© 2018 JETIR October 2018, Volume 5, Issue 10 www.jetir.

org (ISSN-2349-5162)

Enantioseparation of (RS)-Zopiclone
using amino acid through thin layer
chromaography
Manisha Singh*
Department of Chemistry, Lovely Professional University,
Punjab, 144411, India.

Abstract
Resolution of enantiomers of (RS)-Zopiclone was achieved by using direct thin layer
chromatography on silica gel plates impregnated with optically pure (L-Tryptophan) as
chiral selector and as chiral mobile phase additive. The enantiomers were resolved by using
the solvent system (acetonitrile, methanol and water) with different ratio. Spots were
detected by using chamber.

Keywords: Enantioseparation, Thin layer chromatography, Zopiclone

1. Introduction
Almost a large fraction of active pharmaceutical ingrediants are enantiomers and the desired
biological activity is provided by only one of the two enantiomers. The undesirable
enantiomer results into side effects caused due to different therapeutic and pharmacological
activities of one of the enantiomers. With regard to concern at maintaining and enhancing
the global health, among the scientific community the US Food and drug administration
along with other regulatory bodies has created a general awareness. Consequently the
racemic drug dealing has led the regulatory bodies in many countries involved in the
registration process of all new active ingredients to pay a potential focus on the registration
of single enantiomer of a drug along with the stereo selectivity and stereo specificity of
drugs. Stereoselectivity has been encouraged and also well documented in nearly all the
pharmacokinetic processes such as absorbtion, distribution, excretion, etc[1-2]. The living
systems being chirally selective cause the two enantiomers of a racemic drug to have
different chemical reactivity, potency, efficacy, different pharmacokinetic and
pharmacodynamic interaction and also different toxicities. They interact differently towards
one same receptor and enzyme; hence resolution is an important process for mankind [3,4].
Since chiral environment is undoubtedly a primary requirement for chiral separation so the
chiral environment for the chromatographic resolution can be in a way provided through L-
amino acids. L- Amino acids being enantiomerically pure can be used as chiral selectors,
mobile phase additives and also as other chiral auxiliaries to prepare the chiral derivatizing
reagents[5-7]. The chosen drug, Zopiclone(Figure 1) 6-(5-chloro-2-pyridy1)-7-(4-methyl-
lpiperazinyl) carbonyloxy-6,7-dihydro [5~pyrroo [3,4-blpyrazin-5-on], which is a non-
benzodiazepine hypnotic drug belonging cyclopyrrolone class is commercialized as a
racemate[8,9]. It is an anti-depressant. Zopiclone is primarily used for the treatment of
insomnia and both the enantiomers follow a different rate of metabolism. (+)-Zopiclone or
Eszopiclone has 50-60 times greater receptor binding potency towards the benzodiazepine
than (-)-zopiclone [10]. In many countries like Europe and Japan zopiclone is
commercialized as a racemate. FDA has approved the enantiomer (+)–zopiclone having
higher pharmacological activity compared to the other enantiomer [11]. There have been a
few reports on the enantioresolution of Zopiclone using Hydroxy-Propyl-BetaCyclodextrin
as Chiral Selector[12], Chiral αl‐Acid Glycoprotein Column[13], -cyclodextrin bonded
phase [14]. For the enantioseparation of (RS)- Zopiclone, this is the first report following
this approach

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Figure 1: Structure of (RS)- Zopiclone

2. Experimental

2.1. Chemicals and instrumentation:-

UV visible spectrometry was performed by spectrophotometer SHIMADZU-UV-1800. IR
analysis was performed using FTIR by SHIMDZU. Pharmaceutical drug marketed as
zopiclone by ALKEM Laboratories ltd. (Mumbai, India) was purchased from a local drug
store containing 10mg active pharmaceutical ingredient zopiclone. Silica gel was purchased
from LOBA CHEMIE pvt ltd. (Mumbai, India). Amino acid (L-Tryptophan) marketed by
Lobachemie Pvt Ltd. (Mumbai, India) and other reagents marketed by Qualikems Fine
chem. Pvt Ltd. And ALPHA CHEMIKA (Mumbai India) were obtained from nearby drug
store.

