Separation of 1,1'-bi-2-naphthol enantiomers by continuous ...
Evaluation of the enantiomers of centchro- man and their ...
Transcript of Evaluation of the enantiomers of centchro- man and their ...
Indian Journal of ChemistryVol. 36B, April 1997, pp. 372-373
Evaluation of the enantiomers of centchro-man and their quaternary methyl iodide
salts for purity by chiral HPLCt
Amita Tandon, Neeta Srivastava & Suprabhat Ray*Medicinal Chemistry Division, Central Drug Research Institute,
Lucknow 226 001, India
Received 16 July 1996; accepted 18 October 1996
Separation of d- and /-enantiomers of centchromanand its corresponding quaternary methyl iodide saltshas been effected for the first time by chiral high per-formance liquid chromatography (HPLC) to monitortheir enantiomeric purity.
Centchroman, trans-1-[2-[ 4-(7 -methoxy- 2, 2-dime-thyl-3-phenyl-3, 4-dihydro-2H-1-benzopyran-4-yl)-phenoxy]ethyl]pyrrolidine hydrochloride 1, an anties-trogen developed in this Institutehas been recently in-troduced in the market as the first non-steroidal, or-al, weekly pill for female contraception l.It is un-der phase III clinical trials for the treatment ofbreast cancer '. Its activity has also been claimedin the treatment of osteoporosis", psoriasis, res-tenosis", dermatitis", cholesterol lowering" andother estrogen related disorders.
Centchroman has been subsequently resolved?into its d- and i-antipodes. Biochemical and bio-logical studies revealed that i-centchroman bindsto the estrogen receptor with seven fold higher af-finity (RBA= 15.7) as compared to d-centchroman(RBA = 2.1) and is the active compo-nent of di-centchroman in inhibiting implanta-tion? Thus, i-centchroman appears to have agreater potential as a drug. For a detailed study ofi~centchroman, including its clinical investigation,standardisation of enantiomeric purity status ofthe compound obtained on resolution, was re-quired.
Determination of the enantiomeric purity of d-and /-enantiomers was carried out by an enan-tioselective high performance liquid chromatogra-phy (HPLC) method employing a chiral 'chiradex'column, generally used for the separation of basiccompounds" which is reported here.
Further, in order to determine the absolutestereochemistry of centchroman enantiomers,
t CDRI Communication No. 5254
Note
1: R=H, X=CI
~: R = CH3 , X = I
these were converted into their quaternary methyliodide salts 2 for X-ray analysis. Separation of thetwo enantiomeric methyl iodide salts of centchro-man for establishing their purity, was alsoachieved on chiradex column.
Materials and MethodsMaterials di-Centchroman HCl, was synthes-
ized in our laboratory and resolved into its d- andi-forms by optical resolution? Methanol (HPLCgrade) and triethylamine were purchased from Al-drich. Triple distilled water was used throughoutthe experiments.
The HPLC equipment consisted of 501 WatersLiquid Chromatograph equipped with a UV-VISdetector set at 280 nm.
The chiral HPLC column 'chiradex' was pur-chased from merck.
Method. Separation of d/-centchroman was car-ried out under the following conditions. Column:Lichrocart 250-4 'chiradex', particle size: 100A,length of column: 25 em, diameter of column: 4mm, temperature at the time of experiment: 20-2YC Mobile phase: Buffer 65/35 methanollwater+ 1% triethylamine (pH 4.1)Flow rate: 1 mllminuteChart speed: 0.1 cm/minWavelength: 280 nm.
Results and DiscussionsRetention time of d-centchroman, 9.11 min;
retention time of i-centchroman, 18.16 min;retention time of d-centchroman methyl iodidesalt, 9.05 min;retention time of i-centchroman methyl iodidesalt, 17.43 min
A sharp base line separation for the d-and l-centchromans was obtained on the chiradex co-lumn when a buffer of 65:35 methanol/water +1% triethylamine (pH 4.1) was used (Figure 1J.The identity of d- and /-enantiomers was esta-blished by injecting the resolved d- and l-enantiomers separately.
A record of the optical rotation of an HPLCpure sample of /-centchroman having highest ro-tation [a lmax and that of different batches underpurification [alobs, could be used in evaluating thepercentage optical purity of different batches uti-lizing the following equation".
Percentage optical purity = [alob/ [alm3x x 100
The quaternary methyl iodide salts of d- and l-centchromans 210 when run on the above columnunder identical conditions showed similar reten-tion times as observed with d- and l-centchromans (Figure 1). The identity of the saltswas established on the basis of Fab Mass [m/z472(M-127)] and 'H NMR [&3.13 (s, 3H,N+CH))].
AcknowledgementsGrant provided, by the Council of Scientific and
Industrial Research, New Delhi to one of the au-thors (A T) is thankfully acknowledged. We arealso thankful to Mr K P Sirkar for running HPLC.
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.x ,
NOTES 373
-aVI
II'0'Q.2
-eVI
II~'0
S .Qen >.
J:-IIE
-c;VI
tal
~'0
.2
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o c:~ ~~ .
ce~~vCIIU.'0
Figure I-HPLC separation of Centchroman and its methyliodide salts (Time in min)
6 Ray S, Grover P K, Kamboj V P, Setty B S, Kar A B &Anand N, J Med Chem, 19 1976,276.
7 Salman M, Ray S, Anand N, Agarwal A K, Singh M M,Setty B S & Kamboj V p, J Med Chern. 29 1986, 1801.
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