Identification of 1,2,3,4,6-Penta-O-galloyl-ß-d-glucopyranoside as a Glycine N-Methyltransferase Enhancer by High-Throughput Screening of Natural Products Inhibits Hepatocellular Carcinoma.

Glycine N-methyltransferase (GNMT) expression is vastly downregulated in hepatocellular carcinomas (HCC). High rates of GNMT knockout mice developed HCC, while overexpression of GNMT prevented aflatoxin-induced carcinogenicity and inhibited liver cancer cell proliferation. Therefore, in this study, we aimed for the identification of a GNMT inducer for HCC therapy. We established a GNMT promoter-driven luciferase reporter assay as a drug screening platform. Screening of 324 pure compounds and 480 crude extracts from Chinese medicinal herbs resulted in the identification of Paeonia lactiflora Pall (PL) extract and the active component 1,2,3,4,6-penta-O-galloyl-ß-d-glucopyranoside (PGG) as a GNMT inducer. Purified PL extract and PGG induced GNMT mRNA and protein expression in Huh7 human hepatoma cells and in xenograft tumors. PGG and PL extract had potent anti-HCC effects both in vitro and in vivo. Furthermore, PGG treatment induced apoptosis in Huh7 cells. Moreover, PGG treatment sensitized Huh7 cells to sorafenib treatment. Therefore, these results indicated that identifying a GNMT enhancer using the GNMT promoter-based assay might be a useful approach to find drugs for HCC. These data also suggested that PGG has therapeutic potential for the treatment of HCC.

Continuous-Flow Synthesis of Deuterium-Labeled Antidiabetic Chalcones: Studies towards the Selective Deuteration of the Alkynone Core.

Flow chemistry-based syntheses of deuterium-labeled analogs of important antidiabetic chalcones were achieved via highly controlled partial C≡C bond deuteration of the corresponding 1,3-diphenylalkynones. The benefits of a scalable continuous process in combination with on-demand electrolytic D2 gas generation were exploited to suppress undesired over-reactions and to maximize reaction rates simultaneously. The novel deuterium-containing chalcone derivatives may have interesting biological effects and improved metabolic properties as compared with the parent compounds.

Phototoxicity of zinc oxide nanoparticles in HaCaT keratinocytes-generation of oxidative DNA damage during UVA and visible light irradiation.

Zinc oxide nanoparticles (nano-ZnO) are one of the most commonly used nanomaterials in industrial products including paints, cosmetics, and medical materials. Since ZnO is a well-known photocatalyst, it is important to further study if nano-ZnO cause phototoxic effect on skin cells under UVA-irradiation and visible light illumination. Human-derived keratinocytes (HaCaT) were treated with 1-20 microg/mL of nano-ZnO (< 50 nm) and then exposed to UVA (0.5-2 J/cm2). Twenty four hours later, cell viability, membrane integrity, and oxidative DNA damage were determined by MTS assay, lactate dehydrogenase (LDH) release, and the formation of 8-hydroxy-2'-deoxyguanosine (8-OHdG) adduct, respectively. High concentration of nano-ZnO (10-20 microg/mL) significantly induced cytotoxicity, whereas 0.5-2 J/cm2 of UVA irradiation dose-dependently aggravated nano-ZnO-induced cell death via induction of LDH release and DNA damage. The level of photocytotoxicity is mainly dependent on the level of reactive oxygen species (ROS) production. UVA irradiation of nano-ZnO in methanol induced lipid peroxidation in a light dose and substrate dose response manner. Electron spin resonance (ESR) spin trapping studies confirmed that both hydroxyl radical and superoxide anion radical were formed during photoirradiation, while nano-ZnO-induced hydroxyl radical formation is not evolved from superoxide. In addition, nano-ZnO dose-dependently induced single strand DNA break in supercoiled phi x 174 plasmid DNA. Under visible light illumination, nano-ZnO induced the LDH leakage, hydroxyl radical generation, and 8-OHdG formation in a dose-dependent manner. Collectively, these results suggest the photocytotoxic and photogenotoxic effects of nano-ZnO on human skin keratinocytes.

TIPdb: a database of anticancer, antiplatelet, and antituberculosis phytochemicals from indigenous plants in Taiwan.

