Simultaneous high-performance liquid chromatographic determination of puerarin, daidzin, paeoniflorin, liquiritin, cinnamic acid, cinnamaldehyde and glycyrrhizin in Kampo medicines

We report a high-performance liquid chromatographic method to determine the quantities of puerarin, daidzin, paeoniflorin, liquiritin, cinnamic acid, cinnamaldehyde and glycyrrhizin in Kampo medicine. All seven compounds were separated in less than 30 min with a Wakosil-II 5C18 AR column by linear gradient elution using 0.01% (v/v) phosphoric acid acetonitrile (0 min 90:10, 10 min 88:12, 22 min 70:30, 30 min 30:70) as the mobile phase at a flow-rate of 1.0 ml/min(-1), and detection at 250 nm. The detection limits of these compounds are 0.15-0.3 microM with response linearity. This method was applied to determine the quantities in eight Kampo decoctions; Mao-to, Makyo-yokukan-to, Makyo-kanseki-to, Yokuinin-to, Sho-seiryu-to, Keima-kakuhan-to, Kakkon-to and Kakkon-to-ka-senkyu-sin'i. Glycyrrhizin content was lower in both the decoction and the methanol-diluted decoction of Sho-seiryu-to compared with the others. Low pH due to organic acids of Schisandrae fructus in the decoction caused inhibition for glycyrrhizin dissolution in Sho-seiryu-to.     

配伍对麻黄甘草药对中麻黄类生物碱在大鼠体内组织分布的影响

目的：研究麻黄与甘草配伍前后麻黄类生物碱在 SD 大鼠组织分布特点,从药物相互作用角度揭示甘草对麻黄类生物碱组织分布的影响。方法采用超高效液相色谱-串联质谱（ UPLC - MS/ MS）法在多反应监测（MRM）模式下测定麻黄与甘草配伍前后不同时间点大鼠各组织液中麻黄类生物碱的浓度,比较麻黄与甘草配伍前后麻黄类生物碱组织分布的差异。结果与麻黄组相比,麻黄与甘草配伍后,促进了去甲基麻黄碱、麻黄碱和伪麻黄碱在研究组织中的吸收和分布;加快了去甲基麻黄碱、麻黄碱、伪麻黄碱和甲基麻黄碱从脑组织中消除;抑制了去甲基伪麻黄碱在心和脑组织中的吸收和分布,并加快了其从心和脑组织中的消除;不同程度地加快了去甲基麻黄碱从心、肺、肾的消除,加快了麻黄碱从心组织中消除,加快了甲基麻黄碱从肺和肾中的消除。结论麻黄与甘草配伍对麻黄类生物碱在大鼠组织中的分布具有显著的影响。     

关于《中国药典》中麻黄生物碱含量测定指标的探讨

目的能够全面控制麻黄药材的质量。方法对《中国药典》麻黄药材中麻黄类生物碱含量测定指标进行探讨。结果与结论建议在测定单一成分含量的同时,增加测定其他生物碱的含量,从而对麻黄药材质量有更全面的控制和评价。     

Simultaneous determination of baicalin, wogonoside, baicalein, wogonin, berberine, coptisine, palmatine, jateorrhizine and glycyrrhizin in Kampo medicines by ion-pair high-performance liquid chromatography.

An ion-pair high-performance liquid chromatographic method for the simultaneous determination of four flavonoids, namely baicalin, wogonoside, baicalein and wogonin, and four berberine-type alkaloids, namely berberine, coptisine, palmatine and jateorrhizine, and glycyrrhizin in Kampo medicines is described. The analysis can be accomplished within 30 min with a Wakosil-II 5C18 HG column by linear gradient elution using a mobile phase containing aqueous phosphoric acid, sodium dodecyl sulfate and acetonitrile at a flow-rate of 1.0 ml x min(-1), a thermostatic oven at 45 degrees C, and detection at 265 nm. The method was applied to quantifying these components in three Kampo decoctions: Oren-gedoku-to, San'o-shashin-to and Hange-shashin-to. The decoctions were diluted with 65% methanol at the final stage because a large quantity of precipitate, mainly from baicalin and berberine, was formed. The within-day relative standard deviations were less than 2.02% (n=10). The recoveries of these compounds were 90.3-102%. The detection limits of these compounds were 0.02-1.96 microM per injection (5 microl).     

