Subchronic effects of gum arabic (Acacia) in the rat.

Young Wistar rats were fed gum arabic (GA) at dietary concentrations of 0% (two control groups), 1, 2, 4, 8 and 20% for 13 weeks. The criteria studied were body weights, food and water consumption, urinalysis, liver and kidney weights, clinical chemistry, haematology, and histology. No untoward effects were observed at dose levels below those which caused dietary imbalance. At the top dose, female rats showed a small reduction in kidney weight, caecal enlargement, and changes in serum urea and total CO2. Male rats showed no differences from the control groups at dietary concentrations up to approx. 8%, but food and water consumption, body weight, liver and kidney weights all decreased significantly and caecal enlargement was evident at the top dose tested. There were no histological changes and no significant changes in haematological parameters in male or female rats at the top dose tested. The no-untoward effect concentrations were 8.6% (5.2 g/kg/day) and 18.1% (13.8 g/kg/day) for male and female rats respectively.

Refinement of structures previously proposed for gum arabic and other acacia gum exudates.

In the light of advances in structural gum-chemistry, the analytical data available from earlier analytical and structural studies of gum arabic (Acacia Senegal Willd.), and of the gum exudates from Acacia laeta, A. campylacantha and A. seyal, have been re-interpreted. The structures originally suggested were based on random arbitrary assignments of substituent sugars without an attempt to establish regular structures. Modelling considerations and recalculations show that the data can also be interpreted in terms of more ordered structures. These are consistent with almost all of the available experimental data and give a much clearer insight into the nature and extent of the structural differences shown by the two Acacia gums of commercial importance. Acacia senegal (gum arabic, gum hashab) and Acacia seyal (gum talha).

HPLC assay of melatonin in plasma with fluorescence detection.

We report an HPLC assay for melatonin that incorporates automated injection, methanol/water mobile phase, and fluorescence detection. Plasma samples were extracted by solid and liquid phases. Recovery was > 70% for 1-10 mL of plasma extracted, approximately 40 pg-250 ng of melatonin. Samples were dried and reconstituted in 100 mL/L methanol. Injections were 25 microL or 150 microL, depending on sample concentration, and the melatonin peak was eluted in 380 mL/L methanol. The detection limit of the assay was 6 pg on the column, allowing a practical sensitivity in plasma of 11 pmol/L for 8-mL samples and 34 pmol/L for 2-mL samples. More than 100 plasma samples from volunteers and patients were assayed and the results compared with an established RIA. The mean daytime concentration of melatonin was 20.7 pmol/L (SEM = 1.2) and 18.5 pmol/L (SEM = 1.6) for HPLC and RIA, respectively, and the mean nighttime concentration was 82.4 pmol/L (SEM = 6.5) and 82.2 (SEM = 7.3), respectively.

An enzyme immunoassay for 6-sulphatoxy-melatonin in human urine.

We describe a newly developed enzyme immunoassay (EIA) for the determination of 6-sulphatoxy-melatonin (aMT6s) in human urine, using a aMT6s-bovine serum albumin-horseradish peroxidase (aMt6s-BSA-HRP) conjugate as the enzyme label. The assay incorporates a highly specific antibody raised in rabbits. The EIA has a sensitivity of 2 pg/well (40 pg/ml) with intraassay coefficients of variation of 2.3-6.1% in the range of the assay. The material with the highest level of cross-reactivity was N-acetyl serotonin sulphate, with a relative potency of 0.000078%. One hundred thirty-four urine samples from children and adults at different time points were assayed and the results compared with those from an established radioimmunoassay (RIA) and with a newly developed RIA using the same antibody as the EIA. The correlation coefficient, r, comparing the two RIA's was 0.9869, and the regression equation was log (kit) = 0.9340 log (new) + 0.1213. The correlation coefficient, r, comparing the EIA with the newly developed RIA, was 0.9686, and regression equation log (new) = 0.9674 log (EIA) + 0.0600. The EIA for the measurement of aMT6s in urine represents a new approach in the investigation of pineal function.

The purification and characterization of biological 6-sulphatoxymelatonin and comparison with synthetic 6-sulphatoxymelatonin.

We have purified the major metabolite of melatonin, 6-sulphatoxymelatonin, from urine and compared it to its synthetic counterpart. For preparation of the biological material, oral melatonin was administered to human volunteers and their urine extracted onto Amberlite XAD-2 resin to remove urea; the glucuronide metabolites of melatonin were removed by silica chromatography; and 6-sulphatoxymelatonin was separated from N-acetyl serotonin sulphate, the other sulphate metabolite of melatonin, by preparative thin-layer chromatography. Synthetic 6-sulphatoxymelatonin was produced by reacting 6-hydroxymelatonin with chlorosulphonic acid in dimethylformamide; the reaction mixture was purified on Florisil and preparative thin-layer chromatography was used to remove indolic by-products of the reaction. Elemental and X-ray microanalysis of the biological and synthetic products showed that classical methods used for their purification introduced inorganic impurities, such as silicon- and chlorine-containing compounds, which were not detectable by thin-layer chromatography, infrared spectroscopy, nuclear magnetic resonance spectroscopy, or gas chromatography-mass spectrometry. We introduced further purification steps to remove these inorganic impurities, monitoring the process using elemental and X-ray microanalysis. Extensive characterization of the resulting purified products showed that the biological and synthetic compounds were identical.