First direct resolution of gossypol enantiomers on a chiral high‐performance liquid chromatography phase

Abstract

The first direct resolution of gossypol enantiomers has been achieved by HPLC on a chiral stationary phase consisting of cellulose tris-(3,5-dimethylphenyl carbamate) coated onto microporous aminopropyl-silica eluted in the reverse phase mode. Chirality 11: 46-49, 1999. © 1999 KEY WORDS: enantiomers of gossypol; direct resolution; chiral HPLC phase; carbohydrate carbamate coating; cellulose tris-(3,5-dimethylphenyl carbamate)Gossypol (Fig. 1), a binaphthyl pigment isolated from the cotton plant and also present in a number of other members of the Malvaceae, is chiral due to steric hindrance to rotation about the internaphthyl bond.1 The standard isolation procedure developed for the extraction of gossypol from the cotton plant 1 affords racemic gossypol as a 1:1 complex with acetic acid. However, investigations have shown that there are wide variations in the ratio of gossypol enantiomers present in different species of cotton plant (Gossypium species) and even within different tissues in the same plant. 2-5There has been considerable interest in the pharmacological properties of gossypol and it has been shown to be potent as an anti-spermatogenic agent 1,6,7 and anti-tumor agent, 8,9 and to possess activity against the HIV-1 virus 10 and the Entamoeba histolytica organism responsible for amoebiasis. 11,12 In all these cases, the (−)-enantiomer of gossypol has been shown to be substantially more active than the (+)-isomer. [13][14][15][16][17][18][19][20][21] Interest in studies of the varied biological activities of the enantiomers of gossypol [13][14][15][16][17][18][19][20][21] and of its unequal enantiomeric distribution in different Malvaceae species [2][3][4][5] has resulted in a need for simple and rapid methods for the analysis of the enantiomeric composition of gossypol samples, as well as for large-scale methods for the resolution of the racemic compound. Previous efforts to resolve gossypol enantiomers directly by HPLC on chiral stationary phases have been unsuccessful, and as a result it was necessary to develop indirect methods, in which gossypol was converted into Schiff's base derivatives that could be separated on either chiral or achiral phases. [22][23][24][25] For the first time, we have now identified conditions suitable for the direct resolution of gossypol enantiomers, using a chiral carbohydrate carbamate phase under reverse phase conditions. MATERIAL AND METHODS ChemicalsAminopropyl-silica (Hypersil-APS, Shandon, UK) had the following properties: particle size, 5 µm; pore size, 120 Å.Isocyanates were purchased from Aldrich (Milwaukee, WI). Solvents were either HPLC grade from Carlo Erba (Milan, Italy) or were purified as usual. 26 Cellulose used was Avicel from Merck. HPLC dead times (t 0 ) were estimated using acetonitrile. (±)-Gossypol acetic acid was isolated from cotton seeds and its separated (−) and (+) enantiomers obtained as described previously. 22-24 EquipmentThe HPLC system consisted of a Shimadzu LC-10AD pump (Kyoto, Japan), a Rheodyne 7125 injector fitted with a 20-µl ...

Growth, feed utilization and body composition of tilapia (Oreochromis sp.) fed with cottonseed meal-based diets in a recirculating system

“…The retention time for (+) and (−)‐isomer of gossypol were 2.1 and 3.4 min, respectively, with flow rate of 1.8 mL min −1 at 254 nm. The standard gossypol (+)‐and (−)‐isomers were provided by Dr Quezia B. Cass, Departamento de Química, Universidade Federal de São Carlos, São Carlos, Brazil (Cass et al . 1999).…”

A mixture of cottonseed meal, soybean meal and animal byproduct mixture as a fish meal substitute: growth and tissue gossypol enantiomer in juvenile rainbow trout (Oncorhynchus mykiss)

“…The HPLC system consisted of a Beckman 506 A solvent delivery system equipped with a 20 μl injection loop connected to a 4.6‐mm × 150 mm Shodex C‐18 column (Showa‐Denko, Shoko Co. Ltd, Tokyo, Japan) packed with an octadecyl‐bonded porous silica gel (5 μm), and both a UV detector (Programmable detector module 166, Beckman Instruments Inc., San Ramon, CA) set at 254 nm and an electrochemical detector (Model LC‐4C; BAS, West Lafayette, IN) set applied potential at 0.75 V. The mobile phase was made of 80 ml acetonitrile and 2 m m KH 2 PO 4 (final concentration) dissolved in 100 ml distilled water (HPLC grade) adjusted to pH 3.0 with H 3 PO 4 . Standards of (+)‐ and (–)‐enantiomers of gossypol were provided by Dr Quezia B. Cass, Departamento de Química, Universidade Federal de São Carlos, São Carlos, Brazil ( cass et al, 1999). The retention time for (+)‐ and (–)‐enantiomers of gossypol were 2.1 and 3.4 min, respectively, with a flow rate of 1.8 ml/min.…”

Enantiomeric Mixtures in Natural Product Chemistry: Separation and Absolute Configuration Assignment

“…This is usually done by using acetonitrile or methanol as modifiers in water. The use of buffer solutions is necessary only if the analytes require [ 19 ]. It is recommended to start the screening with the Chirobiotic ® CSPs at polar organic mode for nonionizable enantiomers, or at the polar ionic mode for the ionizable ones.…”

“…These enantiomeric ratios were measured indirectly by liquid chromatography (LC) after conversion of gossypol to its Schiff’s base diastereoisomers using l -phenylalanine methyl ester [ 18 ]. The first direct resolution of racemic gossypol using enantioselective chromatography was carried out almost a decade later [ 19 ]. ( R )-gossypol was found, as expected, in modest excess in the seeds of G. barbadense ( Brasiliense tussac ), but ( S )-gossypol was found in excess in the roots and flowers of different cultivars of this variety and in the seeds of other Gossypium species studied [ 20 ].…”