Preparative-layer chromatography reverse phase

Another cultured cell line of Catharanthus roseus (EU4A), which does not produce detectable amounts of vinblastine and other bisindole alkaloids, was also examined for its ability to transform 78 (183). Cell-free extracts of the culture line were prepared, and the 35,000 X g supernatant solution was used. Incubations with 2r-tritioanhydiovinblastine yielded a mixture from which radioactive vinblastine (52) was isolated. The labeled vinblastine was reisolated after unlabeled carrier was added and rigorously purified by successive thin-layer chromatography, reversed-phase HPLC, and crystallization to constant specific activity. Boiled extracts could not produce labeled 52, thus supporting the involvement of enzymes in the conversion process. 

Fractions 8 and 9 from the initial flash vacuum chromatography were combined and vacuum flash chromatographed again on a reverse-phase column using methanol. Further purification of the first fraction by preparative layer chromatography on silica using acetone/hexane (1 1) afforded isomeridine (446). 

Now that reversed phase column packing materials have been used successfully in column liquid chromatography for a number of years, success has also been achieved in thin-layer chromatography in that it is now possible to prepare HPTLC pre-coated plates with the same types of surface-modified sorbents and to use them for separations (2 8, 1, 20). 

The separation and identification of natural dyes from wool fibers using reverse-phase high-performance liquid chromotog-raphy (HPLC) were performed on a C-18 column. Two isocratic four-solvent systems were developed on the basis of the Snyder solvent-selectivity triangle concept (1) 10% acetonitrile, 4% alcohol, and 2% tetrahydrofuran in 0.01 M acetic acid and (2)7% acetonitrile, 8% alcohol, and 5% tetrahydrofuran in 0.01 M acetic acid. Samples were also eluted in 30% acetonitrile. Spot tests and thin-layer chromatography were performed on all samples to confirm HPLC results. The systems also were found to be potentially useful in the identification of early synthetic dyes. A system of sample preparation that minimizes the reaction of samples was discussed. The application of this HPLC separation technique to samples from 20th century Caucasian rugs and American samples unearthed from the foundation of Mission San Jose was examined. 

The evaluation of alkamides, as an indicator for standardization, in Echinacea preparations is commonly completed using HPLC. The method of Bauer (1999b) illustrates the most common method to evaluate alkamides using reverse-phase HPLC. In this method, the lipophilic components are separated by gradient elution using water (eluent A) and acetonitrile (eluent B) linearly from 40% to 80% eluent B at 1 ml/min. The separation was completed on a C18 reverse-phase column and detection was at 254 nm. Thin-layer chromatography using silica 60 plates with... 

RCl by using reverse-phase HPLC for separation and purification and concluded that Chl-RC 1 was a preparation artifact formed during the TLC separation step. The conclusion appeared reasonable as it is known that chlorophyll is easily chlorinated at the C20 position and readily hydroxylated on the silica-gel material used in thin-layer chromatography. 

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