Methamphetamine (MA) is the most common drug of abuse in Japan. MA is produced by chemical synthesis and final products of the drug contain small amounts of precursor chemicals, intermediates and by-products. Chirality is a principal characteristic of MA for evaluating the precursor chemicals. Adulterants such as dimethyl sulfone might be associated closely with trafficking routes of illicit drugs. Drug profiling is a scientific tool to identify the synthesis route, the sources of supply, trafficking routes and connections between seizures, which supports drug law enforcement agencies in their effort to eliminate organized drug crime. This review summarizes methods for characterization and profiling of MA hydrochloride, mainly based on our findings.

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In order to enhance its cationic sorption capacity, waste sawdust was functionalized by monosodium glutamate (MG) to produce potentially biodegradable cationic sorbent. The crude sawdust (CS) and functionalized sawdust (FS) were compared for the sorption behaviors against methylene blue (MB) in a batch system. The effects of various experimental parameters (e.g. initial pH, sorbent dose, dye concentration, contact time, and temperature etc.) were investigated and the sorption kinetic and thermodynamic characteristics were elucidated. The MB removal ratio on CS and on FS increased as the initial pH increased, and reached to the maximum value beyond pH 5 and pH 6 for FS and CS, respectively. For 250 mg/l of MB solution, a removal ratio of greater than 95% could be achieved with 2.0 g/l or more of FS. The MB removal percentage decreased more significantly on CS than on FS with increasing initial MB concentration. The isothermal data of MB sorbed on CS and on FS followed the Langmuir model and the sorption capacities (Q_m) of CS and FS for MB were 87.7 and 188.7 mg/g, respectively. The MB removal on FS and on CS reached to the equilibrium at about 10 and 36 hr, respectively. The MB sorption processes on FS and CS followed the pseudo-first-order rate kinetics. The sorptions of MB on CS and on FS were spontaneous and exothermic processes, and lower temperatures were favorable for the sorption processes.

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Although 1-(2-phenethyl)-4-(N-propionylanilino)piperidine (fentanyl) is controlled by drug control laws, its slightly modified compounds, which show the same analgesic activities, cannot be controlled legally. Among these fentanyl analogues, 1-[2-(2-methylphenethyl)]-4-(N-propionylanilino)piperidine (α-methylfentanyl) is the typical and most widely abused drug and its overdose has caused a number of fatalities. Analysis of the urine of addicts has been widely performed to detect its metabolites and the unchanged compound for proof of its abuse. In this case, the metabolites detected in urine should reflect the structure of the original compound. In the present report, for clarification of α-methylfentanyl abuse, four novel metabolites, which reflect the original structure of α-methylfentanyl, were identified in rat urine. One of these was the p-hydroxy form of the aromatic ring of the α-methylfentanyl phenethyl group (mono-aromatic hydroxy α-methylfentanyl), while the second and third ones were metabolites of ω-1 or ω position hydroxypropionyl of α-methylfentanyl (mono-hydroxypropionyl α-methylfentanyl). The fourth one was a metabolite involving the p-hydroxy form of the aromatic ring of the phenethyl group and ω position of hydroxypropionyl α-methylfentanyl (di-hydroxy α-methylfentanyl). The structures of these compounds were identified by comparisons of their retention times and mass spectra obtained by gas chromatography-mass spectrometry (GC/MS) and mass chromatography of mono- and di-hydroxy α-methylfentanyl with those of the synthesized authentic compounds.

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We have previously reported that genipin, a natural iridoid compound, shows neuritogenic activity in cultured rat pheochromocytoma PC12h and mouse neuroblastoma Neuro2a cells. 6-Hydroxydopamine (6-OHDA) is a dopaminergic neurotoxin putatively involved in the pathogenesis of Parkinson's disease (PD). Here, we studied the protective effects of genipin on 6-OHDA-induced cytotoxicity in Neuro2a cells. 6-OHDA treatment markedly reduced Neuro2a cell viability in a concentration-dependent manner causing DNA condensation and fragmentation. Genipin significantly protected the cells against the 6-OHDA-induced cytotoxicity. Genipin also protected the cells against hydrogen peroxide (H₂O₂)-induced cytotoxicity. It is known that 6-OHDA is rapidly and non-enzymatically oxidized by molecular oxygen to form H₂O₂ and the corresponding p-quinone. These data suggest that genipin is effective at protecting against neurodegeneration that involves oxidative stress, such as PD.