Simultaneous determination of six alkaloids in ephedrae herba by high performance liquid chromatography

A new and rapid analytical method for the simultaneous determination of six EPHEDRA alkaloids (ephedrine, pseudoephedrine, norephedrine, norpseudoephedrine, methylephedrine, and methylpseudoephedrine) in Ephedrae Herba by high performance liquid chromatography (HPLC) has been developed. Recoveries of each alkaloid were 96.0-101.6% with coefficients of deviation of 1.7-3.0%. The contents of six alkaloids in three species of EPHEDRA indigenous to China are determined and reported.     

HPLC-UV法同时测定麻黄中5种麻黄生物碱的含量

目的建立同时分离检测麻黄中麻黄碱(Ephedrine)、伪麻黄碱(Pseudoephedrine)、去甲麻黄碱(Norephedrine)、去甲伪麻黄碱(Norpseudoephedrine)和甲基麻黄碱(Methylephedrine)的含量测定方法。方法采用HPLC-UV法,色谱柱:苯基柱(Alltima Phenyl,250mm×4.6 mm,5μm);流动相:0.02 mol/L磷酸二氢钾溶液-乙腈(96∶4);流速:0.6 ml/min;检测波长:210 nm;柱温:25℃。结果 5种生物碱在13.5 min内即可达到完全分离,E在2.000 8～40.016 0μg/ml线性关系良好;PE在1.003 6～20.072 0μg/ml线性关系良好;NME在0.199 2～3.984 0μg/ml线性关系良好;NMP在0.200 0～4.00 0μg/ml线性关系良好;ME在0.200 4～4.008 0μg/ml线性关系良好。各生物碱的平均回收率均在92%～104%(RSD≤5.76%)。结论本方法可操作性强,简便快速,分离效果好,重现性好,可为麻黄及含麻黄的中西药制剂提供有效的检测手段。     

半夏糖浆质量标准研究

目的研究半夏糖浆（麻黄、枇杷叶、陈皮等）内在质量的控制方法。方法采用薄层色谱（TLC）法对方中的枇杷叶、陈皮、甘草3味药材进行定性鉴别；用高效液相色谱（HPLC）法对方中麻黄的有效成分盐酸麻黄碱进行含量测定。结果定性鉴别薄层色谱特征明显，专属性强；方法学考察表明，盐酸麻黄碱进样量在0．0854-0．5126μg范围内与峰面积具有良好的线性关系（r=0.9999），平均回收率为99．30％（n=9），RSD=0.31％。结论方法准确、可靠、重现性好，可用于半夏糖浆的质量控制。     

HPLC-ESI-TOF-MS法优选麻黄中麻黄碱的提取工艺

目的优选麻黄中麻黄碱的最佳提取工艺。方法水煎法获得麻黄药材总提取物,反相高效液相色谱-电喷雾-飞行时间串联质谱选择离子对法测定总提物中麻黄碱的含量,正交设计优选最佳提取工艺。结果方法检测限为0.05μg·mL-1,线性范围为0.005～0.5mg·mL-1;麻黄碱最佳提取工艺为:加30倍量水,75℃加热40min,提取4次。结论该检测方法灵敏、准确,可用于复杂体系中麻黄碱的测定;水煎法经济、绿色、无污染,可作为麻黄碱等生物碱类物质的提取方法。     

Pharmacokinetic determination of ephedrine in Herba Ephedrae and Wu Tou Tang decoctions in rats using ultra performance liquid chromatography tandem mass spectrometry

A rapid and sensitive ultra performance liquid chromatography tandem mass spectrometry method was developed and validated for the determination and quantification of ephedrine in rat plasma samples. An Acquity UPLC BEH C18 column (1.7 μm, 2.1?mm?×?50?mm) was used for chromatographic separation. Electrospray ionization in the positive mode was used, and the precursor-fragment ion pairs of m/z 166/148 and m/z 289/97 were adopted to characterize ephedrine and testosterone (internal standard), respectively. The method was validated using 10, 100 and 500?ng/mL of ephedrine. It demonstrated adequate levels of precision and accuracy, matrix effect, extraction recovery and stability. Linearity over the concentration range of 0.5-2000?ng/mL was acceptable with a correlation coefficient (r2) better than 0.990. To determine the pharmacokinetic behaviour of this sympathomimetic compound in the Sprague-Dawley rats, ephedrine hydrochloride, Herba Ephedrae single-herb and Wu Tou Tang decoctions were administered orally, and ephedrine hydrochloride was also administered by intravenous injection, and blood samples were collected over 24?h. Ephedrine was measured in plasma and pharmacokinetic parameters were determined by using the standard non-compartmental method and calculated by using Practical Pharmacokinetic Program-Version 87/97. The AUC(0-t) and T(max) values were significantly different (p?相似文献   

