Journal of Chromatography B (v.783, #2)
Editorial Board (OFC).
News Section (N1-N2).
Trace level determination of trichloroethylene from liver, lung and kidney tissues by gas chromatography–magnetic sector mass spectrometry by Stacy D Brown; S Muralidhara; James V Bruckner; Michael G Bartlett (319-325).
Trichloroethylene (TCE) is a common industrial chemical that has been heavily used as a metal degreaser and a solvent for the past 100 years. As a result of the extensive use and production of this compound, it has become prevalent in the environment, appearing at over 50% of the hazardous waste sites on the US EPA’s National Priorities List (NPL). TCE exposure has been linked to neurological dysfunction as well as to several types of cancer in animals. This paper describes the development and validation of a gas chromatography–mass spectrometry (GC–MS) method for the quantitation of trace levels of TCE in its target tissues (i.e. liver, kidney and lungs). The limit of quantitation (5 ng/ml) is substantially lower than currently published methods for the analysis of TCE in tissues. The % RSD and % Error for the assay falls within the acceptable range (<15% for middle and high QC points and <20% for low QC points), and the recovery is high from all tissues (>79%).
High-performance liquid chromatographic assay with ultraviolet spectrometric detection for the evaluation of inhibitors of secretory phospholipase A2 by Melanie Schmitt; Matthias Lehr (327-333).
A non-radioactive spectrometric assay for the evaluation of inhibitors of pancreatic group IB and non-pancreatic group IIA secretory phospholipase A2 (sPLA2) is described. Mixed-micelles consisting of 1 mM of a 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol and 6 mM of sodium deoxycholate were used as substrate. The enzyme activity was determined directly without any sample clean-up by measuring the sPLA2-mediated oleic acid release with reversed-phase HPLC and UV-detection at 200 nm. The known sPLA2 inhibitors MJ33 and AR-C 67047MI were analyzed in this assay for their inhibitory potency. While MJ33 revealed only a very weak inhibition of group IB and IIA sPLA2 at the highest test concentration (33 μM), AR-C 67047MI proved to be a potent inhibitor of both enzymes with IC50-values of 0.36 and 0.14 μM, respectively.
Keywords: Secretory phospholipase A2;
Determination of ximelagatran, an oral direct thrombin inhibitor, its active metabolite melagatran, and the intermediate metabolites, in biological samples by liquid chromatography–mass spectrometry by Marita Larsson; Martin Ahnoff; Anna Abrahamsson; Ulrika Logren; Christina Fakt; Irene Öhrman; Bengt-Arne Persson (335-347).
Analytical methods for the determination of ximelagatran, an oral direct thrombin inhibitor, its active metabolite melagatran, and intermediate metabolites, melagatran hydroxyamidine and melagatran ethyl ester, in biological samples by liquid chromatography (LC) positive electrospray ionization mass spectrometry (MS) using selected reaction monitoring are described. Isolation from human plasma was achieved by solid-phase extraction on octylsilica. Analytes and isotope-labelled internal standards were separated by LC utilising a C18 analytical column and a mobile phase comprising acetonitrile–4 mmol/l ammonium acetate (35:65, v/v) containing 0.1% formic acid, at a flow-rate of 0.75 ml/min. Absolute recovery was ∼80% for ximelagatran, ∼60% for melagatran ethyl ester and >90% for melagatran and melagatran hydroxyamidine. Limit of quantification was 10 nmol/l, with a relative standard deviation <20% for each analyte and <5% above 100 nmol/l. Procedures for determination of these analytes in human urine and breast milk, plus whole blood from rat and mouse are also described.
Keywords: Ximelagatran; Melagatran; Thrombin; Esterases;
Cellobiose and lactulose coupled with mannitol and determined using ion-exchange chromatography with pulsed amperometric detection, are reliable probes for investigation of intestinal permeability by Maddalena Generoso; Mario De Rosa; Roberto De Rosa; Laura De Magistris; Mario Secondulfo; Rafffaele Fiandra; Romano Carratù; Maria Cartenı̀ (349-357).
