Yan-Zin Chang

Integration of GC/EI-MS and GC/NCI-MS for simultaneous quantitative determination of opiates, amphetamines, MDMA, ketamine, and metabolites in human hair.

In this paper, the possibility of using a multiple ionization mode approach of GC/MS was developed for the simultaneous hair testing of common drugs of abuse in Asia, including amphetamines (amphetamine, AP; methamphetamine, MA; methylenedioxy amphetamine, MDA; methylenedioxy methamphetamine, MDMA; methylenedioxy ethylamphetamine, MDEA), ketamine (ketamine, K; norketamine, NK), and opiates (morphine, MOR; codeine, COD; 6-acetylmorphine, 6-AM). This strategy integrated the characteristics of gas chromatography-mass spectrometry (GC-MS) using electron impact ionization (EI) and negative chemical ionization (NCI). Hair samples (25 mg) were washed, cut, and incubated overnight at 25 degrees C in methanol-trifluoroacetic acid (methanol-TFA). The samples were extracted by solid phase extraction (SPE) procedure, derivatized using heptafluorobutyric acid anhydride (HFBA) at 70 degrees C for 30 min, and the derivatives analyzed by GC-MS with EI and NCI. The limit of detection (LOD) with GC/EI-MS analysis obtained were 0.03 ng/mg for AP, MA, MDA, MDMA, and MDEA; 0.05 ng/mg for K, NK, MOR, and COD; and 0.08 ng/mg for 6-AM. The LOD of GC/NCI-MS analysis was much lower than GC/EI-MS analysis. The LOD obtained were 30 pg/mg for AP and MDA in GC/EI-MS and 2 pg/mg in GC/NCI-MS. Therefore, the sensitivity of AP and MDA in GC/NCI-MS was improved from 15-fold compared with EI. The sensitivity of AP, MA, MDA, MDMA, MDEA, MOR, and COD was improved from 15- to 60-fold compared with EI. In addition, the sensitivity of 6-AM increased 8-fold through selection of m/z 197 for the quantitative ion. Moreover, K and NK could dramatically improve their sensitivity at 200- and 2000-fold. The integration of GC/EI-MS and GC/NCI-MS can obtain the high sensitivity and complementary results of drugs of abuse in hair. Six hair samples from known drug abusers were examined by this new strategy. These results show that integrating the characteristics of GC/EI-MS and GC/NCI-MS were not only enhancement of the sensitivity but also avoid wrong results and wrong interpretations of correct results.

Simultaneous quantitative determination of amphetamines, ketamine, opiates and metabolites in human hair by gas chromatography/mass spectrometry

A gas chromatography/mass spectrometry (GC/MS) method was developed and validated for the determination of common drugs of abuse in Asia. The method was able to simultaneously quantify amphetamines (amphetamine; AP, methamphetamine; MA, methylenedioxy amphetamine; MDA, methylenedioxymeth mphetamine; MDMA, methylenedioxy ethylamphetamine; MDEA), ketamine (ketamine; K, norketamine; NK), and opiates (morphine; MOR, codeine; COD, 6-acetylmorphine; 6-AM) in human hair. Hair samples (25 mg) were washed, cut, and incubated overnight at 25 degrees C in methanol/trifluoroacetic acid (methanol/TFA). The samples were extracted by solid-phase extraction (SPE), derivatized using heptafluorobutyric acid anhydride (HFBA) at 70 degrees C for 30 min, and the derivatives were analyzed by electron ionization (EI) GC/MS in selected ion monitoring mode. Confirmation was accomplished by comparing retention times and the relative abundances of selected ions with those of standards. Deuterated analogs of the analytes were used as internal standards for quantification. Calibration curves for ten analytes were established in the concentration range 0.1-10 ng/mg with high correlation coefficients (r2 > 0.999). The intra-day and inter-day precisions were within 12.1% and 15.8%, respectively. The intra-day and inter-day accuracies were between -8.7% and 10.7%, and between -5.9% and 13.8%, respectively. The limit of detection (LOD) and limit of quantification (LOQ) obtained were 0.03 and 0.05 ng/mg for AP, MA, MDA, MDMA and MDEA; 0.05 and 0.08 ng/mg for K, NK, MOR and COD; and 0.08 and 0.1 ng/mg for 6-AM. The recoveries were above 88.6% for all the compounds, except K and NK which were in the range of 71.7-72.7%. Eight hair samples from known polydrug abusers were examined by this method. These results show that the method is suitable for broad-spectrum drug testing in a single hair specimen.

