Study on Kinetic Characteristics of Alliinase

[Objective] The kinetic characteristics of alliinase was studied to select the optimum reaction performance. [Method] Alliinase activity was measured to analysis the influence of temperature, pH, substrate concentration and metal iron. [Result] Alliinase was an enzyme with thermal instability. Its optimum reaction temperature was 29℃ and pH value was 6.1. The Vmax was 0. 439 IU/mg and Km was 0.483 mmol/L by using natural extract as substrate. Alliinase activity was activated when the K^＋ , Mg^2＋ , Na^＋ and Cd^2＋ existed and alliinase activity was inhibited when Cu^2＋ existed. [Condusion] Results showed that the kinetic characteristics of alliinase supply the academic foundation for development and application of garlic medical products.

Pharmacokinetics of Recombinant E. coli L-asparaginase in Rats

The distribution of ^(125)I recombinant E. coli L-asparaginase in tissues ororgans and the excretion in urine, feces and bile were studied with in vivo radioactive tracertechnique. The amount of radioactivity excreted in urine, feces and bile within 24 h afterintravenous administration of ^(125)I recombinant E. col L-asparaginase to rats was 68.95% ,4.44%and 5.36% of the dose respectively. ^(125)I recombinant E. coli L-asparaginase in plasma samples wasdetermined. The levels of structural intact molecule in plasma samples were evaluated by SDS-PAGEand bio-imaging analyzer system. Pharmacokinetic parameters were assessed with a model-dependentmethod. The concentration-time curves of recombinant E. coli L-asparaginase after intravenousinjection at 1 250 IU·kg^(-1), 2 500, IU·kg^(-1), 5 000 IU·kg^(-1) to rats were consistent withthe two-compartment model. The first and terminal elimination t_(1/2) were 0.52 ~ 0.63 h and 2.39 ~2.76 h respectively. AUC was linearly related to the doses. The results of distribution in tissuesor organs and excretion in urine suggested that the metabolites of the enzyme were cleared bymechanisms of urinary excretion. Pharmacokinetics parameters of recombinant E. coli L- asparaginasein rats are warranted for the design of future clinical trials.

Pharmacokinetics and Bioavailability of 2-Amino-6-Cyclopropylamino-9-(2,3 -Dideoxy-β-D-glyceropent-2-enofuranosyl) Purine (Cyclo-D4G) in Rats

AimTo characterize the pharmacokinetics of 2 -amino-6-cyclopropylamino-9-(2,3-dideoxy-β-D-glyceropent-2-enofuran osyl) purine (Cyclo-D4G) following intravenous administration and oral administ ration to rats. MethodsThe concentrations of Cyclo-D4G in rat (Sprague-Dawley male rats) plasma and urine were analyzed by high performance liquid chromatography (HPLC). ResultsFollowing intravenous adm inistration to rats, concentrations of Cyclo-D4G in plasma declined with a term inal phase half-life of 0 78±0 14 h (±s). Total clearance was 0 90±0 21 L·h -1 ·kg -1 . Renal excretion of unchanged Cyclo-D4G accounted for approximately 20% of total clearance. Steady state volume of distr ibution was 0 91±0 07 L·kg -1 . After oral administration to rats, conce ntrations of Cyclo-D4G in plasma declined with a terminal phase half-life of 0 83±0 13 h (±s). Total clearance was 3 81±2 03 L·h -1 ·kg -1 . Renal excretion of unchanged Cyclo-D4G accounted for approximat ely 9% of total clearance. Oral bioavailability of Cyclo-D4G in rat was 26 9%. ConclusionThe favorable pharmacokinetic profiles and lower to xicity provide support for further development of Cyclo-D4G clinical trials.

Acute Toxicity Test of Qiangli Bupi Paste

[Objective] The toxicity of Qiangli Bupi Paste was investigated, to provide a test basis for its further development and safe use. [Method] Forty Kunming mice, half male and half female, were selected and divided into the CK group and the Qiangli Bupi Paste group, each of which included 20 mice. The acute toxicity of Qiangli Bupi Paste was observed by the maximum administration dosage method. No mice and abnormal response were observed within 14 d in various groups. The tested animals were also subjected to anatomical observation. [Result] All the tested animals survived in the test, and behaved normally, with glossy hair. Their body weights accorded with the growth regularity. During the anatomy, no important vis- ceral organs showed pathological changes, and there were no significant differences in body weight between the two groups before administration, on the 7~ after ad- ministration and the 14 d after administration （P〉0.05）. [Conclusion] In the acute toxicity test of Qiangli Bupi Paste, the maximum administration dosage was 48 g/kg （equivalent to 96 times of the daily dose of adult in clinic）, and no obvious toxic response was observed.

