杏香兔耳风总酚酸耐药突变选择窗的体外研究

目的 测定杏香兔耳风总酚酸和对照药绿原酸对金黄色葡萄球菌ATCC26003和大肠埃希菌ATCC44102的防耐药变异浓度(MPC)。方法 采用琼脂平板二倍稀释法,测定杏香兔耳风总酚酸和绿原酸对金黄色葡萄球菌和大肠埃希菌的最小抑菌浓度(MIC)、99 %抑菌浓度(MIC99)、初测MPC(MPCpr)和MPC。结果 杏香兔耳风总酚酸、绿原酸对金黄色葡萄球菌的MIC、MPC和细菌耐药选择指数(MPC/MIC99)分别为0.4,6.0 mg·mL-1;10.8,8.4 mg·mL-1;34.0,1.6。对大肠埃希菌的MIC、MPC和MPC/MIC99分别为12.0,6.0 mg·mL-1;16.8,9.6 mg·mL-1;1.4, 1.7。结论 杏香兔耳风总酚酸限制金黄色葡萄球菌耐药突变菌株作用强于绿原酸,限制大肠埃希菌耐药突变菌株作用弱于绿原酸。     

桑柏生发方的体外抑菌作用研究

目的:探讨桑柏生发方的体外抑菌作用.方法:通过稀释法测定最低抑菌浓度(MIC)和最小杀菌浓度(MBC),菌落法绘制时间-杀菌曲线.结果:桑柏生发方水煎剂对标准株金黄色葡萄球菌、大肠埃希菌、铜绿假单胞菌,临床分离株表皮葡萄球菌和大肠埃希菌的MIC为62.5～250mg/ml,MBC为125～500mg/ml;桑柏生发方搽剂对5种细菌的MIC为3.91～125mg/ml,MBC为15.63～250mg/ml.杀菌曲线表明,桑柏生发方在24h内可完全杀死金黄色葡萄球菌、大肠埃希菌和铜绿假单胞菌.结论:桑柏生发方水煎剂和搽剂在体外对多种细菌具有较强的杀菌作用.     

新疆黑蜂胶对金黄色葡萄球菌生物膜形成的影响

目的研究新疆黑蜂胶提取物对金黄色葡萄球菌生物膜(bacterial biofilm,BF)形成的影响,为临床辅助治疗耐药性感染提供实验依据。方法用醇提法提取新疆黑蜂胶得到黑蜂胶提取物,采用试管二倍稀释法测定蜂胶的最低抑菌浓度(MIC),用微孔板改良法复制金黄色葡萄球菌BF模型,并检测新疆黑蜂胶不同MIC浓度对金黄色葡萄球菌BF形成的影响,同时通过检测蜂胶对成熟金黄色葡萄球菌BF中活菌数的影响评价其对成熟金黄色葡萄球菌BF的作用。结果新疆黑蜂胶提取物对金黄色葡萄球菌的MIC为6.25mg/ml,当浓度为3.125 mg/ml时新疆黑蜂胶就可干扰细菌BF的形成,致活菌数明显少于空白对照组;新疆黑蜂胶还可以抑制并破坏成熟金黄色葡萄球菌BF,高浓度蜂胶抗菌活性与青霉素相比无显著性差异。结论新疆黑蜂胶提取物在体外具有抑制金黄色葡萄球菌BF形成的作用,并能破坏已形成的BF,具备开发应用的潜力。     

五倍子乙醇提取物对金葡菌的体外抗菌研究

目的观察五倍子乙醇提取物对金黄色葡萄球菌(MRSA和MSSA)的体外抗菌活性.方法采用中药抑菌实验方法对五倍子乙醇提取物进行了112株金黄色葡萄球菌的最低抑菌浓度测定.结果五倍子乙醇提取物对84株耐甲氧西林的金黄色葡萄球菌(methicillin-resistant staphylococcus aureus,MRSA)和28株甲氧西林敏感的金黄色葡萄球菌(methicillin-sensitive staphylococcus aureus,MSSA)的MIC50、MIC90分别为0.315、0.315和0.63、0.315 mg/mL.结论五倍子乙醇提取物对金黄色葡萄球菌(84株MRSA和28株MSSA)具有较强的抑菌活性.     

