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.     

A rapid and simple procedure for the determination of ephedrine alkaloids in dietary supplements by gas chromatography-mass spectrometry

A simple method for the determination of ephedrine alkaloids: ephedrine (EF), pseudoephedrine (PE), norpseudoephedrine (NPE), norephedrine (NE) and methylpseudoephedrine (MPE) in dietary supplements by gas chromatography–mass spectrometry is described. After the addition of 3,4-methylenedioxypropylamphetamine as internal standard, a liquid–liquid extraction procedure in alkaline conditions with chloroform/isopropanol (9:1, v/v) was applied to the samples prior to analysis. Chromatography was performed on a fused capillary column and analytes, derivatized with pentafluoropropionic anhydride, were determined in the selected-ion-monitoring (SIM) mode. The method was validated in the range 0.3–10 μg/mg for EP, 0.06–2.5 μg/mg for PE and NPE and 0.04–1 μg/mg NE and MPE. Mean recovery ranged between 65.7 and 81.3% for the different analytes in dietary supplements. The quantification limits were 0.3 μg/mg for EP, 0.06 μg/mg for PE, 0.04 μg/mg for NPE, NE and MPE. The method was applied to analysis of various dietary supplements containing Ma-huang (Ephedra Sinica) and Sida Cordifolia plant extracts promoted for aiding weight control and boosting sports performance and energy.     

Effects of harvesting time and processing on the contents of five alkaloids in the herbaceous stems of Ephedra sinica

In the present study, we studied the changes of the contents of alkaloids in the herbaceous stems of Ephedra sinicaduring different harvest periods as well as before and after processing. The alkaloid contents of 39 batches of ephedra herb, prepared slices ephedra and honey-fried ephedra, 24 batches of ephedra herb with different harvesting periods, which were all collected from cultivation base in Inner Mongolia, and 38 batches of prepared slices ephedra purchased from the market were detected by taking norephedrine (NE), norpseudoephedrine (NPE), ephedrine (E), pseudoephedrine (PE) and methylephedrine (ME) as indicators by using HPLC method. The content of total alkaloid in prepared slices ephedra (1.71%–3.14%) was higher than that in ephedra herb (1.20%–2.53%) and honey-fried ephedra (1.52%–2.99%). Contents of different alkaloids in these three types of samples were significantly different. Prepared slices ephedra and honey-fried ephedra showed significant differences in the contents of NE, NPE and ME (P<0.05), and the contents of E were significantly different between ephedra herb and honey-fried ephedra (P<0.05). The total alkaloid content of ephedra herb was the highest in September (3.10%). Alkaloid contents of prepared slices ephedra collected in the market were uneven and 13%–91% lower than those collected from cultivation base. The results provided a basis for the quality evaluation of ephedra herb and its processed products, and had certain guiding significance for the selection of processed ephedra according to different drug purposes in clinical application. It also provided data support for the harvesting time of ephedra herb.     

十二种国产麻黄的品质评价

本文应用高效液相色谱法对我国24个产地所产的12种麻黄生药进行了六种生物碱的定量分析,这六种生物碱是:麻黄碱(ephedrine),伪麻黄碱(pseudoephedrine),去甲基麻黄碱(norephedrine),去甲基伪麻黄碱(norpseudoephedrine),甲基麻黄碱(methylephedrine)和甲基伪麻黄碱(methylpseudoephedrine)。根据分析结果,对这些麻黄生药的品质作出了评价。     

Comparison of contents of five ephedrine alkaloids in three official origins of Ephedra Herb in China by high-performance liquid chromatography

In the present paper, a fast and economical HPLC method [on a Phenomenex Polar-RP column with a solution of (phosphoric acid:triethylamine:dibutylamine:water = 0.40:0.1:0.2:499.3) and methanol] is developed, and applied for the determination of norephedrine, norpseudoephedrine, ephedrine (E), pseudoephedrine (PE) and methylephedrine (ME) in 64 samples of three species from main habitats in China. Quantitation data showed that total alkaloid content in Ephedra equisetina Bge. (2.708 ± 0.642%) is higher than that in E. sinica Stapf. (1.365 ± 0.624%) and E. intermedia Schrenk et C. A. Mey. (1.537 ± 0.746%), but the range of total alkaloid content in each species is so wide that the ranges of the three species greatly overlap. The contents of E, PE and ME are different among the three species. The ratio E/total alkaloid content and ratio E/PE as well as E and ME contents can be used for identification of E. sinica, E. intermedia and E. equisetina from one another.     