Pharmacokinetics and Biodegradation of Chitosan in Rats

Chitosan, an excellent biomedical material, has received a widespread in vivo application. In contrast, its metabolism and distribution once being implanted were less documented. In this study, the pharmacokinetics and biodegradation of fluorescein isothiocyanate(FITC) labeled and muscle implantation administrated chitosan in rats were investigated with fluorescence spectrophotometry, histological assay and gel chromatography. After implantation, chitosan was degraded gradually during its distribution to diverse organs. Among the tested organs, liver and kidney were found to be the first two highest in chitosan content, which was followed by heart, brain and spleen. Urinary excretion was believed to be the major pathway of chitosan elimination, yet 80% of chitosan administered to rats was not trackable in their urine. This indicated that the majority of chitosan was degraded in tissues. In average, the molecular weight of the degradation products of chitosan in diverse organs and urine was found to be <65 k Da. This further confirmed the in vivo degradation of chitosan. Our findings provided new evidences for the intensive and safe application of chitosan as a biomedical material.

A high performance liquid chromatography method for the quantitative determination assay of sitagliptin in rat plasma and its application in pharmacokinetics study

A new high-performance liquid chromatography （HPLC） method for the quantitative determination of sitagliptin in rat plasma was developed and validated for pharmacokinetics study. The plasma was spiked with the internal standard （hydrocortisone, IS）, treated with sodium hydroxide, and extracted with ethyl acetate. The extracted analyte was injected into an Agilent Zorbax Extend-C18 column （250 mm×4.6 mm, 4 μm） maintained at 30℃ and monitored at 267 nm. The mobile phase consisting of methanol-water （60：40, v/v, containing 10 mM Tris and 10 mM triethylamine） was titrated to pH 9.0 using 1 mol/L hydrochloric acid. The flow rate was 1.0 mL/min. The method showed high specificity. Calibration curves of the peak area ratio of each analyte/IS versus sitagliptin concentration were linear in the range of 0.75-100.0μg/mL （r〉0.9957）. The lower limit of quantification （LLOQ） was 0.75 μg/mL. The intra-day and inter-day coefficient of variation was lower than 10%. The accuracy （relative recovery） at three levels was 105.3% （0.75 μg/mL）, 99.8% （10.0 μg/mL） and 99.0% （100.0 μg/mL）. The extraction recovery was 81.5%, 82.4% and 84.5% at the concentrations of 0.75, 10.0 and 100.0 μg/mL, respectively. The short-term, long-term, freeze-thaw storage stability of plasma samples and the stability of standard solutions were satisfactory. This HPLC method is suitable for determining the concentration of sitagliptin in rat plasma and it was applied to determine the concentration-time profiles of sitagliptin in rat plasma following oral administration of sitagliptin.

Determination of a novel derivative of the PAC-1 anticancer agent in rat plasma by LC-MS/MS and its application to a pharmacokinetics study

A new and sensitive liquid chromatography-tandem mass spectrometry method was developed for the determination of SM-1 in rat plasma. After a simple protein precipitation, SM-1 and internal standard （gefitinib） were separated with gradient elution on a Waters XBridge C18 （50 mm×4.6 mm, 3.5μm） using acetonitrile-methanol-10 mM ammonium acetate （37.5：37.5：25, v/v/v） as mobile phase. The triple quadruple mass spectrometer was set in positive electrospray ionization mode, multiple reaction monitoring was used for quantification. The precursors to produce ion transitions monitored for SM-1 and IS were m/z 407.3→203.4 and 447.3→128.3, respectively. The method validation was conducted over the curve range of 30-6000 ng/mL. The intra- and inter-day precisions were less than 4.7%, the average extraction recoveries ranged from 98.7% to 104.1% for each analyte. SM-1 was proved to be stable during sample storage preparation and analytical procedures. All the results met the acceptance criteria in accordance with the FDA guidance for bioanalytical method. Consequently, this method was successfully applied to determine SM-1 concentrations in rats after oral administrations at the doses of 200, 100 and even 50 mg/kg.

本月妙侃

减肥记 狐猴家族中的大块头小胖近来很苦恼，因为学校里举行运动会，小胖也想参加，但由于它太胖，跑不动也跳不动，所以只能用羡慕的眼光看着同学们比赛。

Determination of metformin in diabetic rat plasma by an improved ion-pair high-performance liquid chromatography: application to a pharmacokinetic study