Lactulose/mannitol and cellobiose/mannitol tests are currently used in the investigation of intestinal permeability (IP) in many gastrointestinal diseases. The aim of this study was to produce a good technique for the determination and comparison of the above-mentioned sugar probes to overcome the problem caused by the presence of significant glycosuria in patients affected by particular metabolic disorders such as diabetes mellitus. Tests were performed in 25 healthy volunteers, using either cellobiose (Ce) (5 g) and mannitol (Ma) (2 g), or lactulose (La) (5 g) and mannitol (2 g), given as oral isosmolar loads. Sugars were recovered in urine collected for 5 h. Analysis was carried out by using anion-exchange chromatography (AEC) with pulsed amperometric detection (PAD). Baseline separation of the above carbohydrates was achieved within 13 min by using a Carbopac PA-100 column and linear gradient elution. Carbohydrate quantification was performed by an internal standard method. The calibration curve for each sugar is linear to 40 mM. The limit of sugar detection is 0.01 mM. Recovery of sugar probes is between 98.2 and 100%.The %La, %Ce, %Ma in urine were evaluated and their ratios (Ce/Ma and La/Ma) were calculated. No significant difference in IP parameters were shown (La/Ma to Ce/Ma 0.018±0.014 vs. 0.012±0.007; the attendant probability of the null hypothesis being P=0.0714). Ce/Ma and/or La/Ma tests result similarly reliable in the clinical investigation of IP and the described new method is also helpful in urine even with high glucose concentration, without any interference.
Keywords: Lactulose; Cellobiose; Mannitol; Glucose;
Quantification of clarithromycin, its 14-hydroxy and decladinose metabolites in rat plasma, gastric juice and gastric tissue using high-performance liquid chromatography with electrochemical detection by J.I.D Wibawa; P.N Shaw; D.A Barrett (359-366).
A rapid, selective and sensitive HPLC assay has been developed for the simultaneous analysis of clarithromycin, its 14-hydroxy-clarithromycin metabolite, and its decladinose acid degradation product, in small volumes of rat gastric juice aspirate, plasma and gastric tissue. Sample were extracted with n-hexane/2-butanol (4:1) and the internal standard was roxithromycin. A Kromasil ODS 5 μm (75×4.6 mm I.D.) column was used with a mobile phase consisting of acetonitrile/aqueous phosphate buffer (pH 7, 0.086 M) (45:55 v/v). The column temperature was 30 °C and coulometric detection was used at 850 mV using a screen voltage of 600 mV. The analysis time was less than 8 min. The limits of quantitation for clarithromycin, 14-OH clarithromycin and decladinose clarithromycin were 0.15 μg ml−1 or lower in plasma (0.05 ml); 0.16 μg ml−1 or lower in gastric juice (0.2 ml); and 0.51 μg g−1 or lower for gastric tissue (0.25 g). The method was linear up to at least 20.3, 15.4 and 12.5 μg ml−1 for clarithromycin, 14-OH-clarithromycin and decladinose, respectively, in gastric juice aspirate and plasma and up to 40.6, 30.9 and 25.0 μg g−1 in gastric tissue. The assay was applied to the measurement of clarithromycin, 14-OH-clarithromycin and, for the first time, decladinose clarithromycin in pharmacokinetic studies of gastric transfer of clarithromycin in individual rats.
Keywords: Clarithromycin; 14-Hydroxy-clarithromycin; Decladinose clarithromycin; Antibiotics;
Simple and accurate determination of bisphenol A in red blood cells prepared with basic glycine buffer using liquid chromatography-electrochemical detection by Junko Sajiki (367-375).
For an accurate determination of bisphenol A (BPA) in red blood cells (RBC), the effect of pH on the concentration of BPA was investigated. Also, BPA recovery using ferric heme, methemoglobin (metHb) and hematin, were investigated to confirm whether BPA binds to ferric heme. BPA recovery in hemolysate was high at alkaline pH and was very low at acidic pH where oxyHb changed to metHb. BPA recovery decreased dose-dependently in metHb and hematin, but inorganic iron ions did not influence the recovery. These results suggested that BPA could be bound to ferric heme in RBC. The use of glycine–NaOH buffer (pH 11) as well as plasma had the highest recovery (97%). BPA was not detected in red blood cells of healthy adult volunteers (n=6). In sheep blood contaminated with BPA, BPA was detected in both plasma and RBC (10 times lower than in plasma), indicating that BPA could have migrated from plasma into RBC.
Keywords: Bisphenol A; Glycine buffer, basic;
Determination of clozapine, and its metabolites, N-desmethylclozapine and clozapine N-oxide in dog plasma using high-performance liquid chromatography by Karen E Mosier; Jiuxue Song; Gordon McKay; John W Hubbard; Jim Fang (377-382).