A comparative study on commercial samples of phellodendri cortex

A total of 31 commercial samples of Phellodendri cortex (Rutaceous plant) which originated from PHELLODENDRON AMURENSE Ruprecht, P. CHINENSE Schneid, P. WILSONII Hayata et Kanehira, and P. AMURENSE Rupr. var. SACHALINENSE Fr. Schm., respectively, were collected from the Taiwan and Japan herbal markets. The contents of five quaternary alkaloids (berberine, palmatine, jatrorrhizine, phellodendrine and magnoflorine) in these samples were determined by capillary electrophoresis. It was found that P. WILSONII (4.10 +/- 0.78%) and P. AMURENSE var. SACHALINENSE (4.18 +/- 1.03%) were superior to P. AMURENSE (1.55 +/- 0.72%) and P. CHINENSE (1.54 +/- 0.60%). Berberine was the major alkaloid in almost all samples. In comprised about 80% of the total alkaloids in the first two of the four herbs named above, but only about 40% in the latter two. From the data on the chemical analysis of the herb's constituents, as well as the herb's texture and color, we can postulate the origin and quality of a herb drug.     

Survey on resources of Ephedra plants in Xinjiang

The resources of wild Ephedra plants in the Xinjiang Uygur Autonomous Region were surveyed. Ephedra plants mainly grow on the fringes of the Taklimakan Desert and Gureban-tonggute Desert. We found six genotypes of Ephedra przewalskii growing widely in Xinjiang. Three genotypes of Ephedra intermedia were limited to the northern and eastern parts, and Ephedra regeliana scattered in the northern part of Xinjiang. These Ephedra specimens were analyzed for DNA sequences of nuclear ribosomal DNA, internal transcribed spacers 1 and 2, chloroplastic DNA, trnL intron and trnL-trnF intergenic spacer. Intraspecific variation of the nucleotide sequence in E. przewalskii was found in different habitats. Norephedrine, ephedrine, pseudoephedrine, and methylephedrine contents of the specimens were determined. Although Ephedra intermedia of all three genotypes contained ephedrine alkaloids, ephedrine alkaloids were not detected in E. regeliana and E. przewalskii.     

基原对半夏急性毒性和主要化学成分含量影响研究

目的对不同基原半夏的生物碱类和有机酸类成分含量和急性毒性大小进行比较研究,探讨基原对半夏质量与毒性的影响。方法运用酸性染料比色法、酸碱滴定法、HPLC法、经典急毒实验法,进行半夏、水半夏药材中总生物碱,总有机酸,麻黄碱、鸟苷、琥珀酸含量测定及急性毒性比较研究。结果不同基原总生物碱、总有机酸、鸟苷、琥珀酸含量均为半夏〉水半夏,麻黄碱含量为水半夏〉半夏。一日内按最大给药量给小鼠灌胃半夏、水半夏生品混悬液分别为34.8g/kg、38.4g/kg（按含生药量计算）,相当于临床70kg人每公斤体重日用量的270.7倍、298.7倍以上,连续观察14天,小鼠一般状况良好,无一死亡。结论不同基原半夏药材中主要化学成分含量和毒性大小存在显著差异,半夏毒性大于水半夏,且其毒性大小与总生物碱和总有机酸含量存在一定相关性。     

Increase in exhaled nitric oxide and protective role of the nitric oxide system in experimental pulmonary embolism

