Evaluation of ketamine abuse using hair analysis: concentration trends in a Singapore population.

This paper presents a method for the detection of ketamine in hair. Hair samples (25 mg) were washed, pulverized, and digested in hydrochloric acid (0.5 M) overnight at 45 degrees C. The samples were extracted by an automated solid-phase extraction procedure, and the extracts were subsequently analyzed using gas chromatography-mass spectrometry in selected ion monitoring mode. Linearity up to 120 ng/mg was obtained for both ketamine and norketamine with correlation coefficients of 0.9987 and 0.9985, respectively. Limit of detection was found to be at 0.4 ng/mg for both drugs while the limit of quantitation was found to be 0.6 and 0.8 ng/mg for ketamine and norketamine, respectively. The validated method was used in the analysis of 91 hair segments obtained from 54 ketamine abusers. Based upon the voluntary confession of the ketamine abusers, a correlation between the amount of ketamine detected and the frequency of abuse was observed.

Practical considerations in acquiring biological signals from confocal microscope: solvent effect and temperature effect.

Fluorescence microscopic imaging (FMI) is one of the fastest growing and most powerful techniques to study cellular activities in a living single cell. FMI has been widely used to monitor the temporal and spatial changes of many important intracellular messengers such as Ca2+, H+ and cAMP. In the course of our study of cellular responses with confocal scanning fluorescence microscopy, we detected two sources of artifacts which may render experimental observations invalid. First, the water content of the DMSO used could affect the efficiency of loading of the fluorescence indicator into cells and also give rise to spurious fluorescence spots. Secondly, apparently spontaneous temperature-dependent oscillations of BCECF fluorescence and cellular pulsations were recorded in cells which might be misinterpreted as natural rhythmic behavior. These were later shown to be artifacts arising from changes in refractive indices of the immersion oil due to small fluctuations in temperature, which in turn leads to random shifts of the focal plane erroneously manifest as signal oscillations. Based on these observations, certain recommendations for the control and elimination of false images are presented.

Purpurogallin inhibits DNA synthesis of murine fibrosarcoma L-929 and human U-87 MG glioblastoma cells in vitro.

We demonstrated, for the first time, that the flavonoid purpurogallin (PPG) at 0.2-0.5 mM inhibits DNA synthesis of murine fibrosarcoma L-929 and human U-87 MG glioblastoma cells in vitro. In the human U-87 MG glioblastoma cell experiments, we found that when cells were incubated with PPG at 0.5 mM for 0.5 and 24 h, about 25 and 50% inhibition of DNA compared with control were observed respectively. In contrast, 0.5 mM Trolox (a more polar analogue of vitamin E) did not inhibit DNA synthesis in both cell lines. These data indicate that PPG inhibits the synthesis of DNA in two distinct tumour cell lines.

Effect of hyperthermia on intracellular pH in human U-87 MG glioblastoma cells.

The effect of hyperthermia at 43 degrees C on intracellular pH (pHi) in human U-87 MG glioblastoma cells was studied by using the fluorescent probe 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein-pentaacetoxymethyl ester. The presence of Na+/H+ antiporter activity in the cells were demonstrated by the Na(+)-dependent increase in intracellular pH (pHi) after cellular acidification in the absence of HCO3-. Hyperthermia at 43 degrees C caused significant decrease in pHi. The acidification was readily reversible by cooling the cells back down to 37 degrees C. The pHi change was inhibited by the addition of 1 mM amiloride in the incubation medium. Amiloride and hyperthermia exhibited a synergistic effect in suppressing thymidine incorporation into the cells.

Effects of hyperthermia on the nucleolar proteins in tumour cells.

The effects of hyperthermia at 41 degrees C and 43 degrees C on the nucleolar protein B23 in Ehrlich ascites tumour (EAT), human glioblastoma U-87 MG and U-373 MG cell lines were studied. Cellular localization of protein B23 was detected by an immunofluorescence technique using monoclonal antibody against protein B23. Diminution of fluorescence in the nucleoli occurred when the cells were treated at high temperature. The decrease in fluorescence level depends on the treatment temperature and duration. Among the three cell lines studied, the U-373 MG glioblastoma was the least responsive to hyperthermia followed by the U-87 MG glioblastoma. The decrease in nucleolar fluorescence of the EAT cells treated at 41 degrees C and 43 degrees C correlated with their subsequent cell survival. Dispersion of the nucleolar argyrophilic granules occurred in EAT cells after heating at 43 degrees C for 1 h. The possible implication of such effect is discussed in relation to the heat-sensitive elements in the nucleolus.

Zinc-deficiency and the development of malignant lymphoma in rats given a single intragastric dose of N-methyl-N-nitrosourea.

Zinc-deficient rats that had received a single dose of N-methyl-N-nitrosourea (MNU) intragastrically developed malignant lymphomas involving the liver, spleen, lung and kidney, as well as the usual epithelial tumours at the site of administration. Interestingly, the incidence of squamous-cell carcinoma of the pharynx was significantly higher in zinc-deficient rats than in control animals. Since MNU does not require metabolic activation, the development of the malignant lymphomas might be related to the generally depressed immunological state of zinc-deficient rats.

The effect of N-nitrosodimethylamine and N-nitroso-N-benzylmethylamine on [3H]thymidine incorporation into the DNA of target and non-target tissues in the zinc-deficient rat.

The influence of dimethylnitrosamine (NDMA), a liver carcinogen and nitrosobenzylmethylamine (NBMA) as esophageal carcinogen on [3H]thymidine incorporation into DNA was studied in the esophagus, liver, forestomach and gastric-stomach of fasted zinc-deficient and pair-fed zinc-sufficient rats, measured 1 h after the thymidine injection. In the untreated animals, dietary zinc deficiency significantly depressed [3H]thymidine incorporation (89%) into the DNA of forestomach only. NDMA, administered 4 h before death at 30 mg/kg, produced 50-55% inhibition in [3H]thymidine incorporation in the esophagus of rats of both dietary groups. This inhibition became more pronounced in the forestomach, reaching 90-94% in the zinc-deficient forestomach and 63-86% in their zinc-sufficient counterparts at NDMA levels ranging from 5 to 20 mg/kg. NBMA at 2 mg/kg produced 60% inhibition in the DNA synthesis of zinc-deficient esophagus and 40% in the corresponding zinc-sufficient ones, this difference being significant at P less than 0.01. On the other hand, [3H]thymidine incorporation in the forestomach DNA was markedly lowered in the presence of NBMA. Recovery of DNA synthesis in the 4 tissues from a single dose of NDMA or NBMA was monitored up to 12 days. Following NDMA injection, [3H]thymidine incorporation in the forestomach of both dietary groups remained inhibited (3% of untreated control) for 5 days, a significant recovery (45% of untreated control) was observed only in the zinc-sufficient animals. Following NBMA injection, [3H]thymidine incorporation was also inhibited in the zinc-deficient esophagus for a longer time than in the zinc-sufficient ones. In autoradiographic studies, the percentage of cells showing 30 or more grains/nucleus was significantly decreased (P less than 0.001) in the NBMA-treated and marginally decreased (P less than 0.05) in the NDMA- or NBMA-treated zinc-deficient and zinc-sufficient rats as compared with the saline-treated zinc-sufficient controls. These results were discussed in the light of our previous findings that NBMA enhanced esophageal tumorigenesis in the zinc-deficient rats and that NDMA, a liver carcinogen produced forestomach tumors in the zinc-deficient but not in the zinc-sufficient rats.