“玉容消痤颗粒”体外抗菌作用研究

目的：研究玉容消痤颗粒体外对金黄色葡萄球菌、表皮葡萄球菌及痤疮丙酸杆菌（P.acne）的抗菌作用。方法：金黄色葡萄球菌和表皮葡萄球菌各取1个菌苔,接种在2 ml MH营养肉汤培养基中,培养6 h;P.acne取1个菌苔,接种在2 ml 硫乙醇酸钠培养基中,厌氧培养48 h;金黄色葡萄球菌和表皮葡萄球菌取10-5、10-6倍稀释度的菌液0.05 ml涂布平皿37℃培养24 h计活菌数,P.acne取10-5、10-6倍稀释度的菌液0.05 ml放入平皿与培养基混匀37℃厌氧培养48 h计活菌数。选30-300个菌落/皿的平板计算结果。以丹参酮为对照药分别测定玉容消痤颗粒和丹参酮胶囊对金黄色葡萄球菌、表皮葡萄球菌和P.acne最低抑菌浓度（MIC）和最低杀菌浓度（MBC）。结果：玉容消痤组,各菌株的5.63-90 mg·ml-1药液的试管未见长菌,其MIC为5.63 mg·ml-1,丹参酮胶囊组,各菌株的45-90 mg·ml-1药液的试管未见长菌。其MIC为45 mg·ml-1;玉容消痤对金黄色葡萄球菌、表皮葡萄球菌和P.acne的MBC均为11.25 mg·ml-1,丹参酮胶囊对金黄色葡萄球菌、表皮葡萄球菌和P.acne的MBC均为90 mg·ml-1。结论：玉容消痤颗粒抗菌作用肯定,最低抑菌浓度及最低杀菌浓度均明显低于丹参酮胶囊。     

立止血与双歧杆菌活菌联合治疗应激性溃疡出血疗效观察

目的:分析立止血与双歧杆菌活菌联合治疗应激性溃疡(SU)的临床疗效。方法:将我院住院的72例SU患者随机分为两组,对照组予以常规治疗;研究组,在常规治疗基础上予以立止血与双歧杆菌活菌联合治疗。两组均为36例,记录两组的呕血、黑便及止血的时间,检测胃粘膜pH值(pHi)以及消化道菌群种类。结果:与对照组相比,研究组的呕血、黑便及止血天数、pHi值明显减少(P<0.05);研究组消化道的幽门螺杆菌、葡萄球菌与大肠杆菌检出率明显较低,而乳酸杆菌和双歧杆菌检出率明显增加(P<0.05)。结论:立止血与双歧杆菌活菌联合治疗SU,临床疗效明显。     

前愈汤剂体外抗菌实验研究

[目的]探讨前愈汤剂体外抗菌效果.[方法]通过液体两倍稀释法分析前愈汤剂对大肠埃希氏菌、金黄色葡萄球菌、木糖葡萄球菌和克雷伯杆菌的抗菌作用,并以前列泰片和前列康作实验对照.[结果]前愈汤剂对以上菌种的最低抑菌浓度(MIC)值分别为62.5、125、125、500g/L.最低杀菌浓度(MBC)值分别为250、125、125、500g/L.[结论]前愈汤剂对大肠埃希氏菌、金黄色葡萄球菌、木糖葡萄球菌和克雷伯杆菌都有明显的抑制和杀灭作用.     

灵芝发酵川芎产物的抑菌性试验

目的研究川芎和灵芎菌质粗提液对大肠杆菌、枯草芽孢杆菌、金黄色葡萄球菌、青霉和黑曲霉供试菌的抑菌作用。方法用水、75%乙醇、95%乙醇、乙酸乙酯和丙酮等5种溶剂分别对川芎和灵芎菌质(灵芝发酵川芎产物)进行提取,以大肠杆菌、枯草芽孢杆菌、金黄色葡萄球菌、黑曲霉和青霉5种菌作为供试菌,采用滤纸片法测定川芎和灵芎菌质粗提液对供试菌的抑菌作用,并用平板稀释法测定最低抑菌浓度(MIC)。结果灵芎菌质的水和95%乙醇提取液对大肠杆菌和枯草芽孢杆菌的抑菌作用极显著优于川芎(P0.01);75%乙醇和丙酮提取液对大肠杆菌的抑菌作用极显著优于川芎(P0.01),对枯草芽孢杆菌的抑菌作用优于川芎(P0.05);灵芎菌质的95%乙醇提取液对青霉的抑菌作用极显著优于川芎(P0.01),丙酮提取液对青霉的抑菌作用优于川芎(P0.05);灵芎菌质的丙酮和乙酸乙酯提取液对金黄色葡萄球菌的抑菌作用显著优于川芎(P0.01)。结论灵芎菌质对大肠杆菌、枯草芽孢杆菌、金黄色葡萄球菌和青霉都具有不同程度的抑制作用且相对优于川芎。     