BACKGROUND AND PURPOSE: Pulmonary embolism (PE) represents a real diagnostic challenge. PE is associated with pulmonary hypertension due to pulmonary vascular obstruction and vasoconstriction. We recently reported that pulmonary gas embolism transiently increases exhaled nitric oxide (FENO), but it is not known whether solid emboli may alter FENO, and whether an intact endogenous NO synthesis has a beneficial effect in experimental solid pulmonary embolism. EXPERIMENTAL APPROACH: We used anaesthetised and ventilated rabbits in these experiments. To mimic PE, a single intravenous infusion of homogenized autologous skeletal muscle tissue (MPE) was given to rabbits with intact NO production (MPE of 60, 15, or 7.5 mg kg(-1); group 1) and to another group (group 2) with inhibited NO synthesis (L-NAME 30 mg kg(-1); MPE of 7.5, 15 or 30 mg kg(-1)). KEY RESULTS: In group 1, after MPE, FENO increased rapidly and dose-dependently and FENO was still significantly elevated after 60 min with the two highest emboli doses. All these animals survived more than 60 min after embolization. In group 2, MPE of 7.5, 15 and 30 mg kg(-1), in combination with NO synthesis inhibition, resulted in 67%, 50% and 25% survival at 60 min respectively, representing a statistically significant decrease in survival. Cardiovascular and blood-gas changes after MPE were intensified by pre-treatment with NO synthesis inhibitor. CONCLUSIONS AND IMPLICATIONS: We conclude that solid PE causes a sustained, dose-dependent increase in FENO, giving FENO a diagnostic potential in PE. Furthermore, intact NO production appears critical for tolerance to acute PE.     

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.     

Simultaneous determination of ephedrine, pseudoephedrine, norephedrine and methylephedrine in Kampo medicines by high-performance liquid chromatography

A simultaneous high-performance liquid chromatographic method for the determination of ephedrine, pseudoephedrine, norephedrine and methylephedrine (ephedrine alkaloids) in Kampo medicines which contain Ephedrae Herba was established. The analysis can be accomplished within 25 min with a Wakosil-II 5C18 HG column by isocratic elution using a mixture of water, acetonitrile and sodium dodecyl sulfate (65:35:0.4) as the mobile phase at a flow-rate of 1.0 ml min(-1), and detection at 210 nm. The detection limits of ephedrine alkaloids are 0.37-1.06 microM per injection (5 microl). This method was applied to analyze 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. The concentration (per Ephedrae Herba gram) of ephedrine alkaloids was higher in the Makyo-kanseki-to decoction than in the others. Calcium sulfate from Gypsum Fibrosum raised ephedrine alkaloids dissolution in the Makyo-kanseki-to decoction.     

The effects of pretreatment with glycyrrhizin and glycyrrhetinic acid on the retrorsine-induced hepatotoxicity in rats.

A wide variety of medicinal herbs contain hepatotoxic pyrrolizidine alkaloids (PAs), and often cause acute and chronic liver damages in man. Liquorice, a known antihepatitis, is commonly used with PA-containing herbs concurrently, and hepatotoxicity induced by such combined uses was not pronounced. The present study is to investigate effects of glycyrrhizin (GL) and 18beta-glycyrrhetinic acid (GA), the major biologically active ingredients of liquorice, against PA-induced hepatotoxicity in rats. Single dose (35 mg/kg, i.p.) of retrorsine (RET), a typical potent hepatotoxic PA, was given to rats to induce liver injury. A single dose pretreatment with GL or GA prior to retrorsine challenge did not show hepatoprotection. However, when rats were pretreated with either GL (200 mg/kg/day, i.p.) or GA (10 mg/kg/day, i.p.) for three consecutive days prior to retrorsine exposure, the elevated serum GOT and GPT levels induced by retrorsine were significantly reduced. Serum levels of transaminases almost returned to normal (GOT: 56+/-2 (control), 104+/-5 (RET), 64+/-3 (GL + RET) and 59+/-3 (GA + RET). GPT: 40+/-2 (control), 90+/-7 (RET), 45+/-2 (GL + RET) and 45+/-4 (GA + RET) SF units/ml). Furthermore, no extensive hepatocellular damages were observed. The results demonstrated that a three-day pretreatment with either GL or GA exhibited protective effect on retrorsine-induced liver damage in rats.     

两种工艺对舒胃颗粒中黄连生物碱含量的影响

目的:比较黄连与其余药材混煎及分煎合并两种工艺对舒胃颗粒中黄连生物碱含量的影响。方法:以黄连生物碱提取量为考察指标,采用紫外分光光度法测定黄连生物碱含量,对两种水提取方法进行比较。结果:按黄连药材投量计,混煎、分煎合并和黄连单煎所得3个样品中生物碱含量以盐酸小檗碱计,分别为3.88%,6.47%,8.37%。结论:分煎合并工艺中黄连生物碱损失量明显少于混煎工艺。     