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Kothala himbutu (KT) is a traditional medicinal plant used in treating diabetes in Ayurvedic medicine. We investigated the effect of the aqueous extract of KT stems (KTE) on energy expenditure in normal mice. Male C57BL/6J mice (n=28) were divided into 4 groups depending on the type of diet they were fed for 9 weeks: normal (N) diet group (N: 13.8% energy in the form of fat), high-fat (HF) diet group (HF: 53.0% energy in the form of fat), 0.1% freeze-dried KTE (KTED)-supplemented N diet group (N+KTED), and 0.1% KTED-supplemented HF diet group (HF+KTED). KTED intake significantly reduced body weight gain in mice in the N and HF groups. Although it did not affect the plasma levels of glucose, triglyceride, and nonesterified fatty acid, KTED significantly decreased the HF diet-induced increased plasma insulin level. The epididymal and perirenal white adipose tissue (WAT) weights were significantly lower in the HF+KTED group than in the HF group. The oxygen consumption (VO₂), measured by indirect calorimetry, of the mice in the KTED-supplemented groups was significantly higher than that of the mice in the N and HF control groups. Moreover, KTED significantly reduced the size of epididymal WAT adipocytes in the N and HF groups. Thus, KTED promoted VO₂ and suppressed WAT accumulation in the mice on the N and HF diets. Therefore, KTE is beneficial in reducing N diet- and HF diet-induced obesity, which may be partly attributable to the stimulation of whole body energy metabolism.

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Dried apple peels were extracted with n-hexane, chloroform, and methanol successively. The portion of the chloroform extract that showed the strongest cytotoxic activity was purified by silica gel chromatography to isolate ursolic acid (UA). The amount of the isolated UA was 0.71% of the dried peels. Normal mouse embryo cells [serum-free mouse embryo (SFME) cells] and tumorigenic human c-Ha-ras- and mouse c-myc-transformed SFME cells [r/m highly metastatic (HM)-SFME-1 cells] were treated with various concentrations of UA (2.5-20 μM) to investigate its effects on cell growth. UA at 10 μM appeared very effective at suppressing the tumor cell growth, affecting more than 82% of r/m HM-SFME-1 cells, while it inhibited cell growth in only about 7% of SFME cells. Tumorigenic r/m HM-SFME-1 cells were also treated with various concentrations (2.5-10 μM) of epidermal growth factor (EGF) or aminoguanidine (AG) in the presence of UA (2.5-10 μM). Neither EGF nor AG seemed to have any effect on UA-inhibited cell growth. In the present study, it is revealed that UA could be a very effective and promising agent for antitumor treatments, as it specifically affects tumorigenic cells yet appears to cause very little harm to normal cells.

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A rapid and simple method to determine taurine in energy drinks by pre-column high-performance liquid chromatography was developed using a derivative of 4-fluoro-7-nitrobenzofurazan (NBD-F) without the need for an exclusive instrument. The reaction of taurine with NBD-F finished in 10 min at 60°C. The derivative was measured on a UV-Visible detector (470 nm) by HPLC using a conventional Octadecyl silane (ODS) column. A mixture of disodium hydrogenphosphate-citric acid buffer solution (pH 5.4) containing 10 mmol/l tetrabutylammonium bromide and acetonitrile (7:3) was used as the mobile phase. The recoveries were in the range of 98.2-99.9%, the precision as standard deviation was in the range of 0.3-0.5%, the linearity as a coefficient of correlation value was 0.999 and the specification was confirmed for taurine added to three commercial energy drinks. The content of taurine measured compared to the labeled amount in five commercial energy drinks containing taurine was 92.9-105.1%.

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The analytical method to determine organochlorine pesticides in natural medicines included in The Japanese Pharmacopoeia (JP), 15th edition,¹⁾ is not adequate because the recovery rates of organochlorine pesticides in Glycyrrhizae radix are very low. In this study, we developed a method to analyze organochlorine pesticides in Glycyrrhizae radix with acceptable recovery rates. The method enables analysis of organochlorine pesticides in all natural medicines for which maximum residue levels are set.

