Yasushi Hori

Enantioselective analysis of glufosinate using precolumn derivatization with (+)-1-(9-fluorenyl)ethyl chloroformate and reversed-phase liquid chromatography.

We have developed a new analytical method to quantify the DL-homoalanine-4-yl(methyl)phosphinate (DL-GLUF) enantiomers in biological specimens using a reversed-phase high-performance liquid chromatography system with a fluorescence detection system. The derivatization of DL-GLUF enantiomers with (+)-1-(9-fluorenyl)ethyl chloroformate was carried out under mild conditions (40 degrees C for 30 min) without inducing racemization. The lower limit of quantitation was 0.01 microg/ml for both D-GLUF and L-GLUF, and the detection limit was 5 ng/ml. When DL-GLUF enantiomers were added to serum to produce concentrations between 0.1 and 100 microg/ml, the mean recovery rate was at least 93.8%. The recovery rate from urine was also satisfactory.

Quantitative determination of glufosinate in biological samples by liquid chromatography with ultraviolet detection after p-nitrobenzoyl derivatization.

We have established a new HPLC method for derivatizing and quantifying glufosinate (GLUF) in human serum and urine using p-nitrobenzoyl chloride (PNBC). The p-nitrobenzoyl derivative of GLUF (PNB-GLUF) was produced quantitatively over 10 min at room temperature. PNB-GLUF possesses the property of ultraviolet (UV) light absorption with a lambda(max) of 272.8 nm, and was isolated from biological specimens by reversed-phase chromatography using Inertsil Ph-3. In experiments at a UV wavelength of 273 nm, GLUF has a quantitative detection limit of 0.005 microg/ml, and when it was added to both serum and urine to yield concentrations of 0.1-1000 microg/ml, its recovery rate was quite satisfactory: at least 93.8% in all cases. Further, the measured amounts of GLUF in 23 serum samples from patients intoxicated by ingestion of GLUF compared favorably with those obtained by fluorescence derivatization-HPLC using 9-fluorenylmethyl chloroformate (R=0.998). This technique of analysis is, in addition, applicable for Glyphosat, which possesses a chemical structure resembling that of GLUF, and it will be of great use in the determination of these two compounds.

Serum Parathyroid Hormone in Healthy Japanese Women in Relation to Serum 25-Hydroxyvitamin D

The aim of this work was to clarify levels of serum parathyroid hormone (PTH) in healthy adult women and to study the relationship between PTH and 25-hydroxyvitamin D [25(OH)D]. One hundred sixty-nine healthy women aged 40 years or older in a community were studied. The average age of the subjects was 65.3 years (SD 8.2). All subjects had normal serum creatinine levels. Serum intact PTH and 25(OH)D were measured in these subjects. The mean intact PTH concentration was 2.19 pmol/l (SD 1.02). High intact PTH levels above the reference range were observed in four women (2.4%), all of whom were aged 70 years or older. Intact PTH increased with age with a correlation coefficient of 0.192 (p = 0.013). However, there was no correlation (r = -0.125, p = 0.105) between intact PTH and 25(OH)D whose concentrations were more than 37.5 nmol/l. In conclusion, PTH levels of healthy adult Japanese women are lower than previous reports from Western countries, which may be due to the high 25(OH)D levels of the present subjects and/or an ethnic difference. In addition, there is no association between serum 25(OH)D and PTH levels in this Japanese population, supporting a cutoff level of 25(OH)D less than 37.5 nmol/l for the elevated PTH level.

Effects of capsaicin on serum triglycerides and free fatty acid in olive oil treated rats.

Male rats were dosed with capsaicin after the administration of olive oil, and the serum was obtained for analysis of triglycerides and free fatty acids (FFAs). The serum triglycerides level was increased at 2 and 4 hours after olive oil treatment. On the other hand, capsaicin significantly lowered this increase after 4 hours of treatment in the dosage of 100 mg/kg. Total FFA level was also lowered which had previously been increased by the administration of olive oil after 2 hours of treatment, solely the increase in oleic acid (C18:1) levels was lowered among the FFA. The present results indicated that capsaicin decreased only the components of the dosed olive oil. Furthermore, elevation of the serum total FFA concentration was significantly inhibited 2 or 4 hours after the treatment but not at 8 hours, suggesting that a single high dose treatment with capsaicin may inhibit the absorption of lipid in the gastrointestinal tract.

Nutritional covariates of dietary calcium in elderly Japanese women:: results of a study using the duplicate portion sampling method☆

OBJECTIVES The purposes of this study were to determine the intake of nutrients related to bone health, including calcium (Ca), phosphorus (P), sodium (Na), potassium (K), protein, and vitamin D, in elderly Japanese women and to examine possible intercorrelations between Ca intake and other nutrients. METHODS Fifty-three elderly women in a rural community in Japan participated by supplying duplicate meal samples of everything they ate during a 24-h period. Dietary Ca, P, Na, K, protein, and vitamin D intakes were examined. The mean (standard deviation) age of the 53 women was 68.2 y (6.4 y). RESULTS The mean (standard deviation) intakes of dietary Ca, P, Na, K, protein, and vitamin D were 670 (219) mg/d, 1019 (267) mg/d, 4203 (1341) mg/d, 2752 (844) mg/d, 65.2 (18.3) g/d, and 9.90 (8,89) microg/d, respectively. Dietary Ca was significantly correlated with dietary P (r = 0.732, P < 0.0001), Na (r = 0.336, P = 0.0140), K (r = 0.571, P < 0.0001), and protein (r = 0.563, P < 0.0001), but not with vitamin D. After adjusting the data to reflect the total dry weight, dietary Ca was still correlated with P (r = 0.696, P < 0.0001), K (r = 0.423, P = 0.0018), and protein (r = 0.405, P = 0.0029). CONCLUSIONS The intake of dietary nutrients relevant to bone health in ambulatory Japanese elderly women was examined and documented. The potential confounding effects of these nutrients, especially P, K, and protein, should be taken into account when evaluating the effects of dietary Ca on bone health in observational studies.