Clozapine and its two major metabolites, N-desmethylclozapine and clozapine N-oxide were quantified using a high-performance liquid chromatographic method with UV detection in dog plasma following a single dose of clozapine. The analysis was performed on a 5-μm Hypersil CN (CPS-1; 250×4.6 mm) column. The mobile phase consisted of acetonitrile–water–1 M ammonium acetate (50:49:1, v/v/v), which was adjusted to pH 5.0 with acetic acid. The detection wavelength was 254 nm. A liquid–liquid extraction technique was used to extract clozapine and its metabolites from dog plasma. The recovery rates for clozapine, N-desmethylclozapine, and the internal standard (I.S.) were close to 100% using this method. The recovery rate for clozapine N-oxide (62–66%) was lower as expected because it is more polar. The quantitation limits for clozapine, clozapine N-oxide, and N-desmethylclozapine were 0.11, 0.05 and 0.05 μM, respectively. Intra-day reproducibility for concentrations of 0.1, 1.0 and 5.0 μM were 10.0, 4.4 and 4.2%, respectively, for N-oxide; 11.2, 4.3 and 4.9%, respectively, for N-desmethylclozapine; and 10.8, 2.2 and 4.9%, respectively, for clozapine. Inter-day reproducibility was <15% for clozapine N-oxide, <8% for N-desmethylclozapine and <19% for clozapine. This simple method was applied to determine the plasma concentration profiles of clozapine, N-desmethylclozapine and clozapine N-oxide in dog following administration of a 10 mg/kg oral dose of clozapine.
Keywords: Clozapine; N-Desmethylclozapine; Clozapine N-oxide;
Determination of urinary 12(S)-hydroxyeicosatetraenoic acid by liquid chromatography–tandem mass spectrometry with column-switching technique: sex difference in healthy volunteers and patients with diabetes mellitus by Naoto Suzuki; Takanori Hishinuma; Toshihide Saga; Jo Sato; Takayoshi Toyota; Junichi Goto; Michinao Mizugaki (383-389).
We developed a determination method for human urinary 12-hydroxyeicosatetraenoic acid (12-HETE) using LC–MS–MS. This method, which includes simple extraction and detection in the SRM mode, allows precise and accurate determination of 12-HETE. There was a significant sex difference in urinary 12-HETE levels. Chiral analysis of 12-HETE using LC–MS–MS with column-switching technique revealed that the major enantiomer was 12(S)-HETE. Furthermore, the urinary level in patients with diabetes mellitus (DM) was analyzed. The present in vivo findings indicate that there could be difference in production of 12(S)-HETE between genders and 12(S)-HETE may play a role in the pathogenesis of DM.
Keywords: 12(S)-Hydroxyeicosatetraenoic acid;
Liquid chromatographic–mass spectrometric method for the determination of α-,β-arteether in rat serum by M Rajanikanth; K.P Madhusudanan; R.C Gupta (391-399).
This study reports the development and validation of a sensitive and selective assay method for the determination of α-,β-arteether in rat serum by liquid chromatography–mass spectrometry. The mobile phase was composed of methanol–0.1 mM sodium acetate (pH 5) (80:20%) at a flow-rate of 1 ml min−1 and chromatographic separations were achieved on a Ultracarb, 5 ODS 20, Phenomenex column (5 μm, 30 mm×4.6 mm I.D.). The total effluent from the column was split so that one-tenth was injected into the electrospray LC–MS interface. ESI-MS analysis was carried out using a Micromass Quattro II Triple Quadrupole Mass Spectrometer equipped with an electrospray source. The MS analysis was carried out at a cone voltage of 52 V with a scan range of 100–400 Da. The analytes were quantified from the [M+Na]+ ion chromatograms of α-,β-arteether at m/z 335 and artemisinin at m/z 305. A simple liquid–liquid extraction with 2×2 ml n-hexane was used to isolate α-,β-arteether from rat serum. The method was validated in terms of recovery, linearity, accuracy and precision (within- and between-assay variation). The recovery from spiked control samples ranged from 88.41 to 96.17% with a maximum CV of 10.8% for α-arteether and 69.83–79.69% with a maximum CV of 17.06% for β-arteether. Linearity in serum was observed over the range 20–320 ng ml−1. Percent bias (accuracy) was well within the acceptable range. Within- and between-assay precision were less than 15%. The assay method described here is being applied to study the pharmacokinetics of CDRI developed intramuscular formulation Emal (α-/β-arteether in the ratio of 30:70) in rats. The method is sensitive enough to monitor α-,β-arteether up to 24 h after a single 30 mg kg−1 i.m. dose.