毛细管区带电泳法测定麻黄中麻黄碱和伪麻黄碱的含量

目的建立测定麻黄中麻黄碱和伪麻黄碱含量的毛细管区带电泳（CZE）法。方法以200 mmol.L-1硼砂-500 mmol.L-1硼酸溶液（1∶1,v/v）含2 mg.mL-1庚烷磺酸钠和0.4%（v/v）乙腈为背景电解质（BGE）,石英毛细管（75 cm×75μm）,有效分离长度60 cm,运行电压15 kV,紫外检测波长210 nm,重力进样20 s（高度10 cm）,以苯甲醇作内标,对麻黄中的麻黄碱和伪麻黄碱进行定量分析,以两者含量为参量对10批麻黄进行系统聚类分析并进行质量评价。结果麻黄碱和伪麻黄碱的相对峰面积（Ai/Ar）与各自的质量浓度C（mg.mL-1）呈良好的线性关系,两者的平均回收率分别为100.4%和100.9%。结论该法准确可靠,操作便捷,可用于麻黄中麻黄碱和伪麻黄碱的含量测定。     

小儿止咳口服液质量标准研究

目的 :建立小儿止咳口服液的质量标准。方法 :采用薄层色谱 (TLC)法对处方中的麻黄、陈皮、甘草等进行定性鉴别 ;用高效液相色谱法测定绿原酸的含量 ,流动相为甲醇 -水 (10∶90) ,流速为1 0ml/min ,检测波长为254nm。结果 :在TLC色谱中均能鉴别出麻黄、陈皮、甘草 ;绿原酸进样量在0 2μg～1 2μg范围线性关系良好 (r=0 9996) ,平均回收率为96 67 %,RSD=1 04 %。结论 :本方法可准确、快速地对小儿止咳口服液进行定性、定量检测 ,可用于该制剂的质量控制。     

甘肃栽培麻黄镇咳、平喘、抗炎、利尿作用研究

目的研究甘肃栽培麻黄的镇咳、平喘、抗炎、利尿作用,并与野生麻黄比较。方法采用氨水诱咳法、气管容积法、耳廓肿胀法及代谢笼法,测定各药理作用指标。结果2种麻黄均可减少小鼠咳嗽反应的次数,对抗气管平滑肌痉挛性收缩,减轻小鼠耳廓肿胀度,增加大鼠尿量。结论甘肃栽培麻黄在镇咳、平喘、抗炎、利尿作用方面与野生麻黄作用相当,可替代野生麻黄药用。     

Ephedra alkaloid contents of Chinese herbal formulae sold in Taiwan

Consumption of Ephedra alkaloids is prohibited in‐competition by the World Anti‐Doping Agency (WADA). In Taiwan, colds are often treated with Chinese herbal formulae containing Herba Ephedrae. We screened products sold in Taiwan and preliminarily assessed their relationships with WADA threshold violations. Fifty‐six concentrated powder products, including 19 Chinese herbal formulae that contained Herba Ephedrae, were collected. The content of Ephedra alkaloids, namely ephedrine (E), methylephedrine (ME), norpseudoephedrine (NPE; cathine), pseudoephedrine (PE), and norephedrine (NE; phenylpropanolamine), was determined using a validated high‐performance liquid chromatography method. The results revealed that the phenotypic indicators of the collected products, E/PE and E/total ratios, were 1.52–4.70 and 0.49–0.72, respectively, indicating that the Herba Ephedrae species in these products was probably E. sinica or E. equisetina, but not E. intermedia. The contents of E, ME, NPE, PE, and NE and the total alkaloid contents in the daily doses of the products were 0.45–34.97, 0.05–4.87, 0.04–3.61, 0.15–12.09, and 0.01–2.00 mg and 0.68–53.64 mg, respectively. The alkaloid contents followed a relatively consistent order (E > PE > ME ≈ NPE > NE), even for products from different manufacturers. We calculated that single doses of 50.0% and 3.6% of the products would result in the WADA thresholds of E and NPE being exceeded, respectively. Our data provide critical information for athletes and medical personnel, who should be wary of using complex Chinese herbal formulae in addition to over‐the‐counter products.     

