a pencil draw a glass or a cover of a Petri dish can be used as a TLC
very light line across the sheet (short dimension) chamber, but it should be large enough so that the
about 1 cm from one end. Then make 4 small light TLC plate can lean against one side.
marks at even intervals along the line for spotting the • Using clean capillary tubes carefully spot four
samples. Draw another light line about 1 cm from samples at four pencil marks.
another end of the plate for the solvent front.
sheet. Place a TLC plates at a time in a TLC chamber. • 2,4-DNPH for Ketone and Aldehydes
drops of ethanol and allow ethanol to drain to the top driven hybrid of medium and short column
of adsorbent. chromatography optimized for rapid separation."
• Fill up the column with ethanol as a polar This approach was pioneered by W.C. Still at
solvent. Columbia University. Flash chromatography utilizes
• When the first band comes down to the neck of a plastic column filled with some form of solid
the pipette column, collect it in a container and stop support, usually silica gel, with the sample to be
adding ethanol. separated placed on top of this support. The rest of
• Allow the solvent to drain to the level of the column is filled with an isocratic or gradient
alumina. Switch to the second eluting solvent, water solvent which, with the help of pressure, enables the
and fill up the column with water. Collect the second sample to run through the column and become
Review Paper
band into another container. separated. Flash chromatography used air pressure
initially, but today pumps are used to speed up the
separation. This technique is considered a low to
medium pressure technique and may be scaled up for
separations from a few mg to many tens or hundreds
of gams. grams.
Gas chromatography (GC)20-23 with and is retained by the polar stationary phase.
Gas chromatography is an analytical technique for Adsorption strengths increase with increased analyte
separating compounds based primarily on their polarity and the interaction between the polar
volatilities. Gas chromatography provides both analyte and the polar stationary phase (relative to the
qualitative and quantitative information for mobile phase) increases the elution time. The
individual compounds present in a sample. interaction strength depends not only on the
Compounds move through a GC column as gases functional groups in the analyte molecule, but also on
Covered in Index Copernicus with IC Value 4.68 for 2010
with their linear velocity and flow rates, because the steric factors. Reversed phase HPLC (RP-HPLC) has
compounds are normally gases or they can be heated a non-polar stationary phase and an aqueous,
and vaporized into a gaseous state. The compounds moderately polar mobile phase. One common
partition between a stationary phase, which can be stationary phase is silica which has been treated with
either solid or liquid and a mobile phase (gas). The RMe2SiCl, where R is a straight chain alkyl group
differential partitioning into the stationary phase such as C18H37 or C8H17. With these stationary phases,
allows the compounds to be separated in time and retention time is longer for molecules which are less
space. polar, while polar molecules elute more readily. The
retention time can be increased by adding more
High performance Liquid chromatography water to the mobile phase; thereby making the
Review Paper
stationary phase. During this movement the A pure supercritical fluid (SCF) is any compound at a
compounds with higher affinity to stationary phase temperature and pressure above the critical values
travel slowly while the others travel faster. Thus (above critical point). Above the critical temperature
separation of components in the mixture is achieved. of a compound the pure, gaseous component cannot
Once separation occurs individual components are be liquefied regardless of the pressure applied. The
visualized as spots at respective level of travel on the critical pressure is the vapor pressure of the gas at
plate. Their nature or characters are identified by the critical temperature. In the supercritical
Covered in Index Copernicus with IC Value 4.68 for 2010
means of suitable detection techniques. environment only one phase exists. The fluid, as it is
termed, is neither a gas nor a liquid and is best
Capillary Electrophoresis (CE)20-23 described as intermediate to the two extremes. This
Capillary Electrophoresis (CE) is a separation phase retains solvent power approximating liquids as
technique based on the differential transportation well as the transport properties common to gases.
velocities of charged species in an electric field
through a conductive medium. Primary candidate for Ultra Performance Liquid Chromatography
CE separation is ions (+/˗). The basic instrumental (UPLC)24-25
set-up consists of a high voltage power supply (0 to High performance liquid chromatography (HPLC) is
30 kV), a fused silica (SiO2) capillary, two buffer a proven technique that has been used in laboratories
Review Paper
reservoirs ,two electrodes and an column detector. worldwide over the past 30-plus years. One of the
primary drivers for the growth of this technique has
been the evolution of packing materials used to effect
the separation. The underlying principles of this
evolution are governed by the van Deemter equation,
which is an empirical formula that describes the
relationship between linear velocity (flow rate) and
plate height (HETP or column efficiency). Since
particle size is one of the variables, a van Deemter
curve can be used to investigate chromatographic
performance. According to the van Deemter
Figure 16: CE equation, as the particle size decreases to less than
2.5 µm, not only is there a significant gain in
efficiency, but the efficiency does not diminish at
increased flow rates or linear velocities. By using
smaller particles, speed and peak capacity (number
of peaks resolved per unit time in gradient
separations) can be extended to new limits, termed
Ultra Performance Liquid Chromatography, or
UPLC. The technology takes full advantage of
chromatographic principles to run separations using
columns packed with smaller particles and/or higher
Figure 17: SCF flow rates for increased speed, with superior
Supercritical fluid chromatography(SCF)20-23 resolution and sensitivity. They shows a stability
indicating assay of five related substances control the constant flow rate of the solvents. The
accomplished in under one minute, proving that the solvents are accessed through tubing from an outside
resolving power of UPLC is not compromised even at reservoir. The flow rate of the solvent is set through
high speed. The current USP lists multiple HPLC computer input and controlled by pumps. There are
methods for the analysis of these same compounds various columns used in liquid chromatography
with run times approaching 20 min, with broad, depending on the type of separation preferred. Each
tailed peaks. column contains a small diameter packing material.
Covered in Index Copernicus with IC Value 4.68 for 2010
Chromatography
separates into various zones of sample components.
These zones are referred to as bands.
Conclusion:
Nowadays it is necessary to have compounds with
maximum purity to get accurate result for analytical
datas (UV, IR, NMR, Mass spectras) & biological
activity both in-vivo and in-vitro. This review
provides depth of knowledge on importance of
Purification of New Drug Substance and New Drug
Product with various techniques of isolation and
purification of intermediate and final compounds
either obtained from natural, synthetic, semi-
synthetic or mineral sources.
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