Infant With In Utero Ketamine Exposure: Quantitative Measurement of Residual Dosage in Hair

BACKGROUND The drug ketamine is frequently abused for recreational use in Asia, but few studies in humans have focused on the effects of ketamine exposure during pregnancy on the health of neonates. Here, we report a neonate whose mother was suspected of ketamine abuse during pregnancy. The case was confirmed by testing hair samples of the neonate. METHODS Hair samples of the neonate were taken on the first day of referral. Levels of common drugs of abuse in Asia were measured in the hair sample by gas chromatography-mass spectrometry using our previously reported method with modifications. This method was developed and validated to simultaneously quantify levels of amphetamine, ketamine and opiate in human hair. RESULTS The neonate was a female baby, born full term, with a low birth weight of 2250 g. Very high levels of ketamine were detected in the neonates hair, even though the mother stated that she had stopped abusing ketamine during the early stage of pregnancy. The neonate suffered from general hypotonia; moderate cerebral dysfunction was found by electroencephalography. Fortunately, her hypotonia improved gradually within 21 days. CONCLUSION This is the first report of ketamine exposure during late pregnancy detected by hair testing. We noted several clinical features in this case, including the infant being small for gestational age, intrauterine growth retardation, remarkable hypotonia, and poor reflex responses. Although the mother denied the use of ketamine during the late stage of her pregnancy, significant amount of ketamine and norketamine was still found in hair samples (only 2 cm long and 25 mg) from the infant.

Exposure to di(2-ethylhexyl) phthalate in premature neonates in a neonatal intensive care unit in Taiwan.

Objective: Neonates are exposed to high levels of di(2ethylhexyl) phthalate through numerous medical procedures in the neonatal intensive care unit. Our aim was to assess the contribution of specific medical devices to the di(2-ethylhexyl) phthalate exposure of neonates. Design: Prospective. Setting: University hospital. Patients: We recruited 32 premature neonates, 20 with very low birth weight (<1500 g) and 12 with low birth weight (<2500 g), and 31 controls at a neonatal intensive care unit from a medical center in central Taiwan. Interventions: Interventions were based on a clinical need and used standard materials and devices, including endotracheal tubes, continuous positive airway pressure, oxygen hood, intravenous injection, intralipid injection, blood transfusion, orogastric tubes, nasogastric tubes, umbilical venous catheterization, umbilical arterial catheterization, chest tube, and isolate. Measurements and Main Results: We recorded the medical procedures of each subject, collected their urine samples, and determined the urinary concentration of three metabolites of di(2-ethylhexyl) phthalate using reversed-phase highperformance liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry. Median levels of di(2-ethylhexyl) phthalate metabolites in premature neonates treated with an endotracheal tube and orogastric tube or nasogastric tube were significantly higher than those not treated with an endotracheal tube, orogastric tube, or nasogastric tube. Median levels of di(2-ethylhexyl) phthalate metabolites in premature neonates treated with intravenous injection were ≥2-fold higher than those of healthy controls who received intravenous injections (p = .01). Median levels of three di(2-ethylhexyl) phthalate metabolites were similar in verylowbirth-weight and lowbirthweight neonates. Conclusions: These data suggest that polyvinyl chloride-containing devices are the major defining factor in di(2-ethylhexyl) phthalate exposure levels in neonates in the neonatal intensive care unit. We urge the use of polyvinyl chloride-free or alternative materials in medical devices, especially for endotracheal tubes, orogastric tubes, nasogastric tubes, and intravenous tubing in the neonatal intensive care unit. The health effects of high di(2-ethylhexyl) phthalate exposure on premature neonates in the neonatal intensive care unit is worthy of further investigation.