基于微生物快速鉴定系统对金黄色葡萄球菌耐药性鉴定新评估方法的建立

目的:探讨微生物快速鉴定系统(MALDI-Biotyper System)用于快速鉴定铜绿假单胞菌(Pseudomonas aeruginosa,P.a),临床分离金黄色葡萄球菌(Staphylococcus aureus,S.a)耐药性情况的可行性。方法:通过采用微生物快速鉴定系统及肉汤稀释法进行鉴定质控、临床分离金黄色葡萄球菌的耐药性情况,并将微生物快速鉴定系统检测结果与肉汤稀释法最低抑菌浓度(MIC)值进行比较分析,最后通过同步鉴定铜绿假单胞菌耐药性情况确定微生物快速鉴定系统的准确性及适用性。结果:微生物快速鉴定系统评分鉴定结果显示敏感质控菌株金黄色葡萄球菌评分值大于2.000,耐药质控菌株耐甲氧西林金黄色葡萄球菌(methicillin-resistant S.a,MRSA)评分值在1.700~2.000。临床分离金葡菌评分鉴定结果均在1.700~2.000,显示耐药,与肉汤稀释法MIC值结果一致。同时,无关质控敏感菌株铜绿假单胞菌的系统评分鉴定值大于2.000,显示敏感,其本身为敏感菌株,结果一致。结论:微生物快速鉴定系统评分方法可以用于金黄色葡萄球菌及铜绿假单胞菌耐药性情况的快速鉴定。     

紫草三黄栓体外抑菌活性的研究

目的研究紫草三黄栓体外抗菌活性。方法采用K-B纸片琼脂扩散法测定紫草三黄栓和阳性对照药马应龙痔疮栓对金黄色葡萄球菌、铜绿假单胞菌、大肠杆菌的抑菌强度;采用稀释法测定紫草三黄栓对金黄色葡萄球菌、铜绿假单胞菌、大肠杆菌的最低抑菌浓度(MIC)。结果紫草三黄栓对三种细菌的抑菌强度与阳性对照药相当;对金黄色葡萄球菌的最低抑菌浓度为16μg.mL~(-1)、铜绿假单胞菌的最低抑菌浓度为64μg.mL~(-1)、大肠杆菌的最低抑菌浓度为8μg.mL~(-1)。结论紫草三黄栓有较为显著的抗菌活性。     

左卡尼汀注射液中三甲胺分离提取及其离子色谱测定法研究

目的 建立分离提取左卡尼汀注射液中三甲胺含量的方法,及采用离子色谱法对三甲胺进行检测研究。方法 以L₉(3⁴)正交试验优化提取条件,采用0.25 mol·L^-1NaOH溶液创造碱性环境,40 ℃加热20 min,连接N₂流量为0.4 L·min^-1,用0.1 mol·L^-1甲磺酸溶液吸收气体,有效实现左卡尼汀注射液中三甲胺的分离提取;采用DionexIonPac^TM CS12A(4.0 mm×250 mm)色谱柱,DionexIonPac^TM CG12A(4.0 mm×50 mm)保护柱,以17 mmol·L^-1甲磺酸溶液为淋洗液,流速0.5 mL·min^-1,柱温30 ℃,电导检测。结果 三甲胺在0.30~48.64 μg·mL^-1内线性关系良好,相关系数为0.999 3,检出限为0.012 μg·mL^-1(S/N=3),定量限为0.3 μg·mL^-1(S/N=10),精密度良好,溶液置于室温24 h或2~8 ℃48 h内稳定。平均加样回收率在97.33~100.82%内,相对标准偏差(RSD)为1.07%(n=9)。结论 该方法能够有效提取分离左卡尼汀注射液中的三甲胺,离子色谱检测方法灵敏度高,回收率和重现性良好。     

