Hiroshi Seno

Liquid chromatographic–mass spectrometric determination of haloperidol and its metabolites in human plasma and urine

Haloperidol and its two metabolites, reduced haloperidol and 4-(4-chlorophenyl)-4-hydroxypiperidine (CPHP) in human plasma and urine were analyzed by HPLC-MS using a new polymer column (MSpak GF-310), which enabled direct injection of crude biological samples without pretreatment. Recoveries of haloperidol and reduced haloperidol spiked into plasma were 64.4-76.1% and 46.8-50.2%, respectively; those for urine were 87.3-99.4% and 94.2-98.5%, respectively; those of CPHP for both samples were not less than 92.7%. The regression equations for haloperidol, reduced haloperidol and CPHP showed good linearity in the ranges of 10-800, 15-800 and 400-800 ng/ml, respectively, for both plasma and urine. Their detection limits were 5, 10 and 300 ng/ml, respectively, for both samples. Thus, the present method was sensitive enough for detection and determination of high therapeutic and toxic levels for haloperidol and its metabolites present in biological samples.

Determination of tricyclic antidepressants in human plasma using pipette tip solid‐phase extraction and gas chromatography–mass spectrometry

A method for the simultaneous extraction of four tricyclic antidepressants from human plasma samples using pipette tip SPE with MonoTip C(18) tips is presented. Human plasma (0.1 mL) containing four tricyclic antidepressants (amitriptyline, amoxapine, imipramine, and trimipramine) and an internal standard (IS), protriptyline, was mixed with 0.4 mL of distilled water and 100 microL 1 M NaOH solution. After centrifugation of the mixture, the supernatant was extracted to the C(18) phase of the tip by 20 repeated aspirating/dispensing cycles using a manual micropipettor. The analytes retained in the tip were eluted with methanol by five repeated aspirating/dispensing cycles. Without evaporation and reconstitution, the eluate was directly injected into a gas chromatograph injector and detected by a mass spectrometer with SIM in the positive-ion electron impact mode. Recovery of the four antidepressants and IS spiked into human plasma was 80.2-92.1%. The regression equations for the four antidepressants showed excellent linearity in the range of 0.2-40 ng/0.1 mL. LODs and LOQs for the four drugs were 0.05-0.2 ng/0.1 mL and 0.2-0.5 ng/0.1 mL, respectively. Intra- and interday CVs for the four drugs in plasma were no greater than 9.5%.

Comparison of SSI with APCI as an interface of HPLC-mass spectrometry for analysis of a drug and its metabolites

Sonic spray ionization (SSI) was compared with atmospheric pressure chemical ionization (APCI) as an interface of high-performance liquid chromatography (HPLC)-mass spectrometry (MS) for sensitive analyses of a neuroleptic drug, haloperidol and its two metabolites, such as reduced haloperidol and 4-(4-chlorophenyl)-4-hydroxypiperidine (CPHP), in biological samples. For both SSI and APCI interfaces, HPLC-MS-MS gave higher sensitivity than HPLC-MS. The sensitivities by HPLC-SSI-MS-MS for haloperidol and reduced haloperidol were 100 and 30 times higher, respectively, than those by HPLC-APCI-MS-MS; no spectrum with recognizable peaks was obtained for CPHP with the APCI interface. Therefore, detection limits and regression equations were examined by the HPLC-SSI-MS-MS for human plasma and urine samples spiked with the above drug and its metabolites. Haloperidol, reduced haloperidol, and CPHP showed good linearity in the ranges of 5–800, 10–800, and 100–800 ng/mL, respectively, for both human plasma and urine; their detection limits were 2.5, 5, and 75 ng/mL, respectively, using a new polymer HPLC column which enabled direct application of biological samples.

Pipette tip solid-phase extraction and gas chromatography-mass spectrometry for the determination of mequitazine in human plasma.

