高效液相色谱法测定吲哚美辛控释胶囊的血药浓度和生物利用度

采用ＯＤＳ柱,甲醇－稀磷酸溶液（７６２４）为流动相,２６０ｎｍ为检测波长,建立了测定血浆中吲哚美辛浓度的高效液相色谱法,并测定了吲哚美辛控释胶囊炎痛康的血药浓度。结果表明,血浆中吲哚美辛浓度在０．１２５～５．０ｍｇ／Ｌ范围内线性关系良好（ｒ＝０．９９９６）,检测限６２．５μｇ／Ｌ（Ｓ／Ｎ＝３１）,平均回收率为１００．４％,日内和日间ＲＳＤ均小于５％。１１位受试者单剂量口服炎痛康后的相对生物利用度为１０２．３８％。     

气相色谱-质谱法测定氯氮平及其去甲基代谢物

建立了测定人血清中氯氮平及其去甲基代谢物的柱前衍生化气相色谱－质谱选择离子监测的分析方法。以三氟乙酸酐作酰化剂,对衍生化条件和样品预处理方法实施了优化。氯氮平和去甲氯氮平的线性范围为１～１２８μｇ／Ｌ,最低检测浓度：氯氮平为０．１μｇ／Ｌ,去甲氯氮平为０．２μｇ／Ｌ,两者的回收率均大于８３％,相对标准偏差都小于１０％。将所建立的方法应用于服用细胞色素氧化酶Ｐ４５０１Ａ２抑制剂前后低剂量氯氮平的药代动力学自身对照试验中,结果显示氯氮平的代谢水平明显受Ｐ４５０１Ａ２活性的影响。     

Measurement of penbutolol and 4-hydroxypenbutolol in plasma or serum by HPLC

A simple HPLC method for penbutolol and 4-hydroxypenbutolol assay has been developed. Plasma or serum (200 microliters) is vortex-mixed (30 s) with Tris solution (2 M, pH 10.6) containing an internal standard (50 microliters) and methyl t-butyl ether (200 microliters). After centrifugation, the extract (100 microliters) is analysed using an unmodified silica column (250 x 5 mm ID) and iso-octane-methanol-methyl t-butyl ether (55:25:20) containing ammonium perchlorate (10 mM, pH 5.7) as eluent and with fluorescence detection. No interference has been encountered and the limit of accurate measurement for both compounds is 5 micrograms/l.     

高效液相色谱法测定血清中头孢噻肟浓度

用固相萃取法处理样品,以扑热息痛为内标物、甲醇－醋酸钠／醋酸缓冲溶液作流动相,采用反相高效液相色谱法测定血清中头孢噻肟的浓度。头孢噻肟和内标物的平均回收率分别为９６．７％和９７．７％,头孢噻肟血清浓度在１０ｍｇ／Ｌ至１５０ｍｇ／Ｌ范围内有良好线性关系,最低检测浓度为２ｍｇ／Ｌ,日内和日间相对标准偏差分别在３．０％和４．１％以内。结果表明方法准确、简便。     

Determination of cholesterol and cortisone absorption in polyurethane. I. Methodology using size-exclusion chromatography and dual detection.

A size-exclusion chromatographic method is described for measuring the absorption of the steroid-based lipids cholesterol and cortisone into Pellethane 2363, a polyurethane used in biomedical implants. The method uses refractometry and ultraviolet diode-array detection, with tetrahydrofuran as the mobile phase. Using an injection volume of 150 microliters, the lower limit of accurate measurement for cholesterol (refractive index detection) was 6 micrograms/ml with a lower limit of detection, based on a 2:1 signal-to-noise ratio, of 0.15 micrograms (1 microgram/ml). For cortisone (ultraviolet detection), the lower accurate limit was 0.6 micrograms/ml with a lower limit of 0.015 micrograms (0.1 micrograms/ml). The results show that after 44 h, 2037 micrograms/g cholesterol and 3131 micrograms/g cortisone were absorbed by the polyurethane. The method eliminates extensive sample manipulation and is sensitive to low levels of lipid in the presence of a high-molecular-mass synthetic polymer.     

