槲皮素对葡萄糖-6-磷酸脱氢酶缺乏者红细胞的体外氧化作用

目的探讨槲皮素对葡萄糖-6-磷酸脱氢酶(G6PD)缺乏者红细胞氧化还原状态的影响。方法分别在体外将低、中、高浓度的槲皮素与G6PD缺乏者及正常者的40%红细胞悬液和全血进行孵育,然后测定红细胞还原性谷胱甘肽(GSH))及高铁血红蛋白(MetHb)水平的变化。结果槲皮素具有较强氧化作用,能明显降低G6PD缺乏者红细胞GSH水平,升高MetHb,且这种氧化作用呈一定浓度依赖性,在中、高浓度表现明显。结论槲皮素对G6PD缺乏者红细胞具有氧化作用,G6PD缺乏者应慎用含槲皮素成分的食物、中草药及制剂。     

染料木素等5种黄酮类物质对小鼠血清尿酸水平及黄嘌呤氧化酶活性的影响

目的探讨黄酮类化合物染料木素、芹菜素、槲皮素、芦丁和落新妇苷体外对黄嘌呤氧化酶活性的影响,对正常小鼠血清和肝脏黄嘌呤氧化酶活性的影响,同时评价对小鼠血清尿酸水平的作用。方法采用改良的紫外分光光度法测定染料木素、芹菜素、槲皮素、芦丁和落新妇苷体外对黄嘌呤氧化酶的抑制作用,采用分光光度法研究对小鼠血清和肝脏黄嘌呤氧化酶活性的影响,以磷钨酸法测定对小鼠血清尿酸水平的作用。结果体外实验表明黄酮类化合物染料木素、芹菜素、槲皮素、芦丁和落新妇苷体外对黄嘌呤氧化酶活性无明显影响。体内实验观察到这5种黄酮类化合物能够显著升高或降低黄嘌呤氧化酶的活性;而且,血清尿酸水平与血清黄嘌呤氧化酶活性密切相关,与肝脏黄嘌呤氧化酶活性无明显关联。用这些黄酮类化合物给药的小鼠血清尿酸水平都高于正常对照组。结论这5种黄酮类化合物不能够作为替代别嘌醇的药物用来降低血清尿酸水平。     

女贞花的化学成分研究

目的 研究女贞花蕾中的化学成分.方法 提取和分离纯化女贞花的化学成分,并通过波谱分析鉴定化合物的结构.结果 从女贞花的丙酮提物中分离出12个化合物,分别是:β-谷甾醇(Ⅰ)、胡萝卜苷(Ⅱ)、麦角甾-7,22-二烯-3β,5α,6β-三醇(Ⅲ)、柚皮素(Ⅳ)、木犀草素(Ⅴ)、芹菜素(Ⅵ)、槲皮素(Ⅶ)、芹菜素7-O-β-...     

黄酮类化合物对耐药金黄葡萄球菌mecA基因的影响

目的:研究黄酮类化合物对人工诱导的耐药金黄葡萄球菌的耐药基因mec A的mRNA表达水平的影响。方法:采用纸片法测定黄酮类化合物对耐药金黄葡萄球菌的逆转作用;利用RT-PCR法测定耐药基因mec A的相对表达量。结果:耐药金黄葡萄球菌分别被芦丁、姜黄素、木犀草素、槲皮素和芹菜素作用后,耐药基因mec A的相对表达量分别为0.55,0.79,0.76,0.92,和0.84。结论:黄酮类化合物能降低金黄葡萄球菌的耐药基因mec A的表达,能对抗金黄葡萄球菌的耐药性。     

黄酮类化合物抑制法舒地尔代谢的体外研究

目的研究13种黄酮类化合物对法舒地尔代谢的抑制作用,为法舒地尔的临床安全合理用药提供参考。方法采用人肝细胞溶质体外孵育法,在孵育体系中加入底物法舒地尔和系列浓度的黄酮类化合物进行共孵育,然后采用LC-MS/MS法测定孵育液中法舒地尔的代谢物羟基法舒地尔的浓度,通过与对照组比较,确定各种黄酮类化合物对法舒地尔代谢的抑制程度,并采用GraphPad Prism 6.0软件计算各化合物的IC50值。结果槲皮素、高良姜素、山奈酚、芦丁、柚皮素、橙皮素、乔松素、表儿茶素、金合欢素及黄豆苷元对法舒地尔均有较强的代谢抑制作用,其IC50值均低于1μmol·L-1;蒙花苷、黄芩苷及灯盏花乙素对法舒地尔代谢的抑制程度较弱。结论 13种黄酮类化合物对法舒地尔代谢为羟基法舒地尔的肝酶活性均有不同程度的抑制作用,槲皮素和高良姜素的抑制程度最高。临床上使用法舒地尔时应关注与富含黄酮的中药或食物间可能发生的代谢性相互作用。     

