VITAMINS

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Three classes of cobalamin-binding proteins are known -intrinsic factor which is secreted from gastric mucosa and is necessary for intestinal absorption of a physiological dose of cobalamin, transcobalamin (formerly designated as transcobalamin II) which transports cobalamin in blood stream to peripheral tissues and haptocorrin (also called as R-binder) which is found in various body fluids and binds endogenous cobalamin. We estimated concentrations of cobalamin and cobalamin-binding proteins in serum of normal and various disorders and observed that both increased markedly in chronic myelogenous leukemia. Physicochemical properties of these cobalamin-binders were investigated by gel filtration and various electrophoretic techniques. Shift of isoelectric point of intrinsic factor toward acidic side on binding of cobalamin was demonstrated by elaborate devices. Haptocorrin was purified by affinity chromatography from human colostrum. Using antiserum produced against the purified haptocorrin, we established a radioimmunoassay system for haptocorrin which enabled estimation of concentration of serum haptocorrin even after an injection of a large dose of cobalamin to patients for the purpose of therapy. Intestinal receptor for intrinsic factor was purified using solidified cobalamin-intrinsic factor. Antiserum obtained by immunizing a rabbit with the purified receptor made a radioimmunoassay for the receptor sensitive and specific. Studies on characteristics of solubilised receptor for transcobalamin from cortex of hog kidney suggested the existence of two mechanisms to take up cobalamin-transcobalamin in kidney - a low-specific mechanism with high capacity reported by Lindemans et al. and a specific mechanism for transcobalamin with low capacity reported by us. Human seminal plasma showed high capacity to bind cobalamin which derived from both transcobalamin and haptocorrin. The haptocorrin comprised of many isoproteins with extremely low isoelectric points. Physiological significances of them are, however,m still largely unknown.

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Since a suitable antibody has not been found, radioimmunoassay of biologically active parathyroid hormone (PTH) in rat serum has not been well performed. The present paper describes a sensitive radioimmunoassay system for determination of rat PTH using an antiserum with specific recognition sites for the amino-terminal region of rat PTH provided by Dr. Slatopolsky. The assay range of this system was 6.25~100 pg/tube. The CV values of inter- and intra-assay were below 10% which were sufficient. This assay system was applied to serum of normal and vitamin D-deficient rats. a diurnal variation and age difference in the PTH levels was observed, but no significant difference in inter-strains and sexes was found in normal rats. The serum PTH levels in normal rats were significantly lower than those in vitamin D-deficient rats. In conclusion, the higher PTH levels by this assay system responded to the low serum Ca levels and reflected secondary hyperparathyroidism in vitamin D-deficient rats.

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We have established a new radioimmunoassay (RIA) system for the determination of biologically active PTH in rat serum. The used anti PTH-antibody has been provided by Dr. Slatopolsky of Washington University, U. S. A., which can recognize the N-terminus of rat PTH, and ^<125>I-labeled Tyr^1 rat PTH (2-34) as a radio-labeled ligand. Based on this RIA system, high PTH levels were observed in vitamin D-deficient rats with decreased serum calcium levels. The levels were similar to those of growing young rats and higher than the normal range. The phenomenon observed in young rats might be presumably attributed to their activated bone turn-over. These findings suggest that our established assay system can be used to determine the serum PTH levels in rats and to evaluate the PTH level in bone diseases related to PTH metabolisms.

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Effect of cinchomeronic acid (pyridine-3,4-dicarboxylic acid; CA) on the growth of various microorganisms, Lactobacillus plantarum, Escherichia coli (3 strains), Bacillus subtilis and Saccharomyces cerevisiae, was investigated. As the results, growth-promoting effect was observed in the each E. coli strain. Most remarkable growth-promoting effect was observed in E. coli B at 0.1-100 mM of CA. The decarboxylation of C-3 or C-4 of CA was investigated by incubating [3,4,7,8-^<14>C]CA with the cell-free extract from E. coli B was analyzed with radio-HPlC. No radioactive peak except CA was detected on this analysis. Hence, CA may promotes the growth of E. coli strains without being metabolized into other compounds.

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