Mequitazine has been found to be extractable from human plasma samples using MonoTip C(18) tips, inside which C(18)-bonded monolithic silica gel was fixed. Human plasma (0.1mL) containing mequitazine and cyproheptadine as an internal standard (IS) was mixed with 0.4mL of distilled water and 25muL of 1M potassium phosphate buffer (pH 8.0). After centrifugation of the mixture, the supernatant fraction was extracted to the C(18) phase of the tip by 25 repeated aspirating/dispensing cycles using a manual micropipettor. The analytes retained on the C(18) phase were then eluted with methanol by five repeated aspirating/dispensing cycles. Without evaporation and reconstitution, the eluate was injected into a gas chromatograph injector and detected by a mass spectrometer with selected ion monitoring in the positive-ion electron impact mode. The separation of mequitazine and the IS from each other and from impurities was generally satisfactory using a DB-1MS capillary column (30mx0.32mm i.d., film thickness 0.25mum). The recoveries of mequitazine and the IS spiked into plasma were more than 90.0%. The regression equation for mequitazine showed excellent linearity in the range of 0.2-200ng0.1mL(-1), and the detection limit was 0.05ng0.1mL(-1)of plasma. The intra-day and inter-day coefficients of variation for mequitazine in human plasma were not greater than 8.16 and 9.24%, respectively. Accuracy for the drug was in the range of 90.0-97.4%. The data obtained from determination of mequitazine in human plasma after oral administration of the drug are also presented.

SENSITIVE ANALYSIS OF TETRODOTOXIN IN HUMAN PLASMA BY SOLID-PHASE EXTRACTIONS AND GAS CHROMATOGRAPHY/MASS SPECTROMETRY

A method to determine tetrodotoxin (TTX) in human plasma by solid-phase extractions and gas chromatography/mass spectrometry (GC/MS) is described. TTX in plasma samples was passed through Sep-Pak PS-2 extraction cartridges. Then the compound was converted to 2-amino-6-hydroxymethyl-8-hydroxyquinazoline (C9-base) under alkaline conditions and extracted with use of an OASIS HLB extraction cartridge. The C9-base was analyzed after trimethylsilyl derivatization by GC/MS-selected ion monitoring. Recovery rates were 35.3–50.8% and their coefficients of variation were not greater than 16.6%. The calibration curve was linear in the range of 0.5–10.0 ng ml−1 and the detection limit was 0.1 ng ml−1. The coefficients of within-day and day-to-day variations were not greater than 11.8%. Actual determination of TTX in rat plasma after intraperitoneal administration was also presented.

High-throughput determination of theophylline and caffeine in human serum by conventional liquid chromatography-mass spectrometry

Automated high-performance liquid chromatography/mass spectrometry (HPLC-MS) with backflush column-switching was established for ultra-fast determination of theophylline and caffeine. A 400-μl portion of serum sample diluted with ultrapure water was injected and transferred to an Oasis HLB cartridge used as a precolumn for extraction. After switching the valves, the analytes trapped in the precolumn were eluted in the backflush mode and separated with a Chromolith Performance RP-18e column (C18-bonded monolithic silica); the compounds in column effluents were then detected by atmospheric pressure chemical ionization (APCI)-MS. The present method successfully provided high-throughput determination of theophylline and caffeine within 2 min. Satisfactory linearity, reproducibility, and sensitivity could be obtained for analysis of therapeutic and toxic levels of both compounds. Because of the very simple procedure and high throughput using the conventional HPLC system, the present method seems to have high potential in the fields of forensic toxicology and emergency medicine.

2. An Unusual Death due to the Impalement of a Gear Stick into the Brain Stem through the Nasal Cavity

A 59-year-old male was driving a car on the road and was involved in a traffic accident, colliding with a tanker and a big lorry. When an ambulance arrived at the scene, he was in a state of cardiopulmonary arrest and was bleeding profusely from his right nostril. He was confirmed dead at hospital. The autopsy showed a laceration of the right upper lip extending to the right nostril. In the basal skull there was a notable perforation at the ethmoid bone together with the central part of the sphenoid bone including the sella turcica. In accordance with the basal skull bone fractures, there were pronounced contusion injuries at the brain stem and a contusion injury was also observed in the right part of the cerebellum. After careful investigation of a causative stick-like item that was present inside the car, it was concluded that a severe movement of the mans body, as a result of the traffic collision, caused the gear stick which was fixed to the steering wheel to become impaled in the mans right nostril. The gear stick passed through the nasal cavity and into the basal skull bones, resulting in fatal brain stem injuries.