Determination of Free Fatty Acids in Human Bile by High-Performance Liquid Chromatography

We developed a high-performance liquid chromatography (HPLC) method for free fatty acids (FFAs) analysis in bile. In this method, FFAs were extracted from bile in a single step using an Isolute™ ODS cartridge, derivatized with 9-anthryldiazomethane (ADAM). ADAM was chosen because of its high reactivity with carboxylic acid at room temperature. Then, HPLC was used for separating and quantifying FFAs. This method proved to be simple and time-saving. The mean recovery of FFA added to human gallbladder bile was 97.6%, and the detection limit was 100–250 pg. Using this method, we determined FFA concentrations in the gallbladder bile of 11 gallstone patients. The mean concentration of total FFA was 0.61 (SD = 0.41) mmol/L, and there was wide variation in the individual FFAs.

Effects of capsaicin on biliary free fatty acids in rats.

The effects of capsaicin, a major pungent agent of capsicum fruits, on biliary free fatty acids (FFAs) were studied in male rats. Animals were dosed 100 mg/kg capsaicin after the administration of olive oil, and the bile was obtained for 6 hours continuously after dosing with capsaicin for analysis of FFAs using HPLC methods. Capsaicin significantly decreased the total biliary FFA concentration in the animals which had been previously increased by the administration of olive oil. The main FFAs in the bile of control rats are lauric and palmitic acids, followed by linoleic, oleic, stearic and palmitoleic acids. Capsaicin alone decreased the values of these main FFAs. While lauric, palmitic, linoleic, stearic and arachidonic acids were increased significantly by the treatment with olive oil, elevation of these FFAs was inhibited by the treatment with capsaicin.

Late Metabolic Acidosis Caused by Renal Tubular Acidosis in Acute Salicylate Poisoning.

A 16-year-old man was transferred to our emergency department seven hours after ingesting 486 aspirin tablets. His blood salicylate level was 83.7 mg/dL. He was treated with fluid resuscitation and sodium bicarbonate infusion, and his condition gradually improved, with a decline in the blood salicylate level. However, eight days after admission, he again reported nausea, a venous blood gas revealed metabolic acidosis with a normal anion gap. The blood salicylate level was undetectable, and a urinalysis showed glycosuria, proteinuria and elevated beta-2 microglobulin and n-acetyl glucosamine levels, with a normal urinary pH despite the acidosis. We diagnosed him with relapse of metabolic acidosis caused by renal tubular acidosis.

Geographical variations in the concentration of biliary free fatty acids with anti-mutagenic action

The concentrations and compositions of free fatty acids (FFAs) in human bile, especially of inhibitory free fatty acids (IFFAs), were analyzed in terms of anti-mutagenic effects in relation to the mutagenic activity of bile. Bile samples were collected from patients with cholelithiasis residing in either Niigata or Kochi prefectures of Japan, regions characterized as the highest and lowest risk areas for gallbladder cancer (GBC), respectively. Biliary FFAs and IFFAs were analyzed by high-performance liquid chromatography and mutagenicity was examined in by the Ames test (TA98+S9mix) after blue rayon treatment. There was a tendency for higher biliary FFA and IFFA concentrations in the Kochi subjects, but the proportion of IFFA to the total FFA concentration did not differ between the two areas. There was an inverse correlation between the concentrations of IFFAs and the numbers of revertant colonies in both Niigata and Kochi subjects. However, at a dose of 591 micromol/l, (calculated based on the average amount of IFFAs absorbed in blue rayon) IFFAs did not exhibit anti-mutagenic actions in the blue rayon extracts. Within this range, more positive samples were seen in Niigata than in Kochi, suggesting the presence of more active mutagen(s) in Niigata samples.

Co-mutagenicity of glyco- and tauro-deoxycholic acids in the Ames test.

Mutagenicity and co-mutagenicity of glyco- and tauro-deoxycholic acids (GDCA and TDCA), which are abundant in human bile, were examined by the Ames test. The two chemicals were not mutagenic for themselves to Salmonella typhimurium TA98 and TA100, with and without S9 mix. They enhanced, however, the mutagenic activities of the pro-mutagens, 2-aminoanthracene (2AA) and benzo[a]pyrene (BaP), for both TA98 and TA100 with S9 mix. They were more co-mutagenic for the pro-mutagens on TA98 than on TA100. On TA98, the mutagenic activities of 2AA with GDCA (5 mumol/plate) and with TDCA (5 mumol/plate) were 9.7-fold and 11.8-fold as high as that of the corresponding control (2AA only), respectively. BaP with GDCA (2.5 mumol/plate) and with TDCA (2.5 mumol/plate) showed 2.8-fold and 3.0-fold increases over the corresponding control level (BaP only), respectively. It is hence concluded that GDCA and TDCA may enhance the activity of some mutagens existing in bile.