Development and characterization of an immunoaffinity column for the selective extraction of bisphenol A from serum samples by Meiping Zhao; Yang Liu; Yuanzong Li; Xinxiang Zhang; Wenbao Chang (401-410).
An immunoaffinity column (IAC) was developed by covalently coupling polyclonal antibodies against estrogenic bisphenols to CNBr-activated Sepharose 4B. The IAC showed high affinity for bisphenol A, while phenol was barely retained. Proteins in the sample matrix showed little nonspecific adsorption on the column. The best binding solvent for bisphenol A was found to be 0.01 mol l−1 phosphate-buffered saline (PBS) and the optimal operating temperature was 4 °C. The bound bisphenol A could be quantitatively recovered by 1 ml of methanol–water (80:20) with an average recovery of 91.8% and a relative standard deviation of 7.1% (n=6). The immunoaffinity column has been successfully used for the isolation and purification of bisphenol A from serum samples.
Keywords: Bisphenol A;
Simultaneous determination of six urinary porphyrins using liquid chromatography–tandem mass spectrometry by Wei Bu; Nichole Myers; Jane D McCarty; Tom O’Neill; Shelly Hollar; Philip L Stetson; Daniel W Sved (411-423).
A liquid chromatographic–tandem mass spectrometric (LC–MS–MS) method without sample pretreatment was developed and validated for determination of porphyrins in samples of canine urine. Acidified urine samples were directly injected into the LC–MS system and a gradient elution program was applied. The mass spectrometer was operated in the multi-reaction monitoring (MRM) mode and six porphyrins were detected with excellent sensitivity and selectivity. The lower limits of quantification were 0.014 nmol/mL for mesoporphyrin IX, coproporphyrin I, 5-carboxylporphyrin, 6-carboxylporphyrin and 7-carboxylporphyrin, and 0.029 nmol/mL for uroporphyrin I. Good ln-quadratic responses of calibration standards over the range 0.01 to 1.0 nmol/mL for mesoporphyrin IX, coproporphyrin I, 5-carboxylporphyrin, 6-carboxylporphyrin and 7-carboxylporphyrin, and 0.02 to 1.0 nmol/mL for uroporphyrin I were demonstrated. This method should be easily adapted through cross-validation for use in determining the effects of chemicals and pharmaceuticals on the urinary excretion profile of porphyrins in preclinical studies with other species, and in assisting the diagnosis of porphyria in clinical studies.
Development of a liquid chromatography–mass spectrometry method for monitoring the angiotensin-converting enzyme inhibitor lisinopril in serum by Andreas Tsakalof; Kyriaki Bairachtari; Manolis Georgarakis (425-432).
In this study, a sensitive, specific, precise and accurate method for lisonopril quantitative determination in human serum was developed and validated. The method comprises lisinopril isolation from serum by means of solid-phase extraction followed by its quantification by liquid chromatography–mass spectrometry. Chromatographic separation was performed at 55 °C on Kromasil C18 5 μm 250×3.2 mm HPLC column with mobile phase composed of 50 mM ammonium formate buffer (pH 3)–acetonirile–methanol (72:7:21, v/v/v). A Finnigan AQA benchtop mass spectrometer with a pneumatically assisted electrospray (ES) interface and a single quadrupole mass filter was used to detect and quantify lisinopril in column effluent. Ion signals were acquired by selected ion monitoring of the protonated lisinopril ion m/z=406.5 (M+1). The detector response was linear with r>0.9993 in the investigated concentration range 6–150 ng/ml. The mean recovery of lisinopril from serum samples was 88%. The limit of quantitation for lisinopril was 6 ng/ml with a signal-to-noise ratio at this concentration level S/N=34.75±3.9 (n=4).
Enantioselective determination of metoprolol acidic metabolite in plasma and urine using liquid chromatography chiral columns: applications to pharmacokinetics by Paula Macedo Cerqueira; Vanessa Bergamin Boralli; Eduardo Barbosa Coelho; Norberto Peporine Lopes; Luis Fernando Lopes Guimarães; Pierina Sueli Bonato; Vera Lucia Lanchote (433-441).