The unique geographic features of Taiwan are attributed to the rich indigenous and endemic plant species in Taiwan. These plants serve as resourceful bank for biologically active phytochemicals. Given that these plant-derived chemicals are prototypes of potential drugs for diseases, databases connecting the chemical structures and pharmacological activities may facilitate drug development. To enhance the utility of the data, it is desirable to develop a database of chemical compounds and corresponding activities from indigenous plants in Taiwan. A database of anticancer, antiplatelet, and antituberculosis phytochemicals from indigenous plants in Taiwan was constructed. The database, TIPdb, is composed of a standardized format of published anticancer, antiplatelet, and antituberculosis phytochemicals from indigenous plants in Taiwan. A browse function was implemented for users to browse the database in a taxonomy-based manner. Search functions can be utilized to filter records of interest by botanical name, part, chemical class, or compound name. The structured and searchable database TIPdb was constructed to serve as a comprehensive and standardized resource for anticancer, antiplatelet, and antituberculosis compounds search. The manually curated chemical structures and activities provide a great opportunity to develop quantitative structure-activity relationship models for the high-throughput screening of potential anticancer, antiplatelet, and antituberculosis drugs.

Agreement between self-reported illicit drug use and biological samples: a systematic review and meta-analysis.

BACKGROUND AND AIMS: Studies often rely upon self-report and biological testing methods for measuring illicit drug use, although evidence for their agreement is limited to specific populations and self-report instruments. We aimed to examine comprehensively the evidence for agreement between self-reported and biologically measured illicit drug use among all major illicit drug classes, biological indicators, populations and settings. METHODS: We systematically searched peer-reviewed databases (Medline, Embase and PsycINFO) and grey literature. Included studies reported 2 × 2 table counts or agreement estimates comparing self-reported and biologically measured use published up to March 2022. With biological results considered to be the reference standard and use of random-effect regression models, we evaluated pooled estimates for overall agreement (primary outcome), sensitivity, specificity, false omission rates (proportion reporting no use that test positive) and false discovery rates (proportion reporting use that test negative) by drug class, potential consequences attached to self-report (i.e. work, legal or treatment impacts) and time-frame of use. Heterogeneity was assessed by inspecting forest plots. RESULTS: From 7924 studies, we extracted data from 207 eligible studies. Overall agreement ranged from good to excellent (> 0.79). False omission rates were generally low, while false discovery rates varied by setting. Specificity was generally high but sensitivity varied by drug, sample type and setting. Self-report in clinical trials and situations of no consequences was generally reliable. For urine, recent (i.e. past 1-4 days) self-report produced lower sensitivity and false discovery rates than past month. Agreement was higher in studies that informed participants biological testing would occur (diagnostic odds ratio = 2.91, 95% confidence interval = 1.25-6.78). The main source of bias was biological assessments (51% studies). CONCLUSIONS: While there are limitations associated with self-report and biological testing to measure illicit drug use, overall agreement between the two methods is high, suggesting both provide good measures of illicit drug use. Recommended methods of biological testing are more likely to provide reliable measures of recent use if there are problems with self-disclosure.

Orai1/CRACM1 overexpression suppresses cell proliferation via attenuation of the store-operated calcium influx-mediated signalling pathway in A549 lung cancer cells.

BACKGROUND: Orai1/CRACM1 is a principal component of the store-operated calcium channels. Store-operated calcium influx is highly correlated with inflammatory reactions, immunological regulation, and cell proliferation. Epidermal growth factor (EGF), which plays an important role in the regulation of cell proliferation, can activate store-operated calcium channels. However, the consequences of Orai1/CRACM1 overexpression in EGF-mediated lung cancer cells growth are not known. METHODS: To investigate the role of Orai1/CRACM1 in EGF-mediated lung cancer cell proliferation, Orai1/CRACM1 plasmids were transfected into cells by lipofection. A cell proliferation assay, immunofluorescence staining, flow cytometry, and real-time polymerase chain reaction were employed to monitor cell proliferation. The calcium influx signals were investigated using a fluorescent-based calcium assay. RESULTS: Transfection of Orai1/CRACM1 plasmids resulted in the inhibition of EGF-mediated cell proliferation. ERK1/2 and Akt phosphorylation were inhibited by Orai1/CRACM1 overexpression. Expression of the cell cycle modulator p21 was induced in the Orai1/CRACM1-overexpressing cells, whereas the expression of cyclin D3 was reduced. Flow cytometry revealed that overexpression of Orai1/CRACM1 resulted in G0/G1 cell cycle arrest. Importantly, Orai1/CRACM1 overexpression significantly attenuated EGF-mediated store-operated calcium influx. In addition, application of 2-APB, a store-operated calcium channel inhibitor, resulted in the inhibition of EGF-mediated cancer cell proliferation. CONCLUSIONS: We conclude that Orai1/CRACM1 overexpression attenuates store-operated Ca(2+) influx that in turn blocks EGF-mediated proliferative signaling and drives cell cycle arrest.