手性荧光衍生化反相高效液相法分离肾上腺素对映体

 以R (- ) /S (+) 4 (N ,N dimethylaminosulfonyl) 7 (3 iso thiocyanatopyrrolidino) 2 ,1,3 benzoxadiazole(DBD PyNCS)为手性荧光衍生化试剂 ,用反相高效液相法 (RP HPLC)对 DL 肾上腺素对映体进行分离。衍生化反应是在含有 7%吡啶的溶剂中 ,6 5℃温度条件下反应 35min ,生成具有荧光特性的肾上腺素非对映体衍生物(λex=4 6 0nm ,λem=5 5 0nm)。生成的非对映体衍生物在流动相为乙腈 水 (体积比为 2 8∶72 ) ,流速为 1 0mL/min时 ,在DiamonsilTMC18柱 (15 0mm× 4 6mmi d ,5 μm)上分离度可达 2 6。     

安培检测-离子色谱法测定乳品中的微量碘

 用安培检测 离子色谱法检测了乳品中的微量碘。以NaOH作淋洗液 ,直接进样 ,优化了碘的离子色谱分离和测定条件 ,方法的检测限达 1μg/L(3倍信噪比 )。测定了婴儿奶粉、孕产妇专用奶粉和鲜牛奶中的微量碘 ,加标回收率分别为 89 0 % ,86 0 %和 84 1%。     

Evaluation of 384-well formatted sample preparation technologies for regulated bioanalysis

The capabilities and limitations of 384-well formatted sample preparation technologies applied to regulated bioanalysis were evaluated by developing two assays for the simultaneous quantitation of lopinavir and ritonavir, the active ingredients of Kaletra. One method used liquid-liquid extraction (LLE), and the other used solid-phase extraction (SPE). The steps and apparatuses employed by the two methods covered most of those used for bioanalysis. Briefly, the previously validated 96-well formatted assays were adapted to the 384-format with minor modifications. Because the wells of a 384-well plate are clustered together, cross-contamination between adjacent wells was evaluated critically, along with sensitivity, assay throughput, and ruggedness. Samples (35 microL) containing plasma samples (15 microL), internal standard (10 microL), and sodium carbonate (0.5 M, 10 microL to basify the sample) were placed in a 384-well microtiter plate that may contain saquinavir or amprenavir as contamination markers. For LLE preparation, the samples were placed in a deep 384-well plate (300-microL well volume) and extracted with 150 microL of ethyl acetate. Approximately 50 microL of the extracts were removed from each well after phase separation for analysis. For SPE preparation, the fortified samples were transferred to a 384-formatted SPE plate (C18, 5 mg packing). The extracts were eluted from the plate with basified 2-propanol. The LLE or SPE extracts were dried and reconstituted for column-switching high-performance liquid chromatography with tandem mass spectrometric detection (HPLC/MS/MS). The lower limit of quantitation and the assay range were the same as the 96-well formatted assay. If combined with appropriate automation, sample preparation in the 384-well format would be up to five times more efficient than the 96-well format.     

Plasma determination of the novel anticonvulsant D,L-3-hydroxy-3-ethyl-3-phenylpropionamide and preliminary pharmacokinetic studies in the rat.

A sensitive gas chromatographic method with flame ionization detection was developed for the analysis in plasma of the novel anticonvulsant D,L-3-hydroxy-3-ethyl-3-phenylpropionamide (HEPP), using D,L-2-hydroxy-2-ethyl-2-phenylacetamide as the internal standard. HEPP was extracted from alkalinized plasma into dichloromethane and quantified after derivatization with bis(trimethylsilyl)-trifluoroacetamide: Standard curves were linear from 0.5 to 50 and from 2 to 100 micrograms/ml of plasma, using 1.5 and 5 micrograms of the internal standard, respectively. The lower limit of detection at a signal-to-noise ratio of 3 standard deviations was 0.33 micrograms/ml of sample. The sensitivity, accuracy and reproducibility of the method were shown to be satisfactory for pharmacokinetic studies of HEPP. After intraperitoneal administration of 50 mg/kg to Wistar rats, the principal kinetic parameters were: absorption half-life = 0.04 h; volume of distribution = 1.32 l/kg; clearance = 4.40 ml/min; peak concentration = 50 micrograms/ml; peak time = 0.25 h; mean residence time = 4.55 h.     

气相色谱/质谱法测定大鼠脑中5-羟色胺的含量

采用气相色谱／质谱法（ＧＣ／ＭＳ）测定了正常大鼠和服药大鼠脑组织中５－羟色胺（５－ＨＴ）的浓度。大鼠脑组织制成匀浆后,先将５－ＨＴ酰化,酰化物经乙酸乙酯提取后,再经七氟丁酸酐衍生化,用ＧＣ／ＭＳ测定。ＭＳ采用电子捕获负离子化学电离（ＥＣＮＩＣＩ）模式。方法的线性范围为０．５０～５０．０μｇ／Ｌ,回归方程为Ｙ＝０．１３４８Ｘ－０．０７９９５（ｒ＝０．９９９６）;平均回收率为９８．２％±３．８％（ｎ＝１０）;检测限为０．５μｇ／Ｌ;相对标准偏差小于１０％。     