Enantioselective separations on chiral stationary phases with or without derivatization were developed and compared for the HPLC analysis of (+)-(R)- and (−)-(S)-metoprolol acidic metabolite in human plasma and urine. The enantiomers were analysed in plasma and urine without derivatization on a Chiralcel OD-R column, and in urine after derivatization using methanol in acidic medium on a Chiralcel OD-H column. The quantitation limits were 17 ng of each enantiomer/ml plasma and 0.5 μg of each enantiomer/ml urine using both methods. The confident limits show that the methods are compatible with pharmacokinetic investigations of the enantioselective metabolism of metoprolol. The methods were employed in a metabolism study of racemic metoprolol administered to a patient phenotyped as an extensive metabolizer of debrisoquine. The enantiomeric ratio (+)-(R)/(−)-(S)-acid metabolite was 1.1 for plasma and 1.2 for urine. Clearances were 0.41 and 0.25 l/h/kg, respectively, for the (+)-(R)- and (−)-(S)-enantiomers. The correlation coefficients between the urine concentrations of the acid metabolite enantiomers obtained by the two methods were >0.99. The two methods demonstrated interchangeable application to pharmacokinetics.
Gas chromatographic–mass spectrometric detection of 2- and 3-hydroxy fatty acids as methyl esters from soil, sediment and biofilm by M.M. Keinänen; L.K. Korhonen; P.J. Martikainen; T. Vartiainen; I.T. Miettinen; M.J. Lehtola; K. Nenonen; H. Pajunen; M.H. Kontro (443-451).
Hydroxy fatty acids (OH-FAs) can be used in the characterization of microbial communities, especially Gram-negative bacteria. We prepared methyl esters of 2- and 3-OH-FAs from the lipid extraction residue of soil, sediment, and biofilm samples without further purification or derivatization of hydroxyl groups. OH-FA methyl esters were analyzed using a gas chromatograph equipped with a mass selective detector (GC–MS). The ions followed in MS were m/z 103 for 3-OH-FAs and m/z 90 and M-59 for 2-OH-FAs. The rapid determination of 3- and 2-OH-FAs concomitantly with phospholipid fatty acids provided more detailed information on the microbial communities present in soil, sediment, and drinking water biofilm.
Keywords: Hydroxy fatty acid methyl esters; PLFA;
Lansoprazole quantification in human plasma by liquid chromatography–electrospray tandem mass spectrometry by Celso H Oliveira; Rafael E Barrientos-Astigarraga; Eduardo Abib; Gustavo D Mendes; Débora R da Silva; Gilberto de Nucci (453-459).
An analytical method based on liquid chromatography with positive ion electrospray ionization (ESI) coupled to tandem mass spectrometry detection was developed for the determination of lansoprazole in human plasma using omeprazole as the internal standard. The analyte and internal standard were extracted from the plasma samples by liquid–liquid extraction using diethyl-ether–dichloromethane (70:30; v/v) and chromatographed on a C18 analytical column. The mobile phase consisted of acetonitrile–water (90:10; v/v)+10 mM formic acid. The method has a chromatographic total run time of 5 min and was linear within the range 2.5–2000 ng/ml. Detection was carried out on a Micromass triple quadrupole tandem mass spectrometer by Multiple Reaction Monitoring (MRM). The intra- and inter-run precision, calculated from quality control (QC) samples, was less than 3.4%. The accuracy as determined from QC samples was less than 9%. The method herein described was employed in a bioequivalence study of two capsule formulations of lansoprazole.
Validated method for the determination of risperidone and 9-hydroxyrisperidone in human plasma by liquid chromatography–tandem mass spectrometry by B.M.M Remmerie; L.L.A Sips; R de Vries; J de Jong; A.M Schothuis; E.W.J Hooijschuur; N.C van de Merbel (461-472).