基于PLS-DA法评价60Co-γ射线对刘寄奴中有效成分的影响

摘要：目的:考察60Co-γ射线辐照灭菌对刘寄奴中6个成分(槲皮素、柚皮素、木犀草素、芹菜素、山奈酚和异泽兰黄素)含量的影响。方法:采用Syncronis C18色谱柱(250 mm×4.6 mm, 5μm),流动相为乙腈-0.2%磷酸水溶液,梯度洗脱,检测波长260 nm,流速为1.0 ml·min-1,柱温为30℃。分别选择剂量为0,3,6,9 kGy对刘寄奴药材进行辐照,比较辐照前后活性成分含量变化,采用偏最小二乘-判别分析(PLS-DA)对其总体质量进行分析评价。结果:刘寄奴药材中槲皮素、柚皮素、木犀草素、芹菜素、山奈酚和异泽兰黄素分别在0.051 0～1.018 0 mg·ml-1、0.021 0～0.427 0 mg·ml-1、0.007 0～0.140 0 mg·ml-1、0.003 0～0.068 0 mg·ml-1、0.005 0～0.108 0 mg·ml-1、0.028 0～0.563 0 mg·ml-1范围内呈良好线性,平均加样回收率分别为99.7%,99.7%,99.3%,98.4%,98.4%和99.8%;辐照剂量不超过6 kGy时,各组分含量变化无统计学意义(P>0.05)。PLS-DA筛选出导致质量差异的3个差异标志物。结论:为保证药品安全有效,采用60Co-γ射线对刘寄奴药材进行灭菌时,辐照剂量不宜超过6 kGy。     

半枝莲的化学成分研究（Ⅰ）

目的研究半枝莲（Scutellaria barbaraD．Don）的化学成分。方法从半枝莲全草的乙醇提取液中分离得到9个化合物。利用理化性质和波谱学分析，鉴定它们的化学结构。结果分离鉴定了9个化合物，分别是6-羟基香豆素（1）、香草酸（2）、异香草酸（3）、4＇-羟基汉黄芩素（4）、6-甲氧基柚皮素（5）、芹菜素（6）、柚皮素（7）、木犀草素（8）、异红花素（9）。结论化合物1为首次从该属植物中分得的已知化合物，化合物2、3为首次从该植物中分得的已知化合物。     

多刺绿绒蒿的化学成分

从多刺绿绒蒿Meconopsis horridula Hook.f.et Thoms的全草中分离鉴定了13个化合物,分别为:木犀草素-7-O-β-D-葡萄糖苷(1)、山奈酚-3-O-β-D-葡萄糖( 1→2)-β-D-葡萄糖苷(2)、槲皮素-3-O-β-D-半乳糖(1→6)-β-D-葡萄糖苷(3)、小麦黄素-7-O-β-D-葡萄糖苷(4)、山奈酚-3-O-β-D-葡萄糖(5)、桂皮酰胺(6)、对羟基桂皮酰胺对羟基苯乙胺(7)、小麦黄素(8)、木犀草素(9)、槲皮素(10)、芹菜素(11)、山奈酚-4'-甲醚(12)、山奈酚(13).其中化合物1、5、6、7、12、13为首次从该属中分离得到,化合物2、3、4为首次从该植物中分离得到.     

染料木素、芹菜素、槲皮素、芦丁和落新妇苷对高尿酸血症小鼠黄嘌呤氧化酶活性及血清尿酸水平的影响

目的探讨黄酮类化合物染料木素、芹菜素、槲皮素、芦丁和落新妇苷体外对黄嘌呤氧化酶活性的影响,对高尿酸血症小鼠血清和肝脏黄嘌呤氧化酶活性的影响,同时评价对小鼠血清尿酸水平的作用。方法采用改良的紫外分光光度法测定染料木素、芹菜素、槲皮素、芦丁和落新妇苷体外对黄嘌呤氧化酶的抑制作用;采用尿酸酶抑制剂氧嗪酸钾诱导小鼠高尿酸血症模型,以分光光度法研究对小鼠血清和肝脏黄嘌呤氧化酶活性的影响,以磷钨酸法测定对小鼠血清尿酸水平的作用。结果体外实验表明这些黄酮类化合物对黄嘌呤氧化酶活性无明显影响。然而,体内实验观察到能够明显升高或降低黄嘌呤氧化酶的活性,而且,血清尿酸水平与血清黄嘌呤氧化酶活性密切相关,与肝脏黄嘌呤氧化酶活性无明显关联。该研究表明用这些黄酮类化合物给药的小鼠血清尿酸水平都高于正常对照组。结论这5种黄酮类化合物不能够作为替代别嘌醇的药物用来降低血清尿酸水平。     