Hydrophilic ionic liquid-passivated CdTe quantum dots for mercury iondetection

A hydrophilic ionic liquid, 1-ethyl-3-methylimidazolium dicyanamide (EMIDCA), was used as a medium for the synthesis of highly luminescent CdTe nanocrystals (NCs) capped with thioglycolic acid (TGA). The synthesis was performed for 8 h at 130 °C, was similar to nanocrystal preparation in an aqueous medium, and used safe, low-cost inorganic salts as precursors. After the reaction, the photoluminescence quantum yield of the CdTe NCs (NC(IL-130)) prepared in EMIDCA was significantly higher than that of the nanocrystals prepared in water (NC(w)) at 100 °C (86% vs. 35%). Moreover, the emission wavelength and particle size of NC(IL-130) were smaller than NC(w) (450 nm vs. 540 nm and 4.0 nm vs. 5.2 nm, respectively). The activation of NC(IL-130) was successful due to the coordinated action of two ligands, EMIDCA and TGA, in the primary steps of the NC formation pathway. An increase or decrease in the synthesis temperature, to 160 °C or 100 °C, respectively, was detrimental to the luminescence quality. However, the quenching effect of Hg²⁺ on the fluorescence signals of the NC(IL-130) was distinctively unique, whereas certain interfering ions, such as Pb²⁺, Fe³⁺, Co²⁺, Ni²⁺, Ag⁺, and Cu²⁺, could also quench the emission of the NC(w). Based on the Perrin model, the quenching signals of NC(w) and NC(IL-130) were well correlated with the Hg²⁺ concentrations in the phosphate buffer (pH 7.5, 50 mM). In comparison with the NC(w), the NC(IL-130) had a high tolerance of the interfering ions coexisting with the Hg²⁺ analyte, high recovery of Hg²⁺ spiked in the BSA- or FBS-containing medium, and high stability of fluorescence quenching signals between trials and days. The NC(IL-130) nanocrystals can potentially be used to develop a probe system for the determination of Hg²⁺ in physiological samples.

Solvent-enhanced microwave-assisted derivatization following solid-phase extraction combined with gas chromatography–mass spectrometry for determination of amphetamines in urine

An approach using microwave-assisted derivatization (MAD) following solid-phase extraction (SPE) combined with gas chromatography-mass spectrometry (GC-MS) was developed to determine amphetamines in urine samples. The parameters affecting the derivatization efficiency - including microwave power and irradiation time - were investigated. Besides, solvent is thought critically important to MAD. Derivatization performance was studied using various solvents and compared with the performance obtained without solvent. Derivatization efficiency was clearly found to be enhanced by the presence of solvent. The highest derivatization efficiencies were obtained in ethyl acetate (EA) under microwave power of 250W for 1min. Calibration curves for all amphetamines were linear over a range from 1 to 1000ng/mL, with correlation coefficients above 0.9992. The intra-day and inter-day precision were less than 15%. The applicability of the method was tested by analyzing amphetamine-abusing subjects urine samples. Accordingly, the solvent-enhanced MAD-GC-MS method appears to be adequate for determining amphetamines in urine.

Antioxidant effect and active components of litchi (Litchi chinensis Sonn.) flower.

The effects of scavenging 2, 2-diphenyl-2-picrylhydrazyl hydrate (DPPH) radicals and inhibiting low-density lipoprotein (LDL) oxidation, and phenolic quantities were used for the activity-guided separation to identify the effective components of litchi flower. The acetone extract of the flower with notable antioxidant capacities was suspended in water and sequentially partitioned with n-hexane, ethyl acetate (EA) and n-butanol. The EA partition with the highest phenolic levels and antioxidant capacities was subjected to silica gel column chromatography. Thirteen fractions (Fr. 1-13) were collected; Fr. 10-12 with higher phenolic levels and antioxidant effects were applied to Sephadex LH-20 column chromatography. Each fraction was further separated into three sub-fractions and the second ones (Fr. 10-II, 11-II, and 12-II) were the best, which two major compounds could be isolated by semi-preparative high performance liquid chromatography (HPLC). Through Mass (MS) and Nuclear Magnetic Resonance (NMR) measurements, they could be identified as (-)-epicatechin and proanthocyanidin A2. Their contents in the litchi flower were 5.52 and 11.12 mg/g of dry weight, respectively. The study was the first time to reveal the effective antioxidant components of litchi flower.