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Helicobacter pylori (H. pylori) infection is common worldwide. Although the seropositivity of H. pylori rates has been unclear in the Turkish population. In this study, anti-H. pylori IgG seroprevalence and anti-cytotoxin-associated gene A (CagA) IgG positivity were evaluated. The sera of 880 people without gastrointestinal symptoms (384 males, 496 females) were tested for anti-H. pylori IgG and anti-CagA IgG antibodies by enzyme linked immunoassay method. Anti-H. pylori IgG antibodies were positive in 263 sera (41%) and their rates increased with age. The seroprevalence of anti-H. pylori IgG was higher in females (43.8%) than in males (38%). Of the anti-H. pylori IgG positive sera, 194 (53%) were also positive for anti-CagA IgG. The anti-CagA IgG positivity did not significantly differ with age. However, the lowest rate (46.6%) was determined among individuals 20-29 years of age and the highest rate (62.5%) among individuals over 60 years age. Anti-CagA IgG positivity rates were higher in males (87.5%) than in females (37.5%).

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Diabetes affects about 4% of the global population and management of diabetes without any side effects is still a challenge to the medical system. The present study investigated the possible protective effects of Costus speciosus (Koen.) sm. (C. speciosus) rhizome extracts on biochemical parameters in streptozotocin (STZ)-induced male diabetic Wistar rats. STZ treatment (50 mg/kg, i.p.) caused a hyperglycemic state that led to various physiologic and biochemical alterations. Hexane, ethyl acetate, and methanol crude extracts administered at the dose of 250 mg/kg, 400 mg/kg, and 400 mg/kg, respectively, for 60 days to STZ-induced hypoglycemic and normoglycemic rats. The plasma glucose concentration was significantly (p<0.05) decreased by all three extracts compared with controls. In addition, oral administration of hexane extract significantly decreased glycosylated hemoglobin (HbA_1c), serum total cholesterol, and triglyceride levels, urea, uric acid, and creatinine and at the same time markedly increased plasma insulin, tissue glycogen, serum protein, and high-density lipoprotein (HDL) cholesterol levels. The hexane crude extract also restored the altered plasma enzymes aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP) and acid phosphatase (ACP) levels to near normal. Glibenclamide used as a reference drug (0.6 mg/kg body weight) also produced a significant reduction in the blood glucose concentration in STZ-induced diabetic rats. In summary, the hexane crude extract was found to be more active in comparision with ethyl acetate and methanol extracts. Thus this study shows that the C. speciosus hexane extract has antihyperglycemic and hypolipidemic activity, is able to ameliorate the diabetic state, and is probably a source of hypoglycemic compounds.

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A novel method that combines inductively coupled plasma atomic emission spectrometry (ICPAES) with a coprecipitation technique to determine trace and sub-trace elements in crude drugs is described herein. Yttrium phosphate quantitatively coprecipitated seven elements (Al, Cr, Fe, Zn, Cd, Pb and Bi) at pH 6 in a solution prepared from microwave-digested samples of crude drugs. Some matrix elements such as Na, K, Mg and Ca were effectively removed by this process. The thus coprecipitated elements were then determined by ICPAES with yttrium as an internal standard element. The detection limits (3σ, n=10) were 0.17 mg/kg for Al, 0.004 mg/kg for Cr, 0.03 mg/kg for Fe, 0.13 mg/kg for Zn, 0.004 mg/kg for Cd, 0.19 mg/kg for Pb and 0.06 mg/kg for Bi. The proposed method was successfully utilized to determine the concentration of the above-mentioned seven elements in standard reference materials [National Institute of Standards Technology (NIST) SRM1515, SRM1547 and SRM1575a] and seven kinds of crude drugs.

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Vibrio mimicus (V. mimicus) is a causative agent of human gastroenteritis and food poisoning. Although several toxic or virulence factors have been isolated from the bacterium, an enterotoxic hemolysin is a sole toxin produced by all clinical isolates. In the present study, we found that the antibody against the hemolysin significantly inhibited the fluid-accumulating action of the living cells inoculated into a rabbit ileal loop, and that the hemolysin gene (vmhA) was probably expressed by the bacterium in the ileal loop. Additionally, in spit of the comparable motility and similar proteome profiles, a vmhA mutant revealed the reduced fluid-accumulating activity. Theses findings suggest that the hemolysin contributes to full virulence of V. mimicus.