齿叶白鹃梅叶中黄酮类成分的分离与结构鉴定

目的研究齿叶白鹃梅叶的化学成分。方法采用硅胶柱色谱、重结晶和Sephadex LH-20凝胶柱色谱的方法进行化学成分的分离和纯化,根据化合物的理化性质和波谱数据鉴定其结构。结果分离得到6个黄酮类化合物,经鉴定为芹菜素(apigenin1,),木犀草素(luteolin,2),槲皮素(querce-tin,3),槲皮素-3-O-β-D-葡萄糖苷(quercetin-3-O-β-D-glucopyranoside,4),芹菜素-7-O-β-D-葡萄糖苷(apigenin-7-O-β-D-glucopyranoside,5),芹菜素-7-O-β-D-新橙皮糖苷(apigenin-7-O-β-D-neosphero-side,6)。结论化合物1-6均首次从此植物中分离得到。     

Cytochrome P450-dependent toxic effects of primaquine on human erythrocytes

Primaquine, an 8-aminoquinoline, is the drug of choice for radical cure of relapsing malaria. Use of primaquine is limited due to its hemotoxicity, particularly in populations with glucose-6-phosphate dehydrogenase deficiency [G6PD(−)]. Biotransformation appears to be central to the anti-infective and hematological toxicities of primaquine, but the mechanisms are still not well understood. Metabolic studies with primaquine have been hampered due to the reactive nature of potential hemotoxic metabolites. An in vitro metabolism-linked hemotoxicity assay has been developed. Co-incubation of the drug with normal or G6PD(−) erythrocytes, microsomes or recombinant cytochrome P₄₅₀ (CYP) isoforms has allowed in situ generation of potential hemotoxic metabolite(s), which interact with the erythrocytes to generate hemotoxicity. Methemoglobin formation, real-time generation of reactive oxygen intermediates (ROIs) and depletion of reactive thiols were monitored as multiple biochemical end points for hemotoxicity. Primaquine alone did not produce any hemotoxicity, while a robust increase was observed in methemoglobin formation and generation of ROIs by primaquine in the presence of human or mouse liver microsomes. Multiple CYP isoforms (CYP2E1, CYP2B6, CYP1A2, CYP2D6 and CYP3A4) variably contributed to the hemotoxicity of primaquine. This was further confirmed by significant inhibition of primaquine hemotoxicity by the selective CYP inhibitors, namely thiotepa (CYP2B6), fluoxetine (CYP2D6) and troleandomycin (CYP3A4). Primaquine caused similar methemoglobin formation in G6PD(−) and normal human erythrocytes. However, G6PD(−) erythrocytes suffered higher oxidative stress and depletion of thiols than normal erythrocytes due to primaquine toxicity. The results provide significant insights regarding CYP isoforms contributing to hemotoxicity and may be useful in controlling toxicity of primaquine to increase its therapeutic utility.     

Understanding the mechanisms for metabolism-linked hemolytic toxicity of primaquine against glucose 6-phosphate dehydrogenase deficient human erythrocytes: evaluation of eryptotic pathway

Therapeutic utility of primaquine, an 8-aminoquinoline antimalarial drug, has been limited due to its hemolytic toxicity in population with glucose 6-phosphate dehydrogenase deficiency. Recent investigations at our lab have shown that the metabolites generated through cytochrome P(450)-dependent metabolic reactions are responsible for hemotoxic effects of primaquine, which could be monitored with accumulation of methemoglobin and increased oxidative stress. The molecular markers for succeeding cascade of events associated with early clearance of the erythrocytes from the circulation were evaluated for understanding the mechanism for hemolytic toxicity of primaquine. Primaquine alone though did not induce noticeable methemoglobin accumulation, but produced significant oxidative stress, which was higher in G6PD-deficient than in normal erythrocytes. Primaquine, presumably through redox active hemotoxic metabolites generated in situ in human liver microsomal metabolism-linked assay, induced a dose-dependent methemoglobin accumulation and oxidative stress, which were almost similar in normal and G6PD-deficient erythrocytes. Primaquine alone or in presence of pooled human liver microsomes neither produced significant effect on intraerythrocytic calcium levels nor affected the phosphatidyl serine asymmetry of the normal and G6PD-deficient human erythrocytes as monitored flowcytometrically with Annexin V binding assay. The studies suggest that eryptosis mechanisms are not involved in accelerated removal of erythrocytes due to hemolytic toxicity of primaquine.     