Mifepristone (RU-486®) as a Schedule IV Controlled Drug-Implications for a Misleading Drug Policy on Women's Health Care.

BACKGROUND: Mifepristone (RU-486) has been approved for abortion in Taiwan since 2000. Mifepristone was the first non-addictive medicine to be classified as a schedule IV controlled drug. As a case of the "misuse" of "misuse of drugs laws," the policy and consequences of mifepristone-assisted abortion for pregnant women could be compared with those of illicit drug use for drug addicts. METHODS: The rule-making process of mifepristone regulation was analyzed from various aspects of legitimacy, social stigma, women's human rights, and access to health care. RESULTS AND DISCUSSION: The restriction policy on mifepristone regulation in Taiwan has raised concerns over the legitimacy of listing a non-addictive substance as a controlled drug, which may produce stigma and negatively affect women's reproductive and privacy rights. Such a restriction policy and social stigma may lead to the unwillingness of pregnant women to utilize safe abortion services. Under the threat of the COVID-19 pandemic, the US FDA's action on mifepristone prescription and dispensing reminds us it is time to consider a change of policy. CONCLUSIONS: Listing mifepristone as a controlled drug could impede the acceptability and accessibility of safe mifepristone use and violates women's right to health care.

New psychoactive substances in Taiwan: The current situation and initiative for rational scheduling.

Use of New Psychoactive Substances (NPS) has posed a global threat to public health and the security of the population. As of December 2019, the NPS items identified in total have outnumbered by three to one the controlled substances listed in the 1961 and 1971 UN Drug Conventions. However, most of these NPS have not been scheduled by the United Nations because of their easy modification on the chemical structures to shun control. Currently, the scheduling and control of NPS is mostly at the national level and a rational scheduling of NPS by objective assessments is essential but often lacking. To rationally schedule NPS, the NPS misuse situation was firstly estimated with the Taiwanese Substance Misuse Monitoring and Reporting Systems (SMMRS) from 2006 through 2019. Then, the assessment of drug-related harms with an expert Delphi procedure for drug scheduling was performed. The epidemiological analysis revealed that among 37 substances commonly misused in Taiwan, heroin posed the highest risk, followed by (meth)amphetamine and ketamine. Of note, misuse of NPS, such as ketamine, synthetic cannabinoids (JWHs, AM-2201, XJR-11), synthetic cathinones (MDPV, bk-MDMA, 4 -MMCetc.), phenethylamines (PMMA, FMA, 2C-B, 2C-E etc.), piperazines (BZP, TFMPP) and tryptamines (5-MeO-DIPT) has been on the rise. Though perceived drug-related harms differed among experts with different professional backgrounds, the differences were not significant. Four dimensions of drug-related harms- addiction, misuse, social harm and physical harm- integrated from Nutt's model and scheduling criteria of Taiwan's Statute for the Prevention and Control of Illicit Drugs (SPCID), were further divided into 11 indicators and applied to assess harms of the 37 substances. Among the 11 indicators that corresponded to the four dimensions, 7 had significant prediction capabilities. Additionally, prevalence of misuse nationally was an important predictor of harm assessment. These indicators of harm assessment of drug misuse can help develop a proper scheduling system for the management of controlled/illicit drugs. In conclusions, drug scheduling is the first step toward proper management of drug use problems. Facing the threats of NPS, it is imperative to implement a rational and effective scheduling system for appropriate management. This study provides a mechanism to scrutinize, and improve, the current evaluation process for NPS scheduling.