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The cascade of adenosine 5'-monophosphate-activated protein kinase (AMPK) is known to be a sensor of cellular energy charge. In this context, a paradigm for obesity treatment via the activation of AMPK has attracted the suppliers of complementary medicines and supplements. It is possible that products that increase the concentration of adenosine 5'-monophosphate (AMP) in the body would be efficacious in reducing body weight. However, since there is currently little information concerning the toxicity of AMP, the cytotoxic action of AMP on rat thymocytes was examined by flow cytometry. Incubation of cells with AMP at a concentration of 30 μM or more for 24 hr significantly increased the populations of dead cells, shrunken cells, and cells containing hypodiploidal DNA in a concentration-dependent manner. Z-VAD-FMK, a pan-inhibitor of caspases, attenuated the AMP-induced changes in cell populations. It is concluded that AMP at a concentration of 30 μM or more exerts a cytotoxic action, which is dependent on the activation of caspases.

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The placenta acts as a barrier that protects the fetus from xenobiotics in the maternal blood. Phase II conjugation reactions in the placenta are considered to play an important role in this protective process by transferring polar hydroxyl groups and thereby imparting water solubility that facilitates the excretion of xenobiotics. Since coffee consumption during pregnancy is reported to affect fetal growth and development, we examined its effects upon the conjugation reactions in the rat syncytiotrophoblast cell line TR-TBT 18d-1. We detected a high level of glucuronidation for 1-naphthol, a model compound, in both a dose- and culture time-dependent manner in the TR-TBT 18d-1 cells. However, no sulfation was detected in any of our analyses. Coffee was found to inhibit the glucuronidation of 1-naphthol with an IC₅₀ of 4.5% (v/v). In contrast however, caffeine, which is a major bioactive constituent of coffee, did not show any inhibition at doses of up to 100 μM. Coffee was also found to inhibit UDP-glucuronosyl transferase (UGT) activity towards 1-naphthol in vitro to a similar extent [IC₅₀=1.5% (v/v)] as in intact cells. Moreover, the expression of the UGT 1A6, which is known to mainly catalyze the glucuronidation of 1-naphthol, was not affected by coffee. If coffee inhibits placental glucuronidation during pregnancy, its consumption would increase the fetal plasma concentrations of harmful xenobiotics, potentially affecting normal fetal growth and development.

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Recently, it has been questioned whether mevalonate pyrophosphate decarboxylase (MPD) is predominantly located in the peroxisomes or cytosol. We previously reported that MPD was predominantly present in the cytosol of rat hepatocytes, normal rat kidney cells, or mouse melanoma cells. In the present study, we examined whether MPD was predominantly present in the cytosol of HepG2 (human hepatoma) cells and Cos7 (monkey kidney) cells using digitonin permeabilization. In HepG2 cells permeabilized with digitonin, 90%and 10%of MPD existed in the cytosol and membrane/organelle (M/O) fraction, respectively, while in Cos7 cells permeabilized with digitonin, 20% and 80% of MPD existed in the cytosol and M/O fraction, respectively. These data suggest that the difference in subcellular distribution of MPD is due to the cell type.

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The nitric oxide (NO)/cyclic guanosine 5'-monophosphate (GMP) pathway remains undefined regarding the regulation of lipoprotein lipase (LPL) release. Here, we investigated whether L-arginine (Arg) stimulates the release of LPL from mouse mammary tumor FM3A cells in a time- and dose-dependent manner. L-Arg-stimulated release of LPL activity was inhibited by N^G-monomethyl-L-Arg monoacetate, which is an endothelial NO synthase (NOS) inhibitor. Furthermore, release of enzyme activity was also suppressed by various inhibitors of guanylate cyclase and adenylate cyclase, as well as cyclic GMP- and cyclic adenosine 5'-monophosphate (AMP)-dependent protein kinases (PKG and PKA). L-Arg also increased intracellular cyclic GMP contents as well as intracellular cyclic AMP contents. In addition, the increase in the cyclic AMP contents was reduced by inhibitors of guanylate cyclase and PKG. These results suggest that the stimulatory release of LPL from tumor cells by L-Arg is partly due to activation of cyclic AMP production and PKA activity caused by elevated cyclic GMP production and PKG activity.

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