UPLC同时测定通宣理肺胶囊中9个成分含量

目的:建立超高效液相色谱法(UPLC)同时测定通宣理肺胶囊中9个有效成分含量的方法。方法:采用Waters ACQUITY UPLCTM BEH C18色谱柱(100 mm×2.1 mm,1.7μm),流动相乙腈(A)-0.05%磷酸水溶液(B)梯度洗脱,流速为0.3 mL·min-1,柱温为30℃,检测波长为210 nm(盐酸麻黄碱、盐酸伪麻黄碱、苦杏仁苷、甘草苷)和278 nm(柚皮苷、橙皮苷、新橙皮苷、黄芩苷、甘草酸铵)。结果:盐酸麻黄碱、盐酸伪麻黄碱、苦杏仁苷、甘草苷、柚皮苷、橙皮苷、新橙皮苷、黄芩苷和甘草酸铵质量浓度的线性范围分别为0.005~0.037μg·mL^-1(r=0.9997)、0.002~0.017μg·mL^-1(r=0.9993)、0.005~0.039μg·mL^-1(r=0.9995)、0.010~0.084μg·mL^-1(r=0.9993)、0.010~0.082μg·mL^-1(r=0.9989)、0.007~0.052μg·mL^-1(r=0.9995)、0.007~0.057μg·mL^-1(r=0.9991)、0.038~0.301μg·mL^-1(r=0.9993)、0.002~0.020μg·mL^-1(r=0.9998);平均加样回收率均在98.2%~102.7%,RSD均小于1.5%。测定的样品中上述9个成分质量分数分别为1.12~1.23、0.39~0.54、1.19~1.31、2.54~2.67、2.49~2.66、1.54~1.66、1.66~1.79、9.19~9.54、0.53~0.69 mg·g^-1。结论:UPLC可用于宣理肺胶囊9个有效成分的含量测定,且方法简单、有效、结果准确。     

HPLC测定枳实薤白桂枝汤颗粒中辛弗林的含量

目的 建立枳实薤白桂枝汤颗粒中辛弗林含量测定方法.方法 采用Agela Technologies Promosil C18色谱柱(5 μm,100 A,4.6×250 mm);以甲醇 磷酸二氢钾溶液(0.6 g磷酸二氢钾,1.0 g十二烷基磺酸钠,1 mL冰醋酸用1000mL水定容)50 ∶ 50为流动相;柱温34℃;...     

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??OBJECTIVE To evaluate the antibacterial activity of levofloxacin, moxifloxacin and nemonoxacin against Staphylococcus aureus in vitro. To assesse the impact of gyrA and parC genes mutant on resistance for quinolones and the sequences of gyrA and parC genes for three quinolones. METHODS The MICs of 50 S. aureus were detected by agar dilution method. The MIC and MPC against four S. aureus which were special gene mutations were detected by agar dilution method. Based on these results, bacterial recovery growth curve of levofloxacin, moxifloxacin and nemonoxacin were traced. RESULTS Nemonoxacin demonstrated activities 8- to 32- fold more potent(MICs at which 90% of isolates were inhibited , 0.5 ??g??mL^-1) than those of moxifloxacin(MIC₉₀, 2 ??g??mL^-1) and levofloxacin (MIC₉₀, 16 ??g??mL^-1) against 50 S. aureus.In condition of the same drug concentrations, the bacterial recovery growth ratios of nemonoxacin was the lowest, while levofloxacin??s was the highest. RN450A3 recovery growth ratio was highest compared with other mutant bacterial strains, while RN450 recovery growth ratio was lowest.CONCLUSION The antibacterial activities of nemonoxacin, moxifloxacin and levofloxacin against S. aureus in vitro are:nemonoxacin?? moxifloxacin ??levofloxacin. Compared with levofloxacin and moxifloxacin, nemonoxacin inhibits bacteria in a lower concentration, and nemonoxacin is utterly efficacious with different genes mutant strains.The target preference of levofloxacin may be the parC gene of topoisomerase ??, while moxifloxacin and nenomoxacin can almost act on the gyrA and parC gene at the same time.     