麻黄中麻黄生物碱的气相色谱测定法

本文应用毛细管气相色谱法,配备氮磷检测器对麻黄中六种生物碱:麻黄碱、伪麻黄碱、去甲麻黄碱、去甲伪麻黄碱、甲基麻黄碱和甲基伪麻黄碱进行分离、测定。对生药样品预处理方法作了较大改进,采用直接碱化醚提法。简化操作步骤。用内标法,线性方程定最测定了国产十二种麻黄,结果与高效液相色谱法基本一致。     

~(60)Co-γ射线辐照贮藏对川贝母质量的影响

目的:研究~(60)Co-γ射线辐照贮藏对川贝母质量的影响。方法:对不同剂量~(60)Co-γ射线辐照处理过的川贝母的霉变、虫蛀情况进行观察,并对辐照和未辐照川贝母的总生物碱和水溶性核苷类成分进行含量比较。结果:经辐照后的川贝母的化学成分、感观形态无明显变化,能有效地防止霉变和虫蛀,明显延长药材的贮藏期;川贝母辐照前后总生物碱和水溶性核苷类成分无含量显著变化。结论:应用~(60)Co-γ射线辐照灭菌杀虫工艺可为富含淀粉及贵重中药材的贮藏提供有效方法。     

不同病因胸腔积液经细管引流前后性质变化特点

摘要：目的 分析不同病因的患者胸腔积液（胸液）引流前后性质及细胞学变化特点。方法 收集天津市胸科 医院呼吸与危重症医学科一病区2016年4月—2017年12月179例中、大量胸液患者资料,根据胸液病理及理化等特 点将其分为结胸组37例、恶性组87例、炎症组39例和心衰组16例。经胸腔置细管引流500~1 000 mL胸液,于引流 前及引流24 h后分别留取标本4 mL,对白蛋白、乳酸脱氢酶（LDH）、白细胞、单核细胞、多核细胞及间皮细胞变化特 点进行统计学分析。结果 结胸组、炎症组、恶性组胸液引流前后基本仍为渗出液。心衰组引流后5例（83.33%）渗 出液变为漏出液。引流前,恶性组、炎症组、心衰组患者胸液白细胞计数均低于结胸组;引流后,该3组仍低于结胸 组,且炎症组、心衰组低于恶性组（P＜0.05）;恶性组引流后白细胞升高（P＜0.05）。引流前,恶性组、炎症组、心衰组 胸液单核细胞计数低于结胸组;引流后,炎症组、心衰组仍低于结胸组;恶性组引流后升高（P＜0.05）。4组患者引流 前、引流后组间,且各组引流前后多核细胞计数差异均无统计学意义（P＞0.05）。引流前,恶性组胸液间皮细胞计数 高于其他3组;引流后,恶性组间皮细胞计数下降,但仍最高（P＜0.05）。结论 慢性充血性心力衰竭伴胸液者引流 前后性质变化较大。肺癌所致恶性胸液者引流前后细胞成分变化较大。对于胸液性质及病理结果的可靠性需结合 标本送检时间谨慎判读。     

Release of prostaglandin E2 from brain of cat: II. In vivo studies on the effects of adrenergic, cholinergic and dopaminergic agonists and antagonists

This study examined the effect of several neurotransmitters, receptor agonists and antagonists on the release of prostaglandin E2 (PGE2) from brain. Levels of PGE2 were measured in consecutive 30-min samples, collected from ventriculocisternal perfusions (100 microliters/min) in chloralose-urethanized cats, before, during and after the addition of carbachol, norepinephrine (NE) and dopamine (DA). Samples were submitted to acid organic extraction and radioimmunoassay (RIA) with a specific antibody. Displacement curves of ligand binding with control and stimulated samples were parallel to those observed with PGE2. The addition of carbachol (10(-4) M) to the perfusate resulted in a significant increase in the levels of PGE2 from 600 +/- 259 pg PGE2/min during the control period up to 1306 +/- 429 pg PGE2/min during the perfusion (P less than 0.02). Though NE (10(-3) M) and isoproterenol (10(-3) M, beta) were without effect, methoxamine (10(-3) M, alpha 1), but not ST-91 (an alpha 2-agonist), produced an increase in secretion of PGE2. This increase was blocked by phentolamine (10(-3) M). The addition of DA (10(-4) M) produced a fall from control levels of 642 +/- 185 pg PGE2/min to 290 +/- 101 pg PGE2/min (P less than 0.02). These results suggest that the central activation of selected populations of receptors will alter basal levels of PGE2 in the extracellular space.     