Determination of chlorophenols in water samples using simultaneous dispersive liquid-liquid microextraction and derivatization followed by gas chromatography-electron-capture detection

Simultaneous dispersive liquid-liquid microextraction (DLLME) and derivatization combined with gas chromatography-electron-capture detection (GC-ECD) was used to determine chlorophenols (CPs) in water sample. In this derivatization/extraction method, 500 microL acetone (disperser solvent) containing 10.0 microL chlorobenzene (extraction solvent) and 50 microL acetic anhydride (derivatization reagent) was rapidly injected by syringe in 5.00 mL aqueous sample containing CPs (analytes) and K(2)CO(3) (0.5%, w/v). Within a few seconds the analytes derivatized and extracted at the same time. After centrifugation, 0.50 microL of sedimented phase containing enriched analytes was determined by GC-ECD. Some effective parameters on derivatization and extraction, such as extraction and disperser solvent type and their volume, amount of derivatization reagent, derivatization and extraction time, salt addition and amount of K(2)CO(3) were studied and optimized. Under the optimum conditions, enrichment factors and recoveries are in the range of 287-906 and 28.7-90.6%, respectively. The calibration graphs are linear in the range of 0.02-400 microg L(-1) and limit of detections (LODs) are in the range of 0.010-2.0 microg L(-1). The relative standard deviations (RSDs, for 200 microg L(-1) of MCPs, 100 microg L(-1) of DCPs, 4.00 microg L(-1) of TCPs, 2.00 microg L(-1) of TeCPs and PCP in water) with and without using internal standard are in the range of 0.6-4.7% (n=7) and 1.7-7.1% (n=7), respectively. The relative recoveries of well, tap and river water samples which have been spiked with different levels of CPs are 91.6-104.7, 80.8-117.9 and 83.3-101.3%, respectively. The obtained results show that simultaneous DLLME and derivatization combined with GC-ECD is a fast simple method for the determination of CPs in water samples.     

Determination of hexazinone in groundwater by direct-injection high-performance liquid chromatography

Hexazinone has been detected at levels ranging from 0.2 to 50 micrograms/L in many groundwater samples from eastern Maine over the past decade. A rapid and inexpensive direct-injection high-performance liquid chromatographic (HPLC) method has been developed to monitor contamination levels of the herbicide. The method is sensitive (limit of quantitation = 0.33 microgram/L) and is linear to 33.0 micrograms/L (R2 = 0.9995). Direct injection results from 50 field samples compared well (R2 = 0.98) with an HPLC method using solid-phase extraction for concentration and cleanup. The technique is very reproducible (coefficients of variation of 0-8.4% within day and 3.0-13.2% between day) and eliminates loss of analyte because of fewer steps in the procedure.     

Development and validation of HPLC method for the determination of α-tocopherol in human erythrocytes for clinical applications

In this work, a simple isocratic reversed-phase HPLC method for determination of alpha-tocopherol in human erythrocytes has been developed and validated. After separation of plasma the erythrocytes were washed three times with 0.9% sodium chloride containing 0.01% butylated hydroxytoluene (BHT) as antioxidant and then were diluted 1:1 (v/v) with the same solution. In the liquid-liquid extraction (LLE) procedure, 2500 microL of n-hexane was added to 500 microL of erythrocytes. After 2 min this mixture was deproteinized by addition of cool ethanol (500 microL, 5 min) denatured with 5% methanol containing alpha-tocopherol acetate (20 micromol L(-1)), as internal standard, and then extracted for 5 min by vortex mixing. After centrifugation (10 min, 1600xg) an aliquot (2000 microL) of the clean extract was separated and evaporated under nitrogen. The residue was dissolved in 400 microL methanol and analysed by reversed-phase HPLC on a 4.6 mmx150 mm, 5 microm Pecosphere C18 column; the mobile phase was 100% methanol, flow rate 1.2 mL min(-1). The volume injected was 100 microL and detection was by diode-array detector at a wavelength of 295 nm. The extraction recovery of alpha-tocopherol from human erythrocytes was 100.0+/-2.0%. The detection limit was 0.1 micromol L(-1) and a linear calibration plot was obtained in the concentration range 0.5-20.0 micromol L(-1). Within determination precision was 5.2% RSD (n=10), between determination precision was 6.1% RSD (n=10). The method was applied successfully in a clinical study of patients with acute pancreatitis and for determination of the reference values in the healthy Czech population.     