Since the first entry of risperidone on to the market in the early 1990s, investigation of the pharmacokinetic behaviour of the compound for which the availability of a bioanalytical method was a conditio sine qua non, has received considerable attention. Most of the published methods, however, did not reach the level of sensitivity and selectivity which can be obtained today since the evolution of liquid chromatography–tandem mass spectrometry (LC–MS–MS) towards a routine technique in the bioanalytical laboratory. Therefore, we developed and validated a new LC–MS–MS method for the determination of risperidone and its active metabolite 9-hydroxyrisperidone in human plasma. This paper describes in detail the bioanalytical procedure and summarizes the validation results obtained. In addition, it focuses on the pitfalls one might encounter when developing similar assays. Despite the particular physicochemical characteristics of risperidone and 9-hydroxyrisperidone, the LC–MS–MS method enabled the quantification of both compounds down to 0.1 ng/ml. The method uses a sample preparation step by solid-phase extraction at pH 6 using a mixed-mode phase. In a short chromatographic run, separation of 9-hydroxyrisperidone from the minor metabolite 7-hydroxyrisperidone is achieved. Detection takes place by (turbo)ionspray tandem mass spectrometry in the positive ion mode. The validated concentration range is from 0.100 to 250 ng/ml, using 500 μl of sample, with accuracy (bias) and precision (coefficient of variation) being below 15%. Although new developments in equipment will allow us to further improve and speed up the method, the assay reported can be used as a routine method to support a wide range of pharmacokinetic studies.
Keywords: Risperidone; 9-Hydroxyrisperidone;
High-performance liquid chromatographic method for determination of amodiaquine, chloroquine and their monodesethyl metabolites in biological samples by O.M.S Minzi; M Rais; J.O Svensson; L.L Gustafsson; Ö Ericsson (473-480).
A high-performance liquid chromatographic method for determination of amodiaquine (AQ), desethylamodiaquine (DAQ), chloroquine (CQ) and desethylchloroquine (DCQ) in human whole blood, plasma and urine is reported. 4-(4-Dimethylamino-1-methylbutylamino)-7-chloroquinoline was used as internal standard. The drugs and the internal standard were extracted into di-isopropyl ether as bases and then re-extracted into an acidic aqueous phase with 0.1 M phosphate buffer at pH 4.0 for AQ samples and at pH 2.5 for CQ filter paper samples. A C18 column was used and the mobile phase consisted of methanol–phosphate buffer (0.1 M, pH 3)–perchloric acid (250: 747.5:2.5, v/v). The absorbance of the drugs was monitored at 333 nm and no endogenous compound interfered at this wavelength. The limit of quantification in whole blood, plasma and urine was 100 nM for AQ and DAQ (sample size 100 μl) as well as for CQ and DCQ in blood samples dried on filter paper. For 1000 μl AQ and DAQ samples, the limit of quantification was 10 nM in all three biological fluids. The within-assay and between-assay coefficients of variations were always <10% at the limits of quantification. Plasma should be preferred for the determination of AQ and DAQ since use of whole blood may be associated with stability problems.
Keywords: Amodiaquine; Chloroquine; Desethylamodiaquine; Desethylchloroquine;
Direct stereoselective assay of fluoxetine and norfluoxetine enantiomers in human plasma or serum by two-dimensional gas–liquid chromatography with nitrogen–phosphorus selective detection by Sven Ulrich (481-490).
A method was developed and validated for the direct enantioselective assay of fluoxetine and norfluoxetine in human plasma or serum by two-dimensional capillary gas–liquid chromatography (GC). A Rtx-1 fused-silica capillary (15 m×0.25 mm I.D., 1.0 μm film thickness) and a hydrodex-β-6-TBDM fused-silica capillary (25 m×0.25 mm I.D., 0.25 μm film thickness) were used. A three-step liquid–liquid extraction was used for sample preparation with fluvoxamine and nisoxetine as internal standards. The method provided linear calibration between about 5 and 250 ng/ml for (R)- and (S)-fluoxetine as well as 15 and 250 ng/ml for (R)- and (S)-norfluoxetine. The limits of detection were about 1.5 and 6 ng/ml, respectively. Intra-day precision (coefficient of variation) was estimated as being between 5.4 and 12.7% at plasma levels of 25, 100 and 200 ng/ml for the four enantiomers. Inter-day precision was between 5.3 and 9.1% at 100 ng/ml. The enantioselective separation of some racemic psychopharmaceuticals was tested with various cyclodextrin GC-capillaries. Advantages and disadvantages of direct enantioselective GC are discussed for the assay of racemic psychopharmaceuticals. Samples from a patient who was treated with racemic fluoxetine were measured. In agreement with literature, plasma levels of the (R)-enantiomers of fluoxetine and norfluoxetine were considerably decreased in comparison to the (S)-enantiomers.
Keywords: Fluoxetine; Norfluoxetine;
Simultaneous quantitative determination of the HIV protease inhibitors indinavir, amprenavir, ritonavir, lopinavir, saquinavir, nelfinavir and the nelfinavir active metabolite M8 in plasma by liquid chromatography by U.S Justesen; C Pedersen; N.A Klitgaard (491-500).