Pharmacological properties of traditional medicines. XXV. Effects of ephedrine, amygdalin, glycyrrhizin, gypsum and their combinations on body temperature and body fluid

Effects of ephedrine, amygdalin, glycyrrhizin, gypsum and their combinations on body temperature and body fluid were studied in rats using the method developed in our previous reports. Ephedrine significantly increased respiratory evaporative water loss and heat loss in response to a marked elevation of body temperature. There was a small but significant increase in body temperature when amygdalin was orally given rats at a dose of 46.32 mg/kg. Glycyrrhizin and gypsum were unable to affect body temperature. However, gypsum was able to prevent the increased action of ephedrine on body temperature, amygdalin exhibited a preventive tendency to it, and glycyrrhizin did not affect it. The results are in good agreement with classical claims of Makyo-kanseki-to and the related crude drugs in traditional medicine. Moreover, a combination of the four components reproduced the effects of Makyo-kanseki-to on body temperature and body fluid. This report suggests that the co-administration of ephedrine and gypsum is physiologically more desirable than ephedrine alone for dry-type asthmatic patients with a fever. Also, it experimentally supports the clinical efficacy of Makyo-kanseki-to.     

金麻杏止咳片快速薄层色谱鉴别与3成分同时定量测定研究

目的 建立简便、快捷的金麻杏止咳片质量控制方法.方法 在5块薄层板上鉴别前胡、黄芩、麻黄、枇杷叶、金银花、苦杏仁和桔梗.采用高效液相色谱法,以乙腈-甲醇-0.1%磷酸(4.5:9:86.5)为流动相,测定样品中盐酸麻黄碱、盐酸伪麻黄碱与苦杏仁苷的含量.结果 通过方法学考察,盐酸麻黄碱进样量在0.0505～0.505 μg(r=0.999997),苦杏仁苷进样量在0.1488～1.488μg(r=0.99996)范围内分别与峰面积呈良好的线性关系.盐酸麻黄碱回收率为100.14%,RSD为1.98%(n=9);苦杏仁苷回收率为97.88%,RSD为1.64%(n=9).结论 所用方法简便、快捷、实用,可用于金麻杏止咳片的质量控制.     

HPLC测定宣肺清热合剂中盐酸麻黄碱与盐酸伪麻黄碱含量

目的建立宣肺清热合剂（麻黄、苦杏仁、浙贝母、黄芩等）中有效成分盐酸麻黄碱和盐酸伪麻黄碱的含量测定方法。方法采用高效液相色谱法,色谱柱Agilent ZORBA×SB-C18（5μm,4.6 mm x 250 mm）,流动相为乙腈-0.02 mol/L磷酸二氢钾（用磷酸调pH至2.7）,（4：100,含0.1%三乙胺）,流速：1.2 ml/min,检测波长：207 nm。结果盐酸麻黄碱在11.04~222.8μg/mL（r=0.999 9）,盐酸伪麻黄碱在8.27~165.4μg/ml（r=0.999 9）的浓度范围内均线性关系良好,回收率均在95%~105%之间,RSD均小于3%。结论本方法操作简便、重现性好,可以更有效地控制制剂的质量。     

慢支咳喘膏的质量标准研究

目的:建立慢支咳喘膏的质量控制方法。方法:采用薄层色谱(TLC)法对方中麻黄、黄芪、甘草进行鉴别;采用高效液相色谱法对制剂中盐酸麻黄碱和盐酸伪麻黄碱进行含量测定。结果:TLC专属性好、斑点清晰;盐酸麻黄碱、盐酸伪麻黄碱进样量分别在1.3～32.5(r=0.9997)、3.4～85μg(r=0.9998)范围内与其峰面积积分值呈良好线性关系,麻黄碱、盐酸伪麻黄碱的平均回收率分别为97.84%、97.85%,RSD分别为0.63%、1.10%(n均为9)。结论:本试验建立的方法简便、准确、重现性好,能有效控制慢支咳喘膏的质量。     