Quantitative analysis of betulinic acid in mouse,rat and dog plasma using electrospray liquid chromatography/mass spectrometry

Betulinic acid is under development as a therapeutic agent for the treatment of metastatic malignant melanoma. In support of pharmacokinetic and toxicological evaluations, a robust assay based on liquid chromatography/mass spectrometry (LC/MS) was developed for the quantitative analysis of betulinic acid. Sample preparation consisted of deproteinization of the plasma by the addition of three volumes of acetonitrile and one volume of methanol followed by centrifugation. Aliquots of the supernatant were analyzed using an isocratic reversed-phase high-performance liquid chromatography (HPLC) system coupled to a negative ion electrospray mass spectrometer. Deprotonated molecules of betulinic acid and the isomeric internal standard oleanolic acid were detected using selected ion monitoring at m/z 455. The limit of detection of betulinic acid was 0.5 pg (1.1 fM) injected on-column (50 pg/mL, 10 microL injection volume), and the limit of quantitation was 2 pg (4.4 fM, 200 pg/mL, 10 microL injection volume). Betulinic acid was stable in plasma samples at -20 degrees C for at least 3 weeks. The intra-day and inter-day coefficients of variation of the assay were < or =6.4 and < or =9.0%, respectively. The utility of the assay was demonstrated by analyzing betulinic acid spiked into mouse, rat and dog plasma, by determining the extent of binding of betulinic acid to plasma proteins, and by measuring betulinic acid in mouse and rat plasma following intraperitoneal or intravenous administration in vivo. At 15 and 25 microg/mL in mouse, rat or dog plasma, betulinic acid was 99.99% bound to serum proteins, and, at 5 microg/mL, betulinic acid was > or =99.97% bound.     

Insecticidal activity against Bemisia tabaci biotype B of peel essential oil of Citrus sinensis var. pear and Citrus aurantium cultivated in northeast Brazil

The fumigant action of peel essential oils of Citrus sinensis var. pear (pear orange = PO) and C. aurantium (bitter orange = BO) from the northeast of Brazil were evaluated against Bemisia tabaci biotype B and compared with eugenol as a positive control. The oil concentration in the PO at 8.5 microL/L of air caused 97% mortality, while the oil concentration of BO at 9.5 microL/L of air caused 99% mortality. However, the LC50 estimates for both oils (LC50 = 3.80 microL/L of air for PO and LC50 = 5.80 microL/L of air for BO) did not differ from each other, but they did when compared with eugenol (LC50 = 0.20 microL/L of air). Regarding their effects on oviposition, the Citrus oils showed concentration-response dependence, reducing the number of eggs as the concentration increased, which was not observed for eugenol. The minimum concentrations of the oils that caused a significant reduction in the egg lay were 3.5 and 7.0 microL/L of air for BO and PO, respectively. These results suggest that oils from PO and BO peels may be promising as models to develop new insecticides that might be applied into the integrated management of whiteflies.     

High-throughput LC-MS/MS assay for 6-methoxy-2-naphthylacetic acid, an active metabolite of nabumetone in human plasma and its application to bioequivalence study

A simple, precise and accurate assay for the determination of 6-methoxy-2-naphthylacetic acid (6-MNA), an active metabolite of nabumetone in human plasma, was developed and validated using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The analyte (6-MNA) and propranolol (internal standard, IS) were extracted from 200 microL aliquot of human plasma via solid-phase extraction employing HLB Oasis cartridges and separated on a Discovery HS C18 (50 x 4.6 mm, 5 microm) column. Detection of analyte and IS was done by tandem mass spectrometry with a turbo ion spray interface operating in positive ion and multiple reaction monitoring acquisition mode. The total chromatographic runtime was 3.0 min with retention time for 6-MNA and IS at 1.97 and 1.26 min, respectively. The method was validated over a dynamic linear range of 0.20-60.00 microg/mL for 6-MNA with mean correlation coefficient r > or = 0.9986. The intra-batch and inter-batch precision (%CV) across five validation runs (lower limit of quantiation, low-, medium- and high-quality controls and upper limit of quantitation) was less than 7.5%. The accuracy determined at these levels was within -5.8 to +0.2% in terms of percentage bias. The method was successfully applied for a bioequivalence study of 750 mg nabumetone tablet formulation in 12 healthy Indian male subjects under fasted condition.     