Inhibition effects of some antidepressant drugs on pentose phosphate pathway enzymes

The glucose metabolism in the pentose cycle is essential to the source of NADPH. Deficiency of these enzymes have been linked to depression and psychotic disorders. Depression is an increasingly prevalent mental disorder which may cause loss of labor. Antidepressant drugs are commonly employed in treatments of mood disorders and anxiety treatment. The purpose of this study is to investigate the effects of aripiprazole, mirtazapine, risperidone, escitalopram and haloperidol on the activity of 6-phosphogluconate dehydrogenase (6PGD) and glucose-6-phosphate dehydrogenase (G6PD) enzymes purified from human erythrocytes. It was found that aripiprazole, mirtazapine, risperidone, escitalopram and haloperidol show effective inhibitor properties on purified G6PD and 6PGD enzymes. The IC₅₀ values of these drugs were found in the range of 26.34 μM-5.78 mM for 6PGD and 16.26 μM-3.85 mM for G6PD. The K_i values of the drugs were found in the range of 30.21 ± 4.31 μM-4.51 ± 1.83 mM for 6PGD and 14.12 ± 3.48 μM-4.98 ± 1.14 mM for G6PD. Usage of drugs with significant biological effects may be a hazard in some conditions.     

Effects of Some Antibiotics on Activity of Glucose-6-Phosphate Dehydrogenase from Human Erythrocytes In Vitro and Effect of Isepamicin Sulfate on Activities of Antioxidant Enzymes in Rat Erythrocytes

The purpose of this study was to investigate effects of some antibiotics on glucose-6-phosphate dehydrogenase (G6PD), antioxidant enzymes, and malondialdehyde (MDA). Initially, for in vitro studies, G6PD was purified from human erythrocyte, 9811-fold in a yield of 42.4% by using ammonium sulfate precipitation and 2′,5′ ADP-Sepharose 4B affinity gel. The purified enzyme showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The effects of four different antibiotics (isepamicin sulfate, meropenem, chloramphenicol, and thiamphenicol glisinat hydrochloride) were investigated on the purified enzyme. K_i value and type of inhibition were determined by means of Lineweaver–Burk graphs and regression analysis graphs. Isepamicin sulfate inhibited the enzyme activity (I₅₀ value, 2.1 mM; K_i value, 1.7 mM), whereas thiamphenicol glisinat hydrochloride activated the G6PD dose dependently. Other drugs showed no inhibition and activation effect. In addition, the effects of isepamicin sulfate on the activities of G6PD, glutathione reductase (GR), superoxide dismutases (SOD), glutathione peroxidase (GPx), catalase (CAT), and glutathione S-transferase (GST) and MDA contentrations were examined in Sprague-Dawley rat erythrocytes in vivo. A marked alteration in the activities of these enzymes and MDA levels may be the result of oxidative stress in the rats receiving isepamicin sulfate.     

Pharmacokinetics of morphine-6-glucuronide following oral administration in healthy volunteers

Aim After oral administration, morphine-6-glucuronide (M6G) displays an atypical absorption profile with two peak plasma concentrations. A proposed explanation is that M6G is hydrolysed to morphine in the colon, which is then absorbed and subsequently undergoes metabolism in the liver to morphine-3-glucuronide (M3G) and M6G. The aims of this study were to confirm and elucidate the biphasic absorption profile as well as clarify the conversion of M6G to morphine after a single oral administration of M6G in healthy volunteers. Methods The study was conducted accordingly to a nonblinded, randomised, balanced three-way crossover design in eight healthy male subjects. The subjects received 200 mg oral M6G, 50 mg oral M6G and 30 mg oral morphine. Blood samples were collected until 72 h after M6G administration and until 9 h after morphine administration. Paracetamol and sulfasalazine were coadministered with M6G as markers for the gut contents reaching the duodenum and colon, respectively. Results The plasma concentration peaks of M6G were seen at 4.0 (2.0–6.0) and 18 (12.0–24.0) h after 200 mg M6G and at 3.5 (2.0–6.0) and 21.3 (10.0–23.3) h after 50 mg M6G, which was in agreement with previously published results. The K_{M6G_abs}/K_{M6G_M6G} ratio was found to be 10. Conclusion The pharmacokinetic profile of M6G after oral administration was confirmed and with the presence of M3G and morphine in plasma after oral administration of M6G, proof seems to be found of the constant and prolonged absorption of M6G. The K_{M6G_abs}/K_{M6G_M6G} ratio of 10 indicates that the second absorption peak of M6G consists of approximately 10 times more absorbed M6G than reglucuronidated M6G. However, further studies are required to determine the precise kinetics of the second absorption peak.     