Determination of Di-(2-ethylhexyl)phthalate (DEHP) metabolites in human hair using liquid chromatography–tandem mass spectrometry

BACKGROUND Di-(2-ethylhexyl)phthalate (DEHP) is an endocrine disrupting chemical that is widely used as the major plasticizer for worldwide plastic products. It can cause several toxic effects to human with high dose exposure. In response to the need of human exposure assessment, different biological specimens are taken into account. Compared to blood, urine and other specimens, hair is unique in that it could determine the time period of chemical exposure after several months to years. METHOD The developed method consists of solution incubation, liquid-liquid extraction and stable isotope dilution liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. RESULTS A reliable and sensitive analytical method was developed and validated for the determination of 5 metabolites, mono-(2-ethylhexyl)phthalate (MEHP), mono-(2-ethyl-5-hydroxyhexyl)phthalate (MEHHP), mono-(2-ethyl-5-oxy-hexyl)phthalate (MEOHP), mono-(2-ethyl-5-carboxypentyl)phthalate (5cx-MEPP) and mono-[2-(carboxymethyl)hexyl]phthalate (2cx-MMHP) in human hair. Ten authentic hair specimens were successfully determined and quantitated by the developed method. CONCLUSION The developed LC-MS/MS method can successfully determine specific DEHP metabolites in human hair and has a great potential to assess the long term DEHP exposure of human.

High-Throughput Simultaneous Analysis of Five Urinary Metabolites of Areca Nut and Tobacco Alkaloids by Isotope-Dilution Liquid Chromatography-Tandem Mass Spectrometry with On-Line Solid-Phase Extraction

Background: Areca nut and tobacco are commonly used drugs worldwide and have been frequently used in combination. We describe the use of on-line solid-phase extraction and isotope-dilution liquid chromatography-tandem mass spectrometry for the simultaneous measurement of five major urinary metabolites of both areca nut and tobacco alkaloids, namely, arecoline, arecaidine, N-methylnipecotic acid, nicotine, and cotinine. Methods: Automated purification of urine was accomplished with a column-switching device. After the addition of deuterium-labeled internal standards, urine samples were directly analyzed within 13 minutes. This method was applied to measure urinary metabolites in 90 healthy subjects to assess areca nut/tobacco exposure. Urinary time course of arecoline, arecaidine, and N-methylnipecotic acid was investigated in five healthy nonchewers after oral administration of areca nut water extracts. Results: The limits of detection were 0.016 to 0.553 ng/mL. Interday and intraday imprecision were <10%. Mean recoveries of five metabolites in urine were 97% to 114%. Mean urinary concentrations of arecoline, arecaidine, N-methylnipecotic acid, nicotine, and cotinine in regular areca nut chewers also smokers were 23.9, 5,816, 1,298, 2,635, and 1,406 ng/mg creatinine, respectively. Time course study revealed that after administration of areca nuts extracts, the major urinary metabolite was arecaidine with a half-life of 4.3 hours, followed by N-methylnipecotic acid with a half-life of 7.9 hours, and very low levels of arecoline with a half-life of 0.97 hour. Conclusions: This on-line solid-phase extraction liquid chromatography-tandem mass spectrometry method firstly provides high-throughput direct analysis of five urinary metabolites of areca nut/tobacco alkaloids. Impact: This method may facilitate the research into the oncogenic effects of areca nut/tobacco exposure. Cancer Epidemiol Biomarkers Prev; 19(10); 2570–81. ©2010 AACR.

High throughput screening various abused drugs and metabolites in urine by liquid chromatography-heated electrospray ionization/tandem mass spectrometry

An integrated method of liquid chromatography-heated electrospray ionization/tandem mass spectrometry was evaluated for high throughput screening of various abused drugs in urine. Chromatographic analysis was performed on a C18 reverse phase column using a linear gradient of 10mM ammonium acetate containing 0.1% formic acid-methanol as mobile phase and the total separation time was 7 min. A simple and rapid sample preparation method used was by passing urine samples through a 0.22 microm PVDF syringe filter. The detection limits of the studied abused drugs in urine were from 0.6 ng mL(-1) (ketamine) to 9.0 ng mL(-1) (norcodeine). According to the results, the linear range was from 1 to 1200 ng mL(-1) with relative standard deviation (R.S.D.s) value below 14.8% (intra-day) and 24.6% (inter-day). The feasibility of applying the proposed method to determine various abused drugs in real samples was examined by analyzing urine samples from drug-abused suspects. The abused drugs including ketamines and amphetamines were detected in suspected urine samples. The results demonstrate the suitability of LC-HESI-MS/MS for high throughput screening of the various abused drugs in urine.