手性毛细管色谱法测定人尿中美芬妥英对映体的含量及其在代谢分型中的应用

 采用手性毛细管色谱柱和ＦＩＤ检测器建立了人尿中美芬妥英（ＭＰ）对映体的定量分析方法。尿样用二氯乙烷提取，用酸、碱洗涤得以纯化，测得各对映体的最低检测限为６０μｇ／Ｌ。在１１５～６９０μｇ／Ｌ浓度范围内，标准曲线呈良好的线性关系，ｒ＞０．９９，日内、日间精密度ＲＳＤ＜６．５％，Ｓ－ＭＰ的平均回收率为７４．４１％，Ｒ－ＭＰ的平均回收率为７３．７８％。并以ＭＰ为探针药物，对３２名志愿者的尿样进行了ＭＰ氧化代谢分型研究。     

Reference materials at emission levels of NH3 and HCl

Due to instability in static mixtures, in the past only dynamic preparation techniques for mixtures of NH3 and HCl were maintained; however, recent developments of new passivation techniques for cylinder treatment and commercially available nitrogen with a very low content of impurities have opened up the possibility of preparing stable mixtures in cylinders by gravimetry. Mixtures of NH3 (300 microL/L and 30 microL/L) and HCI (300 microL/L) were prepared in nitrogen in cylinders with different passivation treatments. The mixtures were compared with dynamically prepared mixtures using Photo Acoustic detection (PAS) and FT-IR. The uncertainty is about 3.5% with PAS analysis for NH3 at the 300 microL/L level. For the analysis of HCI with FT-IR the initial uncertainty in the 300 microL/L mixtures is about 7%. Two of the HCI mixtures show significant instability; for the other three mixtures the uncertainty in the measurement is too large to discriminate between stability and instability. The NH3/N2 mixtures are stable within 3.5% for a one-year period. The stability of the 30 microL/L mixtures of NH3 cannot be underpinned by measurement results, but a systematic effect in the measurement procedure seems more likely than instability. Improvement in the measurement procedure should clarify this point.     

Simple method for the determination of rosiglitazone in human plasma using a commercially available internal standard

To the best of our knowledge, bioanalytical methods to determine rosiglitazone in human plasma reported in literature use internal standards that are not commercially available. Our purpose was to develop a simple method for the determination of rosiglitazone in plasma employing a commercially available internal standard (IS). After the addition of celecoxib (IS), plasma (0.25 mL) samples were extracted into ethyl acetate. The residue after evaporation of the organic layer was dissolved in 750 microL of mobile phase and 50 microL was injected on to HPLC. The separation was achieved using a Hichrom KR 100, 250 x 4.6 mm C(18) with a mobile phase composition potassium dihydrogen phosphate buffer (0.01 m, pH 6.5):acetonitrile:methanol (40:50:10, v/v/v). The flow-rate of the mobile phase was set at 1 mL/min. The column eluate was monitored by fluorescence detector set at an excitation wavelength of 247 nm and emission wavelength of 367 nm. Linear relationships (r(2) > 0.99) were observed between the peak area ratio rosiglitazone to IS vs rosiglitazone concentrations across the concentration range 5-1000 ng/mL. The intra-run precision (%RSD) and accuracy (%Dev) in the measurement of rosiglitazone were <+/-10.69 and <-12.35%, respectively across the QC levels (50-1000 ng/mL). The extraction efficiency was >80% for both rosiglitazone and IS from human plasma. The lower limit of quantitation of the assay was 5 ng/mL. In summary, the methodology for rosiglitazone measurement in plasma was simple, sensitive and employed a commercially available IS.     

Determination of sulfamethazine in milk by biosensor immunoassay

A biosensor based on surface plasmon resonance (SPR) measurement was developed for use in an immunoassay for detection of sulfamethazine (SMZ) in milk. The biospecific surface was a carboxymethyl dextran-modified gold-surface sensor chip to which SMZ was covalently bound. The assay was based on inhibition of the binding of polyclonal antibodies to immobilized SMZ by SMZ in the sample. The SPR response changed inversely in relation to the antibiotic concentration in the sample. Calibration curves were constructed for SMZ in buffer and in milk at a concentration which included the maximum residue limit (0 to 200 micrograms/kg). The analysis time per sample varied from 8 to 30 min. Different flow rates and antibodies were modified alternatively during the study to assess their influence on the performance of the assay. The active antibody concentration was calculated at approximately 1880 and 180 nM for the antibody anti-SMZ 1 and the antibody anti-SMZ 2, respectively. No cross-reactivity of antibodies with other antibiotics was found. Under optimal conditions, the detection limits in milk for SMZ were 8 and 1.7 micrograms/kg, respectively, for antibody 1 and antibody 2, at a flow rate of 20 microL/min.