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A method is described for enhancing retention of arginine on a pellicular anion-exchange column. Arginine exhibits adjustable increases of retention time that are dependent on the acidity of standard or sample matrix. This effect is based on interactions of the protonated form of arginine with the residual cation-exchange groups on the core beads of pellicular particles. The relative magnitude of retention time shift of arginine is evaluated for identical concentrations of hydrochloric, sulfuric, and perchloric acids. Although the direct addition of acid is very effective in influencing the retention of arginine, it affects peak shapes and retention of other peaks in the chromatographic separation. The new technique—acid coinjection—achieves a similar retention enhancement for arginine with only a minimal effect on the rest of the separation. Detection limits, reproducibility results, and calibration data are presented for the chromatography of amino acids with acid coinjection. Improved resolution of arginine is demonstrated with chromatograms of soybean hydrolysate and cell culture samples.

Oligodeoxynucleotide-containing phosphorothioate backbones have been used to regulate viral as well as cellular gene expression. The studies carried out in tissue culture have shown promising results on the use of oligonucleotide phosphorothioates as antiviral agents and, at present, study is underway to develop these oligonucleotide analogues as chemotherapeutic agents. To analyze and purify oligonucleotide analogues, high-performance liquid chromatography using weak anion exchange column has been described. The separation of oligonucleotide phosphorothioate is found to be length dependent.

Neuraminidase substrates of high specific activity (>300 μCi/μmol) were prepared by reduction of sialyllactose with NaB3H4, followed by separation of the 2 → 3 and 2 → 6 isomers of [3H]sialyllactitol by paper chromatography. Hydrolysis of sialyllactitol by neuraminidase was monitored by measuring the radioactivity in the neutral reaction product, which was separated from the charged substrate by passage over a small anion exchange column. The assay was applied to the neuraminidase activity of cultured human skin fibroblasts. The Km was found to be 1.1 m m for both substrates; the pH optimum, 4.0; the 2 → 3 isomer was hydrolyzed twice as fast as the 2 → 6. In several genetic disorders associated with neuraminidase deficiency, the activity toward both isomers was reduced almost completely (mucolipidoses I and II; Goldberg syndrome), or only partially (mucolipidosis III; adult myoclonus syndrome); however, the relative activity towards the two isomers remained approximately the same in all cases.

A simple and fairly rapid procedure is described for separation of a mixture of five 3′, 5′-cyclic ribonucleotides. The separation was achieved by high pressure liquid chromatography on a Vydac pellicular anion exchange column using a linear gradient of 0.5 to 1.0 m acetic acid as the eluting solvent.

A method is presented for the assay of γ-carboxyglutamate with an amino acid analyzer equipped with an anion-exchange column. The polyacidic nature of γ-carboxyglutamate results in very good separation from other ninhydrin-reactive material.

The quantitative analysis of 2-keto- l -gulonic acid (2-KLG) produced by microbial fermentation is described. 2-KLG is separated from other aldonic and ketoaldonic acids by high-performance liquid chromatography on an Aminex anion exchange column with ammonium formate or potassium phosphate as the eluant. This is a rapid and simple method for routine analysis of a large number of samples generated by fermentation studies. Gas chromatography—mass spectrometry permits the qualitative and quantitative analysis of nanogram levels of 2-keto- l -gulonate in complex media and provides confirmation of the HPLC results. The methodologies presented are useful for the analysis of a number of aldonic and ketoaldonic acids.