Induction of alkoxyresorufin O-dealkylases and UDP-glucuronosyl transferase by phenobarbital and 3-methylcholanthrene in primary cultures of porcine ciliary epithelial cells

Our previous studies have shown that drug-metabolizing activities in the eye are highest in the ciliary body, a tissue responsible for aqueous humor production. In this work, we have separated nonpigmented epithelial (NPE) cells and pigmented epithelial (PE) cells from porcine ciliary body and determined basal and induced activities of 7-ethoxyresorufin (ER) O-dealkylase, 7-pentoxyresorufin (PR) O-dealkylase, and UDP-glucuronosyl transferase (UDP-GT) using primary cultures of separated cells. ER and PR activities were associated primarily with NPE cells and were very low in PE cells. Treatment of NPE cells with phenobarbital (PB) for 48 hr resulted in about a 4-fold increase in PR O-dealkylase activity but only a 1.3-fold rise in ER O-dealkylase activity. Conversely, 3-methylcholanthrene (MC) treatment augmented the ER O-dealkylase activity of NPE cells 6 times over the basal activity in 48 hr but had little effect on PR O-dealkylase activity. Both NPE and PE cells had low basal UDP-GT activities. UDP-GT activity increased about 5-fold in PB-treated PE cells and about 4-fold in PB-treated NPE cells in 48 hr. The results of MC treatment were similar to those of PB treatment; enhancement of UDP-GT was more pronounced in PE cells than in NPE cells. Induction by PB and MC of ER O-dealkylase, PR O-dealkylase and UDP-GT activities in ciliary NPE and PE cells was inhibited almost completely by 3.5 microM cyclohexamide and 40 nM actinomycin D. The heterogeneous distribution of these enzymes suggests that a harmonious interplay between NPE and PE cells is important for metabolic detoxification of blood plasma prior to aqueous humor formation.     

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。     

Caffeine-herbal ephedra combination increases resting energy expenditure, heart rate and blood pressure

1. The purpose of the present study was to determine whether the consumption of an acute dose of caffeine and Ma Huang increases resting energy expenditure (REE), heart rate (HR) and blood pressure (BP) over a 3 h period. 2. A randomized, double-blind cross-over study was performed evaluating the acute effects of caffeine (150 mg)/herbal ephedra (Ma Huang; 20 mg ephedra alkaloids) versus a placebo. A total of eight healthy subjects (four males and four females) with a mean (+/-SD) age of 23.4+/-0.8 years (mean ages for males and females: 25.3+/-0.7 and 22.0+/-0.7 years, respectively) and 22.5+/-3.1% body fat (15.7+/-1.2 and 27.6+/-3.5% body fat for males and females, respectively) were recruited to the study. Participants were moderate caffeine users (approximately 150-300 mg/day). 3. Subjects reported to the laboratory following a 12 h fast and 48 h of a caffeine-free diet. Resting energy expenditure was measured prior to supplementation and for 15 min every 30 min for 3 h following supplementation. Heart rate and BP were obtained every 15 min. Blood samples were obtained every 30 min following the measurement of REE and analysed for caffeine, ephedrine, free fatty acids and glucose. 4. By 3 h, HR was 22.7+/-5.5% higher (P<0.05) than baseline for the caffeine/ephedra trial compared with 8.9+/-2.2% higher for the placebo group. At 3 h, systolic BP was 9.1+/-2.2% higher (P<0.05) than baseline for the caffeine/ephedra trial compared with only 1.9+/-2.9% different from baseline for the placebo trial. There was no effect of the caffeine/ephedra combination on diastolic BP. Resting energy expenditure during the last 30 min was 4.5+/-2.5% higher in the placebo trial and 10.7+/-2.5% higher (P<0.05) in the caffeine/ephedra trial; REE was 8.5 +/- 2.0% higher (P<0.05) in the caffeine/ephedra trial compared with the placebo trial. Free fatty acids increased over time in the placebo and caffeine/ephedra trials (from 0.5+/-0.05 to 0.63+/-0.05 mEq/L and from 0.48+/-0.06L to 0.8+/-0.05 mEq/L, respectively). 5. Caffeine and herbal ephedra, at doses of 150 mg and 20 mg (ephedrine), respectively, result in a significant elevation in REE, HR and BP. Although significant, the increase in energy expenditure is negligible in terms of weight loss.