Tohru Imamura

Simultaneous determination of paraquat and diquat in human tissues by high-performance liquid chromatography

A simple, sensitive, reliable, and economical method for simultaneous determination of paraquat dichloride and diquat dibromide in human biological materials has been developed, using high-performance liquid chromatography. The drugs were extracted from the sample with a Sep-Pak C18 cartridge and applied to a chromatograph with the internal standard, L-tyrosine. Paraquat and diquat were clearly separated on the octadecylsilica column with a mobile phase of 0.5% potassium bromide in 5% methanol solution, containing triethylamine (1 ml/l). The pH of the mobile phase was adjusted to 3-4 with 1.3 M phosphoric acid. Two ultraviolet wavelengths were selected, 256 nm for paraquat as well as the internal standard, and 310 nm for diquat. The calibration curves were linear in the concentration range 0.1-10 micrograms/g, and the lower limit of detection was 0.05 microgram/g. We used this method to examine the concentrations of paraquat and diquat in tissues of an individual at autopsy.

Sensitive determination of methomyl in blood using gas chromatography-mass spectrometry as its oxime tert.-butyldimethylsilyl derivative

A sensitive, selective and reliable method was developed to determine methomyl ¿methyl-N-[(methylcarbamoyl)oxy]-thioacetimidate¿, a carbamate insecticide in human blood, using gas chromatography-mass spectrometry. Dimethylglyoxime served as an internal standard (I.S.). Methomyl in the blood was converted to its oxime form by sodium hydroxide. The solution made acidic with hydrochloric acid was poured into a column packed with Extrelut. Methomyloxime and I.S. were eluted from the column with a mixture of dichloromethane-ethyl acetate-chloroform (65:25:10), transformed to tert.-butyldimethylsilyl derivatives, and analyzed by gas chromatography-mass spectrometry in the electron impact mode. The calibration curves were linear in the concentration range from 1 ng/g to 100 ng/g and 100 ng/g to at least 5000 ng/g. The lower limit of detection was 0.5 ng/g. The absolute recoveries were 72-93% and within-day coefficients of variation were 3.1-5.6% at blood concentrations of 10 and 1000 ng/g. Two practical forensic applications are described.

Distribution of drugs in various tissues in a brain dead man.

We examined the distribution of drugs in a 49-year-old brain-dead man. Our objective was to determine the possibility of diagnosing how and at what point the patient became brain dead. The presence of mepivacaine, pentazocine, lidocaine and thiamylal in various tissues, including seven regions of the brain were confirmed, using gas chromatography/mass spectrometry. Tissue-to-blood concentration ratios of mepivacaine, pentazocine and lidocaine in the brain were higher than these ratios in other tissues, while ratios of thiamylal were lower. Therefore, cerebral blood flow was likely to have ceased between the administration of the former drugs and that of the latter drug, in agreement with clinical records. Among seven regions of the brain, the ratios of the former three drugs were high in occipital and parietal lobes, and were low in the cerebellum and medulla oblongata. On the other hand, the ratios of the latter drug were high in the cerebellum and the medulla oblongata. Therefore, cerebral blood flow presumably ceased first in occipital and parietal lobes, and last in the cerebellum and the medulla oblongata. Based on these results, assessment of concentrations of drugs in human tissues, including various regions of brain is useful to determine the time and progression of brain death.

Selective determination of sultopride in human plasma using high-performance liquid chromatography with ultraviolet detection and particle beam mass spectrometry

We developed a sensitive and selective method for determining levels of sultopride, a neuroleptic drug of the substituted benzamide, in human plasma using high-performance liquid chromatography (HPLC) combined with UV detection and particle beam mass spectrometry (PBMS). Sultopride was extracted with tert.-butylmethyl ether using a salting-out technique. Tiapride served as an internal standard (I.S.). Sultopride and I.S. were separated by HPLC on a silica column with a mobile phase of acetonitrile-0.1 M ammonium acetate (94:6, v/v). The calibration curves were linear over the concentration range from 5 to 1000 ng/ml by HPLC with UV detection and from 10 to 1000 ng/ml with PBMS detection. The limit of quantitation was 5 ng/ml with UV detection and 10 ng/ml with PBMS detection. The absolute recovery was 92% and the within-day coefficients of variation were 2.9-7.1% at plasma concentrations from 50 to 500 ng/ml, determined by HPLC with UV detection. Using this method, we measured the plasma concentrations of sultopride with replicate analyses in four hospitalized patients and steady-state plasma levels were determined to be 161.6 +/- 30.8, 321.1 +/- 93.7, 726.5 +/- 143.1 and 1273.6 +/- 211.2 ng/ml, respectively.

Sensitive and selective determination of bromisovalum by high-performance liquid chromatography/particle beam mass spectrometry

A specific procedure using high-performance liquid chromatography/particle beam mass spectrometry (HPLUPBMS) has facilitated determination of bromisovalum in human plasma and whole blood. Bromisovalum, a sedative and hypnotic, was effectively extracted with SepPak C,, cartridges and selectively determined by HPLUPBMS of EI mode. An originally synthesized 2-bromohexanoylurea served as the internal standard (IS). The calibration curve was linear over the concentration range 0.5-5.0 &g and the lower limit of detection was 0.1 r&g (10 ng at the time of direct injection). This method could be practically applied to detect bromisovalum in the whole blood of an autopsied individual.

Identification of vegetable species in gastric contents using HPLC.

Identification of 16 vegetables, focusing on the influence of digestion in the stomach, was carried out on the basis of the types of flavonoids detected on chromatograms using HPLC. Among the 12 vegetables for which flavonoids were detected, the chromatographic patterns of the flavonoids in digested vegetables were the same as those of the corresponding raw vegetables, making it possible to identify the species of vegetables even after digestion. In our analysis, 5 mg of a freeze-dried sample was found to be an adequate quantity to enable the detection of flavonoids. Brief practical cases are also described.