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A mouse hepatic microsomal enzyme that catalyzes the oxidation of cannabinol (CBN) to 11-hydroxycannabinol (11-OH-CBN) was characterized. 11-OH-CBN formed was determined by HPLC. The reaction required NADPH as a cofactor and the optimal pH for the reaction was at 7.8. The Km and Vmax values for the reaction were 122 μM and 8.82 nmol/min/mg protein, respectively. The reaction was inhibited by inhibitors of cytochrome P450 such as SKF 525-A and by cannabidiol. The pretreatment of mice with phenobarbital significantly increased the enzyme activity but not with 3-methylcholanthrene, whereas that with cobaltous chloride significantly decreased the activity. These results indicate that cytochrome P450 plays a major role in the formation of 11-OH-CBN in the mouse hepatic microsomes.

The GC-MS and TLC methods for the determination and identification of Δ9-tetrahydrocannabinol-11-oic acid, a major urinary metabolite of Δ9-tetrahydrocannabinol which is one of main constituents of marihuana, were established. Δ9-Tetrahydrocannabinol-11-oic acid was detected on TLC by using Fast Blue BB salt as a color reagent. The metabolite was determined as trimethylsilyl derivative of its methyl ester by using electron impact ionization GC-MS and 5'-nor-Δ8-tetrahydrocannabinol-4'-oic acid as an internal standard. The assay provides excellent sensitivity, reproducibility and specificity with detection limit of 1 ng/ml human urine.

Tetrahyrocannabinol and cannabinol were identified and determined in cannabis seeds sold as feed for birds by thin-layer chromatography, gas chromatography and gas chromatography-mass spectrometry.

Wet digestion in a microwave oven for rapid determination of total arsenic was examined. Organic matter in samples was digested very rapidly in comparison with other heating methods. We then investigated Hirayama's method, in which nickel is added to samples to prevent loss of arsenic due to volatilization, and confirmed the effectiveness of nickel ion. Thus loss, of arsenic during wet digestion can be prevented by addition of nickel and by shortening the digestion time by the use of a microwave oven. By this method, arsenic contents in various foods could be determined with a recovery of 85.9-104% and a coefficient of variation of 1.8-5.7%. This microwave oven-digestion (MOD) method might also be applicable to the determination of other metals in foods.

A simple colorimetric method for the determination of cannabinoids was developed, using Fast Blue BB salt. Fast Blue BB salt produced stable diazo compounds with cannabinoids in alkaline media. The absorption maxima of Δ8-tetrahydrocannabinol, Δ9-tetrahydrocannabinol, cannabidiol and cannabinol were at 488, 489, 470 and 490 nm, respectively. The molar extinction coefficients of the above cannabinoids were estimated to be 3.3, 2.9, 5.2 and 2.5×104, respectively, and the coefficients of variation (10 analyses) were 0.7, 0.4, 0.4 and 2.7 (%), respectively. The contents of these cannabinoids in extracts from six kinds of cannabis leaves were determined by the colorimetric method in combination with thin-layer chromatography. The results obtained were in good agreement with those obtained by the gas chromatographic method.

A fluorometric determination of H2O2 based on the formation of amprochrome was developed. Amprochrome is an oxidation product of amprolium, an agent used for treating coccidiosis of chickens, and has a strong fluorescence. Though potassium ferricyanide has been used for oxidizing amprolium, H2O2 is also considered to be useful for the reaction. The analytical procedure was as follows. To a sample solution containing a suitable quantity of H2O2, 1 ml of amprolium (500 μg/ml), NaOH (10%) and peroxidase solution (20 units/ml) were added. The mixture was shaken for 45 min at room temperature and amprochrome produced was extracted with 10 ml of isoBuOH after saturating the solution with sodium sulfate. Relative fluorescence intensity of the separated isoBuOH layer was measured (excitation wavelength 400 nm and emission wavelength 460 nm) against that of 10-6M quinine sulfate in 0.1 N H2SO4. The calibration plot was linear up to H2O2 0.2 μg/tube. On the basis of the proposed method, remaining H2O2 in Japanese noodles (udon) could be determined with a recovery of 82.4-108%.

Wet digestion in a microwave oven was examined for rapid determination of Zn, Cu, Mn, Pb and Cd. Analytical results for metals in foods obtained by this microwave oven-digestion (MOD) method agreed well with those obtained by the conventional wet digestion technique (Kjeldahl furnace method) and further, the analytical results for National Bureau of Standards (NBS) standard reference material agreed with the certified values. The values of recovery and coefficient of variation of the metals in food samples were 84.3-113% and 0.1-6.0%, respectively. The results indicate that the MOD method could be useful for the determination of various metals in foods.