Herbal medicine and false-positive results on lymphocyte transformation test

In vitro mitogenic activity of 16 herbs and 3 Kampo (herbal medicine) formulae have been reported in experimental studies. It is not known how many herbs and Kampo formulae in total have mitogenic activity. Lymphocyte transformation test (LTT) is generally utilized to diagnose drug-induced liver injury. In LTT, mitogenic activity is assessed by measuring 3H-thymidine incorporation. The objective of the present study was to determine which herbs and which Kampo formulae caused false-positivity on LTT. We examined 2496 summaries of all admission records from 1979 to 1999 in our department. We selected patients in whom liver injuries were diagnosed as definitely unrelated to Kampo medication. In these patients, LTT was performed for some herbs contained in the suspect Kampo medicines, resulting in positive LTT for 17 herbs: Evodiae Fructus (Goshuyu), Zizyphi Fructus (Taiso), Ginseng Radix (Ninjin), Zingiberis Rhizoma (Shokyo), Hoelen (Bukuryo), Aconiti Tuber (Bushi), Angelicae Radix (Toki), Cnidii Rhizoma (Senkyu), Rehmanniae Radix (Jio), Ephedrae Herba (Mao), Anemarrhenae Rhizoma (Chimo), Cinnamomi Cortex (Keihi), Bupleuri Radix (Saiko), Artemisiae Capillari Spica (Inchinko), Persicae Semen (Tonin), Moutan Cortex (Botanpi) and Paeoniae Radix (Shakuyaku). These results were considered false-positive, because the results were observed in the "definitely unrelated" patients. Mitogenic activity inherent to some herbs and Kampo formulae may sometimes cause false-positivity on LTT in clinical situations. These examples suggest that LTT for Kampo formulae may be unreliable as a diagnostic method for drug-induced liver injury.     

HPLC法同时测定百喘朋中盐酸麻黄碱和盐酸苯海拉明的含量

目的 :建立HPLC法同时测定百喘朋中盐酸麻黄碱和盐酸苯海拉明的含量。方法 :采用DiamonsilC18(5μ,4.6mm×150mm)色谱柱 ,流动相为乙腈 -水 -磷酸 (60∶40∶0 1 ,每1000ml含4 0g 十二烷基硫酸钠 ) ,检测波长为254nm ,采用外标法。结果 :盐酸麻黄碱、盐酸苯海拉明的线性范围分别为2 69μg～32.28μg、2 54μg～30.48μg ,回收率 (n=5)分别为100 4 % (RSD=1.1 % )和99 8 % (RSD=0.9 % )。结论 :本法可同时准确测定百喘朋中盐酸麻黄碱和盐酸苯海拉明的含量。     

Determination of ephedrine alkaloids in Ephedra natural products using HPLC on a pentafluorophenylpropyl stationary phase

In this study a pentafluorophenylpropyl (PFPP) stationary phase was applied to the fast and reliable qualitative and quantitative analysis of ephedrine alkaloids in Ephedra plant material and derivatives. A Discovery HS F5 column (150mmx4.6mm i.d., 5microm) was used, with an isocratic mobile phase composed of ammonium acetate (7mM) in acetonitrile-water (90:10, v/v), at a flow rate of 1.0ml/min. The column temperature was set at 45 degrees C. UV detection was set at 215 and 225nm. The total analysis time was 16min. The validation parameters, such as linearity, sensitivity, accuracy, precision and specificity, were found to be highly satisfactory. Sonication and microwave extractions were compared in order to optimize the yield of the target analytes. Under the optimized extraction conditions (based on two cycles of sonication with methanol at 40 degrees C for 15min), different matrices containing Ephedra were successfully analyzed for their alkaloid content. The method was applied to the analysis of standard reference materials (SRMs) containing Ephedra. Furthermore, the developed technique allowed the simultaneous determination of ephedrine alkaloids and synephrine, the main phenethylamine alkaloid of Citrus aurantium, that has replaced Ephedra in dietary supplements used in the treatment of obesity. The results indicated that this procedure is suitable for the phytochemical analysis of Ephedra plant material and extracts, and can be applied to demonstrate the label claims for product content, including the absence of ephedrine alkaloids in Ephedra-free products.