An efficient and sensitive ion-pair HPLC-UV method using atenolol as internal standard （IS） was developed and validated for the determination of metformin in the plasma of diabetic rats. Plasma samples were deproteinated with 10% （v/v） perchloric acid. Separation was achieved on a UltimateTM AQ-C18 column （250 mm×4.6 mm, 5 μm） with a mobile phase （pH 5.05） composed of acetonitrile-water （31：69, v/v, containing 0.002 M sodium dodecyl sulfate, 0.0125 M potassium dihydrogen phosphate, 0.015 M triethylamine） at a flow rate of 1.0 mL/min. The calibration curve was linear （r〉0.994） between 7.5 and 4000 ng/mL. The lower limit of quantification （LLOQ） was 7.5 ng/mL. The precision was validated and the relative standard deviation was in the range of 1.87% to 15.70%; the accuracy was between 93.98%-106.89%. The mean recoveries were 95.40% and 95.31% for metformin and IS, respectively. The relative error （RE） of stability at different storage conditions was within ±9.00%. This method was used to determine the concentration-time profile of metformin in diabetic rat plasma following an oral administration of metformin at the dose of 10 mg/kg. Our results indicated that ion-pair HPLC-UV method using UltimateTM AQ-C18 column was effective for the pharmacokinetic studies of high polarity compounds like metformin.

Distribution and pharmacokinetics of five Rhubarb anthraquinones in rabbits and rats

The main active components of Rhubarb are anthraquinones（AQs）, most of which are glycosides and others are free. The concentrations of AQs derivatives（rhein, aloe-emodin, emodin, chrysophanol and physcion） in plasma and homogenate were assayed with a high performance liquid chromatography（HPLC） method. The pharmacokinetic parameters and distribution of Rhubarb AQs in rabbits or rats were studied after administration of different formulas. Elimination of AQs was fit to a two-compartment model in rats and rabbits. There were no significant difference in the main pharmacokinetic parameters between rhein and AQs in rats. AQs were distributed progressively in the kidney, liver, blood, and heart. The AQs were mainly composed of rhein in vivo and was excreted by the kidney. For formulas that contained Rhubarb, rhein could be used as a probe for in vivo pharmacokinetic studies.

Simultaneous determination of two bioactive components of Huangqi Guizhi Wuwu Decoction in rat plasma using UPLC-MS/MS and its application to a pharmacokinetic study

A rapid, sensitive and selective UPLC-MS/MS method was developed to determine paeoniflorin and astragaloside IV, This method was validated via a pharmacokinetic study using rat plasma. The internal standard was clarithromycin. A simple one-step deproteinization procedure was used to prepare plasma samples. Separation was achieved on a CAPCELL CORE ADME CI8 column with a gradient mobile phase consisting of solution A （water containing 0.1% formic acid） and solution B （acetonitrile） at a flow rate of 0.3 mL/min. Multiple reaction monitoring （MRM） was used with an electrospray ionization source （ESI） in positive mode. A good linear response was observed within the ranges of 0.01 to 5.00 ~g/mL for paeoniflorin and 0.000l to 0.05 ~tg/mL for astragaloside IV. The accuracy （RE） was within the range of-3.5% to 6.3%, and the intra- and inter-day precisions （RSD） were within 14.2%. The extraction recoveries were all above 78.9%. The pharmacokinetic study of the two analytes in rats after oral administration of Huangqi Guizhi Wuwu Decoction （HGWD） was successfully completed through this method. The method develooed in this studv will fill a gap in oharmacokinetic studies of HGWD.

Pharmacokinetic investigation on nteraction between hydrophilic lithospermic acid B and lipophilic tanshinone IIA in rats:an experimental study

OBJECTIVE:To elucidate the interaction between hydrophilic lithospermic acid B and lipophilic tanshinone Ⅱ A in rats.METHODS:A reliable high-performance liquid chromatography method was adopted for simultaneous determination of lithospermic acid B and tanshinone Ⅱ A in rat plasma,through which the pharmacokinetic interaction between lithospermic acid B and tanshinone Ⅱ A by intravenous injection was investigated.RESULTS:The simultaneous intravenous injection of tanshinone Ⅱ A and lithospermic acid B significantly altered the pharmacokinetic parameters of both compounds when compared with the individual intravenous administration of each compound.The area under the concentration-time curve of tanshinone Ⅱ A and lithospermic acid B increased by 18.35 and 59.31%,respectively.The mean retention time of tanshinone Ⅱ A and lithospermic acid B increased,respectively,from 9.3 to 32.8 h and20.2 to 49.1 h.The concomitant use of tanshinoneⅡ A magnified the volume of distribution at steady state(V_(ss)) and time for the drug in the plasma to reduce the highest concentration by half(t_(1/2)) of lithospermic acid B,while at the same time the V_(ss) and t_(1/2)of tanshinone Ⅱ A changed significantly in the presence of lithospermic acid B.CONCLUSION:Lithospermic acid B and tanshinone D A interact with each other following simultaneous intravenous injection in rats and this observation may expand the clinical use of Danshen(Radix Salviae Miltiorrhizae).