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The production of cyanobacterial toxins as anatoxin-a in the UK by blue—green algae such as Oscillatoria and Anabaena flos-aquae is a potential health hazard, especially to animals, birds and fish. A sensitive reversed-phase ion-pair high-performance liquid chromatographic (RP-HPLC) method is described for the measurement of the neurotoxins anatoxin-a (AnTx) and homoanatoxin (HomoAnTx) in the presence of lyophilized algal extract. A base-deactivated C18 column material was used with acetonitrile—phosphate buffer (pH 3) and sodium dodecyl sulfate as ion-pair reagent with ultraviolet detection at 227 nm. Linear calibrations were obtained between 2 and 93 ng on-column for AnTx·HCI and 2–112 ng on-column for HomoAnTx·HCI with limits of detection of 1 and 2 ng on-column, respectively. A sample of Oscillatoria bloom material collected from Loch Insh, Scotland, in 1991 was found by this method to contain approximately 0.8 mg of AnTx·HCl per gram of lyophilized extract. Homoanatoxin was not present. The identity of AnTx was confirmed by the isolation of the AnTx peak by RP-HPLC, its derivatization to N-butylAnTx and its analysis by RP-HPLC and gas chromatography—mass spectrometry.

The separation of organochlorine pesticides and polychlorinated biphenyls from a high concentration of lipid has been achieved using a polymeric column designed for high-performance liquid chromatography. The method, which can be used as a clean-up step in residue analysis procedures, has a high lipid capacity but low solvent consumption. Recoveries of both lipid and selected organochlorine compounds were routinely over 90% at p.p.m. levels.

Codeine and norcodeine have been extracted together from a 7-l batch of microbial transformation liquor using an XAD-4 resin column. The extract was divided equally and used to compare normal-phase (NP) and reversed-phase (RP) preparative high-performance liquid chromatography for the recovery of pure codeine and norcodeine. The conditions for each column were optimised for maximum sample throughput with complete resolution of both compounds. The aqueous fractions of codeine and norcodeine from the RP column were also passed through an XAD-4 resin column to extract the compounds finally, whereas the organic solvent fractions from the NP column did not require this step. The RP procedure gave recoveries of 71.25% of codeine (98.6% purity) and 80.69% of norcodeine (97.5% purity) and the NP procedure gave recoveries of 79.20% of codeine (96.7% purity) and 83.97% of norcodeine (97.2% purity). Owing to its superior sample throughput and lower operating costs, the RP procedure is recommmended for routine use.

A previously described routine procedure involving the use of a silica gel column in determining diosgenin and yamogenin has been improved by using water-containing solvents. The advantages are that there is less variation between duplicate results, that each column can be used at least five times and that component bands are eluted in predictable volumes of solvents. An apparatus and solvent sequence is described that allows twelve columns to be developed simultaneously.The method has been successfully applied to crude extracts from Dioscorea deltoidea tuber and to oily crude extracts from the seeds of Trigonella foenumgraecum(fenugreek) and Balanites aegyptiaca. The over-all error of the procedure, including sampling, extraction and infrared spectrophotometric determination for duplicate analyses of 2·5-g samples of the fenugreek seed used, expressed as a 95 per cent. confidence interval of the mean sapogenin value, was 1·04 ± 0·025 per cent. for diosgenin plus yamogenin, 0·64 ± 0·019 per cent. for diosgenin and 0·40 ± 0·023 per cent. for yamogenin.As the method does not permit the separation of tigogenin from diosgenin nor that of neotigogenin from yamogenin, the results indicate maximum yields for diosgenin and yamogenin in fenugreek seed. The results exclude sterols, steryl esters, dihydroxysapogenins and spirostadienes.

A routine procedure is described for the determination of diosgenin and yamogenin in fenugreek seed. A crude extract is obtained by acidic hydrolysis of the seed, neutralisation and extraction of the insoluble matter with light petroleum. By means of column chromatography this extract is freed from fixed oil and other substances, thus yielding a mixture of diosgenin and yamogenin suitable for infrared spectrophotometric analysis. The results calculated for duplicate analyses of 2·5-g samples of commercial Moroccan seed, expressed as a 95 per cent. confidence interval of the mean sapogenin value, are 0·96 ± 0·017 per cent. for diosgenin plus yamogenin, 0·58 ± 0·008 per cent. for diosgenin and 0·38 ± 0·016 per cent. for yamogenin. The procedure has been found to be satisfactory for column loadings of up to 75 mg of diosgenin plus yamogenin in the presence of up to 600 mg of fixed oil, which is approximately three times the weight of extractive from 2·5 g of Moroccan seed.