LC-MS/MS检测早产儿咖啡因血清浓度及其临床应用

目的 建立液相色谱串联质谱法测定早产儿咖啡因血清浓度的方法,并对66例早产儿给药7 d后的咖啡因血清谷浓度进行监测及疗效评估。方法 采用咪达唑仑作为内标,样品经过蛋白沉淀法处理后上样。色谱柱为UPLC BEH C₁₈(2.1 mm×50 mm,1.7 μm),柱温设为30 ℃,流动相由甲醇和0.1％甲酸组成,梯度洗脱,流速为0.4 mL·min^-1,洗脱时间为4 min,进样量10 μL。收集连续使用咖啡因7 d后早产儿血清,进行咖啡因血清谷浓度测定,并观察患儿在用药第8~12天疗效和不良反应发生情况。结果 血清中内源性物质不干扰待测物定量,咖啡因血清浓度在0.5~64 μg·mL^-1内呈良好的线性关系(R²为0.999 5)。66例早产儿给药7 d后咖啡因血清浓度为(8.92±6.31) μg·mL^-1,治疗有效率和不良反应发生率分别为68.18%和10.60%。咖啡因血清浓度水平与治疗有效率和不良反应发生率均有一定的关系,具有统计学意义(P值分别为0.033和0.011)。结论 本方法灵敏度高,重复性好,所需样本量少,能快速准确测定咖啡因血清浓度,为临床安全有效使用咖啡因治疗早产儿呼吸暂停提供保障和依据。     

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??OBJECTIVE To establish a determination method of molecular-size distribution of human serum albumin (HSA) by ultra performance liquid chromatography (UPLC).METHODS An UPLC method was developed to specifically determine the polymers and other components in HSA on UPLC PROTEIN BEH SEC analysis column with Waters Alliance UPLC system and Waters UPLC TUV detector. The separation was performed using a mobile phase consisting of PBS at a flow rate of 0.6 mL??min^-1 and the UV detection wavelength was set at 280 nm. HSA samples were diluted to different concentrations (0.5-20 mg??mL^-1) to confirm the optimal concentration range of the injection. The change of component percentage and the linear relationship between HSA concentration and chromatographic peak height were confirmed and the molecular-size distribution was calculated by area normalization method. Within the optimum injection concentration range, the national control sample for HSA was diluted to 12 mg??mL^-1 and tested by UPLC method and the methodology was confirmed. Twenty batches of HSA samples were determined by both UPLC and existing HPLC methods, and the samples were determined in parallel. The consistency of the methods was compared and the method was reconfirmed.RESULTS The UPLC retention time of HSA polymer was 1.469 min, of dimer was 1.972 min, and of monomer was 2.267 min, respectively. The resolution of dimer and monomer was 2.20 and the USP tailing of monomer peak was 1.18 respectively. In the range of the injection concentrations, 0.5-20 mg??mL^-1, there was linear relationship between the concentrations of the components of 11 HAS samples, including polymer, dimer and monomer peak, and the peak area%, peak height, peak area, and the squares of linear correlation coefficient were all greater than 0.997 0. The components peak area percentage of HSA samples remained relatively stable within the concentration range of 10-16 mg??mL^-1 (total injection amount of 100-160 ??g). The RSDs of the percentage of polymers were 0.00% (n=3, 10 mg??mL^-1), 0.10% (n=3, 12 mg??mL^-1), and 0.10% (n=3, 16 mg??mL^-1), respectively. The UPLC method was used to determine the national control sample for human albumin of 12 mg??mL^-1, and the mean value of peak area% was 5.62% (n=10). The results were consistent with those of the parallel determination by HPLC (5.58%), both of which were in accordance with the quality control range of the national standard for human albumin. The RSD of the percentage of the peak area of the polymers in national standard for human albumin by UPLC was 0.40% (n=10). The HPLC and UPLC methods were used to determine the polymer peak area percentage of 20 batches of HSA samples from 7 domestic and foreign enterprises at the concentration of 12 mg??mL^-1. The correlation coefficient of the two methods was 0.996 0 (P> 0.05) and there was no significant difference between the two methods (P>0.05).CONCLUSION Compared with the traditional HPLC method, the detection time of HSA SEC by the proposed UPLC method is shortened by at least 10 times, and the accuracy and repeatability of the determination are satisfactory. UPLC method can save much more analysis time, is simple and much faster, and can be used for high-throughput determination of molecular-size distribution of human serum albumin.     