Pharmaceutical Assessment Suggests Locomotion Hyperactivity in Zebrafish Triggered by Arecoline Might Be Associated with Multiple Muscarinic Acetylcholine Receptors Activation.

Arecoline is one of the nicotinic acid-based alkaloids, which is found in the betel nut. In addition to its function as a muscarinic agonist, arecoline exhibits several adverse effects, such as inducing growth retardation and causing developmental defects in animal embryos, including zebrafish, chicken, and mice. In this study, we aimed to study the potential adverse effects of waterborne arecoline exposure on zebrafish larvae locomotor activity and investigate the possible mechanism of the arecoline effects in zebrafish behavior. The zebrafish behavior analysis, together with molecular docking and the antagonist co-exposure experiment using muscarinic acetylcholine receptor antagonists were conducted. Zebrafish larvae aged 96 h post-fertilization (hpf) were exposed to different concentrations (0.001, 0.01, 0.1, and 1 ppm) of arecoline for 30 min and 24 h, respectively, to find out the effect of arecoline in different time exposures. Locomotor activities were measured and quantified at 120 hpf. The results showed that arecoline caused zebrafish larvae locomotor hyperactivities, even at a very low concentration. For the mechanistic study, we conducted a structure-based molecular docking simulation and antagonist co-exposure experiment to explore the potential interactions between arecoline and eight subtypes, namely, M1a, M2a, M2b, M3a, M3b, M4a, M5a, and M5b, of zebrafish endogenous muscarinic acetylcholine receptors (mAChRs). Arecoline was predicted to show a strong binding affinity to most of the subtypes. We also discovered that the locomotion hyperactivity phenotypes triggered by arecoline could be rescued by co-incubating it with M1 to M4 mAChR antagonists. Taken together, by a pharmacological approach, we demonstrated that arecoline functions as a highly potent hyperactivity-stimulating compound in zebrafish that is mediated by multiple muscarinic acetylcholine receptors.

Validation study of a new reconstructed human epidermis model EPiTRI for in vitro skin irritation test according to OECD guidelines.

Following the global trend of reducing animal testing, various reconstructed human epidermis (RHE) models for skin irritation test (SIT) have been developed, verified, validated and included in OECD TG 439. We developed a new RHE called EPiTRI and a SIT method using EPiTRI (EPiTRI-SIT model) following the OECD guidelines. EPiTRI possesses morphological, biochemical and physiological properties similar to human epidermis with well-differentiated multilayered viable cells with barrier function. The EPiTRI-SIT model was tested for 20 reference chemicals in Performance Standard of OECD TG 439 (GD 220), showing good predictive capacity with 100% sensitivity, 70% specificity and 85% accuracy. EPiTRI had sensitivity in detecting di-n-propyl disulphate, as an irritant chemical (UN GHS Category 2), whereas most validated reference methods detected it as a non-irritant. An international validation study of EPiTRI-SIT was conducted in four laboratories to confirm the within- and between-laboratory reproducibility, as well as predictive capacity. The phase I/II within-laboratory and between-laboratory reproducibility was 100%/95% and 95%, respectively. The overall sensitivity, specificity and accuracy of EPiTRI-SIT was 96%, 70% and 83%, respectively, which fulfilled the OECD criteria. Thus, EPiTRI, meets the criteria of Performance Standards of OECD TG 439 (GD 220) and is suitable for screening irritating chemicals in vitro.

Sub-maximal exercise altered the prednisolone absorption pattern.

PURPOSE: Prednisolone has been widely used due to its relatively short pituitary inhibition and its moderate potency. Exercise is one of the non-pharmacological interventions for glucocorticoid- induced osteoporosis prevention. However, the effects of exercise on the pharmacokinetics of prednisolone were unclear. The purpose of this study was to evaluate the effects of sub-maximal exercise on the pharmacokinetics of prednisolone. METHODS: A randomized, double-blind, cross-over experimental design was used. Subjects needed to undertake two trials: a non-exercise trial (NE) and an exercise trial (E). After the first blood sampling, the subjects consumed 5 mg of prednisolone with 100 mL of water and then took a rest for 30 min. In the E trial, subjects cycled at 70% VO(2max) intensity until exhaustion. When subjects underwent the NE trial, they remained seated for the duration of the experiment. Serial blood samples were collected at 0.25, 0.5, 0.75, 1, 1.25, 1.5, 2, 3, 4, 5, 6, 8, 10 and 12 hr after prednisolone administration. Prednisolone and cortisol concentration were analyzed with a validated high performance liquid chromatographic method. RESULTS: This study indicated that the maximum concentration of the E trial was significantly lower than the NE trial (p<.05). However, the area under the plasma concentration-time, half life, the time-averaged total body clearance, and the apparent volume of the distribution showed no significant difference between two trials. The mean percentage change of cortisol of E trail was significantly higher than NE trail (p<.05). CONCLUSIONS: The results suggested that sub-maximal exercise altered the prednisolone absorption pattern.