Comparison of sonic spray lonization with atmospheric pressure chemical lonization as an interface of liquid chromatography-mass spectrometry for the analysis of some local anesthetics

SummarySonic spray ionization (SSI) was compared with atmospheric pressure chemical ionization (APCI) as an interface for liquid chromatography (LC)-mass spectrometry (MS) for the analysis of some local anesthetics. Peaks at [M+H]+ constituted the base peaks for all compounds by both SSI and APCI, except for prilocaine. The sensitivities by SSI for tetracaine, benzoxinate, dibucaine, bupivacaine and mepivacaine were 4–16 times higher than those by APCI; those by SSI for procaine and lidocaine were equivalent to those by APCI. Only for prilocaine was the sensitivity by SSI two times lower than that by APCI. In view of the higher sensitivities obtained for many local anesthetics by SSI, we established a detailed procedure for the assay of these drugs in human plasma and urine by LC-MS with SSI in combination with a diol-bonded silica gel HPLC column that enabled direct injection of crude biological samples without complicated pretreatment. The recoveries, sensitivities, accuracies and precisions were found satisfactory to quantitate them at their therapeutic levels.

Sensitive determination of pethidine in body fluids by surface ionization organic mass spectrometry.

We have presented a simple and sensitive method for determining pethidine, a narcotic analgesic drug in body fluids by gas chromatography (GC)/surface ionization organic mass spectrometry (SIOMS). Good linearity was obtained in the range of 0.625-25 ng/ml of whole blood and urine by mass chromatography, and in the range of 0.05-2 ng/ml of whole blood by selected ion monitoring (SIM). Pethidine and diphenylpyraline (internal standard) were extracted from body fluids with Bond Elut Certify cartridges; their recoveries were above 95%. The detection limits (signal-to-noise ratio=3) were estimated to be 0.2 ng/ml of whole blood or urine by mass chromatography, 0.02 ng/ml of whole blood by SIM.

Molecularly imprinted solid‐phase extraction for the selective determination of methamphetamine, amphetamine, and methylenedioxyphenylalkylamine designer drugs in human whole blood by gas chromatography‐mass spectrometry

A novel method is described for the extraction of methamphetamine, amphetamine, and methylenedioxyphenylalkylamine designer drugs, such as 3,4-methylenedioxy-methamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxyethylamphetamine, N-methyl-1-(3,4-methylenedioxyphenyl)-2-butanamine, and 3,4-(methylenedioxyphenyl)-2-butanamine, from human whole blood using molecularly imprinted solid-phase extraction as highly selective sample clean-up technique. Whole blood samples were diluted with 10 mmol/L ammonium acetate (pH 8.6) and applied to a SupelMIP-Amphetamine molecularly imprinted solid-phase extraction cartridge. The cartridge was then washed to eliminate interferences, and the amphetamines of interest were eluted with formic acid/methanol (1:100, v/v). After derivatization with trifluoroacetic anhydride, the analytes were quantified using gas chromatography-mass spectrometry. Recoveries of the seven amphetamines spiked into whole blood were 89.1-102%. The limits of quantification for each compound in 200 μL of whole blood were between 0.25 and 1.0 ng. The maximum intra- and inter-day coefficients of variation were 9.96 and 13.8%, respectively. The results show that methamphetamine, amphetamine, and methylenedioxyphenylalkyl-amine designer drugs can be efficiently extracted from crude biological samples such as whole blood by molecularly imprinted solid-phase extraction with good reproducibility. This extraction method will be useful for the pretreatment of human samples before gas chromatography-mass spectrometry.