UPLC-MS/MS快速同时测定犬血浆中补骨脂的10个活性成分及补骨脂犬药代动力学研究

建立了同时测定犬血浆中补骨脂的10个活性成分——补骨脂素、异补骨脂素、5,7,4′-三羟基黄酮、5,7,4′-三羟基异黄酮、补骨脂宁、新补骨脂异黄酮、补骨脂甲素、补骨脂二氢黄酮甲醚、补骨脂苷、异补骨脂苷的UPLC-MS/MS分析方法,并对比格犬灌胃给药补骨脂水提取物后不同时间点血浆中的各成分的浓度进行了测定,绘制了血药浓度-时间曲线,并用WinNonlin软件计算了补骨脂活性成分在犬体内的药代动力学特征。液相色谱采用Waters HSS-T3色谱柱(2.1 mm×100 mm,1.8μm);流动相为乙腈-水(含0.004%甲酸),梯度洗脱;质谱检测采用电喷雾离子源,正离子多反应监测模式,分析时间8.5 min。方法学经专属性、准确度、精密度、线性范围、回收率、基质效应和样品稳定性等考察,符合要求,可满足犬灌胃补骨脂提取物后药代动力学研究。犬灌胃给药后,补骨脂10个化学成分的T_max为1.92~5.67 h;其中补骨脂素、异补骨脂素、补骨脂苷和异补骨脂苷的Cmax为383~3613 ng·mL^-1,AUC0-∞为3556~18949 ng·h·mL^-1,半衰期t1/2为2.45~4.83 h;其余6个化合物的Cmax为0.81~19.9 ng·mL^-1,AUC0-∞为6.54~178 ng·h·mL^-1,半衰期t1/2为2.95~7.29 h。该研究建立的UPLC-MS/MS分析方法快速、准确、灵敏,适合于比格犬灌胃给药补骨脂提取物后的药代动力学研究,为合理解释中药补骨脂的有效性和毒副作用以及其后续相关研究提供了药代动力学信息。     

UPLC-MS/MS测定大鼠血浆中奥拉帕利的浓度及其药动学研究

目的 建立一种具有高灵敏度和高选择性测定大鼠血浆中奥拉帕利药物浓度的超高效液相色谱-串联质谱(UPLC-MS/MS)检测方法,并研究奥拉帕利在大鼠体内的药动学特征。方法 大鼠血浆样品采用乙腈沉淀法去除蛋白,色谱柱为Acquity UPLC BEH C₁₈柱 (2.1 mm×50 mm,1.7 μm),流动相为0.1%甲酸水溶液-乙腈,梯度洗脱。质谱采用电喷雾离子源,多反应监测正离子模式,奥拉帕利定量离子对为m/z 435.19→m/z 367.1。以50 mg·kg^-1奥拉帕利经大鼠灌胃给药后于不同时间点采集血样,利用建立的方法进行检测分析,并用DAS2.0软件计算药动学参数。结果 血浆中奥拉帕利在(0.6~1 821) ng·mL^-1内线性关系良好,定量限为0.15 ng·mL^-1,日内、日间精密度RSD均小于5.5%,准确度在95.4%~99.2%内,平均提取回收率为84.2%~96.0%,基质效应在91.4%~108.8%内。结论 建立的方法灵敏度高、结果准确,可用于大鼠血浆中奥拉帕利质量浓度的测定及其药动学研究。     

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??OBJECTIVE To characterize the metabolism of genistin and study its enzymatic kinetics in rat liver microsome by HPLC-MS. METHODS Genistin was incubated with rat liver microsomal incubation system. HPLC-MS method was used to characterize the metabolites. A metabolite generation method was established for quantitative analysis of genistein with sulfamethoxazole as internal standard.The enzyme kinetics parameters V_max and K_m was calculated by the GraphPad Prism 5.0 software. RESULTS The metabolites in vitro incubation system was identified as genistein.The optimal time in rat liver microsomes incubation time of 40 min,the optimal protein concentration of 1 mg??mL^-1,a substrate concentration of 50 ??mol??L^-1.The enzyme kinetics parameters of genistein were as follows: V_max=(0.104 2??0.003 3) ??mol??min^-1??mg(pro)^-1, K_m=(28.96??2.80) ??mol??L^-1. CONCLUSION The results indicate that genistin can be metabolited as the form of hydroxylation in rat liver microsome. Metabolite generation method is a reliable and simple method for determination of kinetic parameters of hepatic microsomal enzymes, and enzyme kinetic parameters obtained of genistin provide important parameters for further study.