New psychoactive substances in Taiwan: challenges and strategies.

PURPOSE OF REVIEW: Abuse of new psychoactive substances (NPS) has been a new global concern. So far, there has been no international consensus on legislative control of NPS. Scrutiny of Taiwan's illegal drug use history reveals that legislation and policy play an important role in tackling the drug issues. RECENT FINDINGS: Since the early 2000s, use of club drugs (mostly NPS) has become popular in local rave parties and dance clubs in Taiwan. Some NPS, such as ketamine, synthetic cathinones, and para-methoxymethamphetamine, have posed a risk to public health and a challenge to drug policy. SUMMARY: The illegal drug use history in Taiwan was firstly briefly reviewed, and the recent NPS use situation was depicted. Heroin and methamphetamine have been the most predominant drugs, but NPS such as ketamine and synthetic cathinones have become a new issue. The toxicological profiles of commonly abused NPS in Taiwan, although limited, were discussed. By comparison of the legislative mechanism for NPS control between Taiwan, South Korea, and Japan, it was found that timely and flexible legislative mechanism(s) is essential for early identification, surveillance, and comprehensive evaluation. In addition, researches on NPS epidemiology and toxicology are needed to firm up evidence-based strategies for effective prevention, treatment, and harm reduction measures.

Comparison of legislative management for new psychoactive substances control among Taiwan, South Korea, and Japan.

For decades, the three United Nations drug conventions have served as the basis for member states' obligations and international cooperation on drug control. However, the emergence of new psychoactive substances (NPSs) poses a new risk to public health and a challenge to drug policy because of their unknown toxicological effects and easy modification of chemical structures to shun legal control. So far, there is no international consensus on legislative control of NPSs. Therefore, we compared the legislative management on NPS control among Taiwan, South Korea, and Japan. Drug-related information was obtained from the authorities of these three countries. The results indicate that despite geographic proximity and similar legal attitudes toward illegal drug use, the legislative criteria, and responses for NPS control in these three countries were quite different. Ketamine has been the major used NPS in Taiwan but seldom found in South Korea and Japan. The difference in the number of controlled NPSs in Taiwan (91) and South Korea (245) might be due to the implementation of temporary designation systems and analog controls in South Korea. The recent surge of newly controlled NPSs in Japan was because of the promulgation of designated drug regulation and subsequent control of "dangerous drugs." Although NPS use has become a potential social and medical problem among these three countries, the outcomes of NPS legislation control remain to be scrutinized. To minimize harm from NPS use, development of legislative mechanism(s) on NPS scheduling is the first step for early identification and control of NPS problems.

HIV risk and the association with accessibility coverage to medical facilities and socioeconomic status among heroin users in Kaohsiung, Taiwan (2011-2015): A GIS approach.

Illegal drug use has caused considerable health and social costs. Access to medical treatment has been one of the most cost-effective interventions to reduce problematic drug use. Accessibility to treatment, which can be analyzed by the spatial approach such as geographic information system (GIS), has thus been utilized as a measure of the proportion of population that reaches appropriate health services. In this study, the association of accessibility coverage to medical facilities and socioeconomic status with human immunodeficiency virus (HIV) risk for heroin users was evaluated by GIS analysis in Kaohsiung, Taiwan. Data of 7890 heroin users were collected from 2011 to 2015 and categorized into five risk groups according to their income and distance to treatment settings. The results of this GIS-based analysis show that the areas with over 50% accessibility coverage rate had less amounts of HIV cases with statistical significance. Inconvenient access to medical facilities could result in poor treatment outcomes such as higher HIV incidences. Therefore, in order to effectively reduce HIV incidences among the heroin users, the accessibility coverage and locations of medical treatment facilities should meet the needs of drug users. It is also advised that the formulation of policies associated with illegal drug use problems should be evidence-based and geographical indicators could serve for this purpose from either prevention or intervention perspective.