2.2. Isolation and purification of (RS)- Zopiclone

30 tablets of racemic zopiclone were taken for purification and was purified by grinding it
to fine powder using pistil. The powder so obtained was dissolved into methanol (maximum
soluble) and the solution formed in the beaker was stirred properly to get the maximum
drug dissolved. Then the solution was sonicated for nearly 20 minutes on the sonicator
about 5 to 6 times. The filtration was done using 190µm thick Whatman filter paper having
pore size 8µm.The filtrate obtained after the completion of the sonication process was
evaporated in air and crystals of pure sample were achieved at room temperature . With the
help of IR and UV the sample was analyzed and the peaks of the drug were obtained
confirming us the purification of the sample. Also, melting point was taken to check out the
purity and was found out to be 129 0C.

2.3. Thin Layer Chromatography

Separation was carried out through direct method using amino acid L-tryptophan as chiral
reagents through two different approaches.
Approach A:- Amino acid was impregnated into the TLC plates and resolution of racemic
zopiclone was carried out using different achiral solvent systems. The TLC plates were
dried and activated for nearly 8 to 10 hours at an appropriate temperature of 50 to 60 0C.
Also, to get the desired results, different binary and ternary solvents of methanol, water and
acetonitrile were used in different ratios.
Approach B:- Chiral mobile phase was prepared and separation was performed by putting
an appropriate amount of chiral selector into the ternary solvent system. Spots of chiral drug
were put on the plain TLC plates and this way employed for resolution respectively.

3. Results and Discussion

In approach A, TLC plates were impregnated with L-Tryptophan. Slurry of silica gel was
prepared by taking 50ml of distilled water and a 10-2M pure amino acid solution was mixed
in it. Also, 25g of silica gel were dissolved in the chiral solution prepared and the slurry
formed was used to prepare the TLC plates. The TLC plates prepared were dried at room
temperature and activated at nearly 60 0C in an oven for 8 hours. After heating the TLC
plates, they were cooled to room temperature and used for separation. In approach B, the
chiral mobile phase was prepared initially by preparing a 10-2M amino acid solution in 95:5
(water : methanol) and added in appropriate ratio to the binary or ternary solvent system.
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Chromatograms were developed in iodine chamber for both approach A and B. Different
binary and ternary solvent systems using acetonitrile, methanol and water were used.
Resolution was not achieved using the binary solvent systems of acetonitrile and methanol.
Separation was achieved using a ternary solvent system of acetonitrile, methanol and water.
The different ratios tried for resolution were 4:3:3, 5:1:1, 5:4:5 etc. The successful ratios
of resolution are given in the Table 1. Spots were located in an iodine chamber.
Chromatograms achieved showing resolutions through this method are given below in
Figure 2. It was observed that via approach A resolution was found to be 4.21 and via
approach B it was 3.89. Approach A was found to good as compare to Approach B. Between
16 and 32 °C analyze with effective solvent systems, were done. For this purpose, those
chromatographic chambers were set inside an incubator to accomplish the particular
temperature. Those best determination to (RS)- Zopiclone for constantly on L- tryptophan
for both approach acquired during 28 °C. Expand about temperature should 32°C brought
about tailing from claiming spots Also adiminish clinched alongside temperature to 16°C
demonstrated no determination or eight molded structures.

(a) (b)

Figure 2: Pictures of chromatograms showing resolution using (a) approach

A, (b) approach B successful solvent system:-Acetonitrile:Methnaol:Water

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Table 1: For enantioseparation of (RS)-Zopiclone using L- Tryptophan the

RS and hRf (Rf ×100) values are shown below:

Approach
Solvent system hRf RS

Rf1 Rf2
A Acetonitrile-methanol-water(4.3:3 v/v) 32 64 4.21

B Acetonitrile-methanol-water(4.4:2 v/v) 14 93 3.89

Acknowledgments
Author is thankful to Lovely Professional University for providing lab for experimental
work.

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Authors
Author’s Name, Dr. Manisha Singh
Assistant Professor
Lovely Professional University
Punjab, India

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