A simple HPLC method that quantitates all six currently available protease inhibitors and the nelfinavir active metabolite M8 in one assay is presented. A 500-μl plasma sample was treated by liquid–liquid extraction with a mixture of heptane and ethyl acetate. After evaporation, the residue was redissolved in sodium dihydrogenphosphate and acetonitrile and washed twice with heptane. Chromatography was performed with an analytical C18 column. Ultraviolet detection at 210 and 239 nm was used. The present method is associated with high accuracy and low imprecision in the concentration range of 25–5000 ng/ml of all six protease inhibitors and M8. This makes it suitable for monitoring purposes.
Keywords: Indinavir; Amprenavir; Ritonavir; Lopinavir; Saquinavir; Nelfinavir;
Detection of bacterial pathogen DNA using an integrated complementary metal oxide semiconductor microchip system with capillary array electrophoresis by Joon Myong Song; Joel Mobley; Tuan Vo-Dinh (501-508).
In this paper, we show an integrated complementary metal oxide semiconductor (CMOS)-based microchip system with capillary array electrophoresis (CAE) for the detection of bacterial pathogen amplified by polymerase chain reaction (PCR). In order to demonstrate the efficacy of PCR reaction for the heat-labile toxin producing enterotoxigenic Escherichia coli (E. coli), which causes cholera-like diarrhea, 100 bp DNA ladders were injected along with the PCR product. Poly(vinylpyrrolidone) (PVP) was used as the separation medium and provided separation resolution which was adequate for the identification of PCR product. The miniaturized integrated CMOS microchip system with CAE has excellent advantages over conventional instrumental systems for analysis of bacterial pathogens such as compactness, low cost, high speed, and multiplex capability. Furthermore, the miniaturized integrated CMOS microchip system should be compatible with a variety of microfabricated devices that aim at more rapid and high-throughput analysis.
Keywords: DNA; Poly(vinylpyrrolidone);
Determination of midecamycin by capillary zone electrophoresis with electrochemical detection by Daxing Liu; Wenrui Jin (509-515).
Capillary zone electrophoresis was employed for the determination of midecamycin using an end-column amperometric detection with a carbon fiber micro-disk bundle electrode at a constant potential of +1.15 V vs. saturated calomel electrode. The optimum conditions of separation and detection are 1.00×10−3 mol l−1 Na2HPO4–3.49×10−4 mol l−1 NaOH (pH 11.4) for the buffer solution, 20 kV for the separation voltage, 5 kV and 5 s for the injection voltage and the injection time, respectively. The limit of detection is 5.0×10−7 mol l−1 or 0.41 fmol (S/N=3). The linear range of the calibration curve is 1.00×10−6–1.00×10−3 mol l−1. The relative standard deviation is 1.4% for the migration time and 4.9% for the electrophoretic peak current. The method could be applied to the determination of midecamycin in human urine. In this case, a separation voltage of 14 kV was used.
Gas chromatography–mass spectrometry profile of urinary organic acids of Wistar rats orally treated with ozonized unsaturated triglycerides and ozonized sunflower oil by Daniel Jardines; Teresa Correa; Oscar Ledea; Zullyt Zamora; Aristides Rosado; Jesús Molerio (517-525).
The main products in the ozonolysis of unsaturated triglycerides or vegetable oils are peroxides, aldehydes, Criegee ozonides and carboxylic acids. Some of these compounds are present in different concentrations in the biological fluids. The aim of this work is to study, using gas chromatography–mass spectrometry (GC–MS), the organic acid excretion in urine of rats orally treated with ozonized sunflower oil (OSO), ozonized triolein or ozonized trilinolein. Oral administration of OSO to Wistar rats has produced changes in the urinary content of dicarboxylic organic acids. Among others heptanedioic (pimelic acid) and nonanedioic acids (azelaic acid) were the major increased dicarboxylic acids found. The urinary dicarboxylic acid profiles of rats which received ozonized triolein only showed an increase in heptanedioic and nonanedioic acids. However, when ozonized trilinolein is applied, the profile is similar to that obtained when OSO is administered. A biochemical mechanism is proposed to explain the formation of dicarboxylic acids from ozonated unsaturated triglycerides.
Keywords: Dicarboxylic acids; Ozonized triglycerides;
Author Index to Vol. 783 (527-530).
Compound Index to Vol. 783 (531-535).