Characterization of cocaine-elicited cell vacuolation: the involvement of calcium/calmodulin in organelle deregulation.

The sizes of organelles are tightly regulated in the cells. However, little is known on how cells maintain the homeostasis of these intracellular compartments. Using cocaine as a model compound, we have characterized the mechanism of deregulated vacuolation in cultured rat liver epithelial Clone 9 cells. The vacuoles were observed as early as 10 min following cocaine treatment. Removal of cocaine led to vacuole degeneration, indicating vacuolation is a reversible process. The vacuoles could devour intracellular materials and the vacuoles originated from late endosome/lysosome as indicated by immunofluorescence studies. Instant calcium influx and calmodulin were required for the initiation of vacuole formation. The unique properties of these late endosome/lysosome-derived vacuoles were further discussed. In summary, cocaine elicited a new type of deregulated vacuole and the involvement of calcium/calmodulin in vacuolation could shed light on prevention or treatment of cocaine-induced cytotoxicity.

New psychoactive substances of natural origin: A brief review.

Plant-based drugs of abuse are as old as recorded human history. Although traditional addictive substances, such as opium, cannabis and coca, have been controlled by the United Nations anti-drug conventions, many, if not most, natural plants with addictive or abuse liability remain elusive. Therefore, the United Nations Office on Drugs and Crime (UNODC) has warned the emerging threat from new psychoactive substances (NPS), which are mostly derived or modified from the constituents of natural origin. For example, synthetic cannabinoids and synthetic cathinones are derived from the cannabis and khat plant, respectively. In this review, we briefly discussed the chemistry, pharmacology and toxicology of five common NPS of natural origin, i.e., khat, kratom, salvia, magic mushroom and mandrake. Through the review, we hope that professionals and general public alike can pay more attention to the potential problems caused by natural NPS, and suitable control measures will be taken.

Immunomodulatory Effects of Taiwanese Neolitsea Species on Th1 and Th2 Functionality.

Neolitsea species, medicinal plants belonging to Lauraceae, contain rich alkaloids, steroids, sesquiterpenoids, and triterpenoids which possess antimicrobial, antioxidant, and anti-inflammatory bioactivities. However, species differences in the immunomodulatory effects and evidence pertaining to the effects of Neolitsea species on adaptive immunity are scarce. This study aimed to evaluate the immunomodulatory properties of ten Taiwanese Neolitsea plants on T helper (Th) cell functionality, especially Th1 and Th2. Most of the 29 crude extracts of Neolitsea were not toxic to splenocytes, except N. buisanensis roots. N. aciculata and N. villosa leaf extracts possessed differential immunomodulatory effects on Th1/Th2 balance. N. aciculata var. variabillima and N. hiiranensis leaf extracts attenuated both Th1 and Th2 cytokines while N. konishii dramatically suppressed IFN-γ production. As N. aciculata var. variabillima and N. konishii leaf extracts significantly attenuated Th1 functionality, we further evaluated their effects on CD4 cells under CD3/CD28 stimulation. N. aciculata var. variabillima significantly suppressed IFN-γ, IL-10, and IL-17, demonstrating the broad suppressive effects on T helper cells; N. konishii significantly suppressed IFN-γ and IL-10 production, while the production of IL-17 was not altered. Collectively, these data demonstrated that leaf extracts of Taiwanese Neolitsea species contain phytochemicals with potentials to be developed as selective immunomodulators.

Prevalence of new psychoactive substances in Northeast Asia from 2007 to 2015.

The proliferation of new psychoactive substances (NPS) has been a global trend in drug abuse and its regulation has been a worldwide concern. There is no doubt that it is necessary to share information related to these emerging substances between countries and continents for the effective regulation of NPS. With efforts for the efficient regulation of NPS, many studies and information have been published for the prevalence of NPS in the United States and other countries in Europe and Oceania. However, there is lack of information available for the prevalence of NPS in Asian and African countries. Therefore, this research was focused on the investigation of legal status of certain NPS in Northeast Asian countries, including China, Japan, South Korea and Taiwan, in order to provide information on the prevalence and trend of emerging NPS in these countries. The results showed that a total of 940 NPS was reported in 4 Northeast Asian countries from 2007 to 2015. Among 940 NPS, 882 NPS are legally restricted in at least one country (94%) and 96 substances were not currently under control (6%) in these countries. The number of controlled NPS that are currently controlled in all 4 countries was only 25 (or 28%) out of 882 NPS. Each substance was categorized in 9 groups according to the classification proposed by the United Nations Office on Drugs and Crime (UNODC). In Northeast Asia, the most commonly controlled NPS were synthetic cannabinoids, synthetic cathinones, and phenethylamines. It was found that Japan is the most proactive country in terms of the NPS regulation with 41% of the total number of controlled NPS in Northeast Asia, followed by South Korea (21%), China (28%), Taiwan (10%). Comparing the number of NPS newly regulated in each country every year, NPS has been broadly scheduled in 2011 and the number of scheduled NPS has dramatically increased from 2013 to 2015. It was shown that Northeast Asia is also in danger of these emerging NPS and the effective regulation across countries is important for the prevention of NPS. Also, this study will bring attention to local law enforcement in the construction of local drug crime prevention network sharing information for these controlled substances.

Direct and sensitive analysis of methamphetamine, ketamine, morphine and codeine in human urine by cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography.

Cation-selective exhaustive injection and sweeping micellar electrokinetic chromatography (CSEI-Sweep-MEKC) was directly used to test some abuse drugs in human urine, including morphine (M), codeine (C), ketamine (K) and methamphetamine (MA). First, phosphate buffer (50 mM, pH 2.5) containing 30% methanol was filled into uncoated fused silica capillary (40 cm, 50 microm I.D.), then high conductivity buffer (100 mM phosphate, 6.9 kPa for 99.9 s) was followed. Electrokinetic injection (10 kV, 500 s) was used to load samples and to enhance sensitivity. The stacking step and separation were performed at -20 kV and 200 nm using phosphate buffer (25 mM, pH 2.5) containing 20% methanol and 100 mM sodium dodecyl sulfate. Using CSEI-Sweep-MEKC, the analytes could be simultaneously analyzed and have a detection limit down to ppb level. It was unnecessary to have sample pretreatments. During method validation, calibration plots were linear (r>or=0.9982) over a range of 150-3,000 ng/mL for M and C, 250-5,000 n g/mL for MA, and 50-1,000 ng/mL for K. The limits of detection were 15 ng/mL for M and C, and 5 ng/mL for MA and K (S/N=3, sampling 500 s at 10 kV). Comparing with capillary zone electrophoresis, the results indicated that this stacking method could increase 6,000-fold sensitivity for analysis of MA. Our method was applied for analysis of 28 real urine samples. The results showed good coincidence with immunoassay and GC-MS. This method was feasible for application to detect trace levels of abused drugs in forensic analysis.

Chemopreventive effect of punicalagin, a novel tannin component isolated from Terminalia catappa, on H-ras-transformed NIH3T3 cells.

Terminalia catappa and its major tannin component, punicalagin, have been characterized to possess antioxidative and anti-genotoxic activities. However, their effects on reactive oxygen species (ROS) mediated carcinogenesis are still unclear. In the present study, H-ras-transformed NIH3T3 cells were used to evaluate the chemopreventive effect of T. catappa water extract (TCE) and punicalagin. In the cell proliferation assay, TCE and punicalagin suppressed the proliferation of H-ras-transformed NIH3T3 cells with a dose-dependent manner but only partially affected non-transformed NIH3T3 cells proliferation. The differential cytotoxicity of TCE/punicalagin on the H-ras-transformed and non-transformed NIH3T3 cells indicated the selectivity of TCE/punicalagin against H-ras induced transformation. TCE or punicalagin treatment reduced anchorage-independent growth that could be due to a cell cycle arrest at G0/G1 phase. The intracellular superoxide level, known to modulate downstream signaling of Ras protein, was decreased by punicalagin treatments. The levels of phosphorylated JNK-1 and p38 were also decreased with punicalagin treatments. Thus, the chemopreventive effect of punicalagin against H-ras induced transformation could result from inhibition of the intracellular redox status and JNK-1/p38 activation.