Glucose 6-phosphate dehydrogenase: in vitro and in vivo effects of dantrolene sodium

In our study, effects of dantrolene sodium on glucose 6-phosphate dehydrogenase (G6PD) were examined in the human erythrocytes in vitro and in rat erythrocytes in vivo. Human erythrocyte G6PD was purified using ammonium sulfate fractionation and 2',5'-ADP Sepharose 4B affinity chromatography. The enzyme activity was determined by Beutler's method. The overall purification procedures gave the human G6PD having the specific activity of 97.6 EU/mg of protein, which was purified 9760-fold with a yield of 39%. Dantrolene sodium inhibited the enzyme activity under in vitro conditions and the I(50) value (drug concentration which produces 50% inhibition) of this drug was 0.91 mM. In vivo studies were performed in rats (Sprague-Dawley). Dantrolene sodium at 10 mg/kg inhibited the enzyme activity significantly (p < 0.05) 3 h after dosing. We conclude that dantrolene sodium showed inhibitory effect on G6PD activity both in vitro and in vivo.     

If Vitamins Could Kill: Massive Hemolysis Following Naturopathic Vitamin Infusion

Introduction

Hemolysis from naturopathic remedies remains poorly reported in the medical literature, although it is most commonly noted in the patients with glucose 6-phosphate dehydrogenase (G6PD) deficiency. We report a case of massive intravascular hemolysis following the infusion of a naturopathic preparation that contains vitamins.

Case Report

A 47-year-old African-American man presented to the hospital with 3 days of fever, dyspnea, emesis, dark urine, and progressive confusion. His symptoms began 1 day following an infusion of a vitamin complex. His physical examination was significant for lethargy and scleral icterus. Initial laboratory studies were notable for anemia (hemoglobin, 3.3 g/dL and hematocrit, 11%), brisk reticulocytosis (33%), acute renal insufficiency (creatinine, 2.8 mg/dL), and indirect hyperbilirubinemia (total bilirubin, 4.4 mg/dL). His peripheral smear demonstrated "blister cells," erythrocytes that have been left devoid of precipitated hemoglobin by the spleen, which are commonly seen in patients with G6PD deficiency. His physician revealed that the infusion contained vitamins B and D complex, free amino acids, magnesium, and taurine. The patient clinically improved and was discharged to home. G6PD concentration was significantly reduced to 4.7 U/g Hb upon recovery.

Discussion

Life-threatening intravascular hemolysis may occur following a naturopathic vitamin infusion and may identify previously unknown G6PD deficiency. Since most properly formulated naturopathic treatments have few toxic ingredients, the possibilities of improper formulation, toxic diluents, or contaminants should be considered. Inadequate regulatory oversight of naturopathic remedies has the potential to allow serious toxicity especially in genetically predisposed individuals.     

亨氏小体和G6PD检测对地中海贫血筛查的应用价值

目的评价亨氏小体和葡萄糖6磷酸脱氢酶（G6PD）检测在地中海贫血筛查中的应用价值,为临床寻找一种更理想的地贫筛查的检验方法。方法在血红蛋白电泳、红细胞平均体积（MCV）、红细胞渗透脆性试验（脆性试验）的基础上增加亨氏小体和G6PD检测。结果用血红蛋白电泳、MCV、脆性试验联合检测对轻型α-地贫的准确度为90．5％,用Hh电泳、MCV、脆性试验、亨氏小体、G6PD对轻型α-地贫的准确度为96．8％,两项检测差异有显著性。结论血红蛋白电泳、MCV、脆性试验、亨氏小体、G6PD联合测定是地贫筛查的较理想的试验方法。     

Effects of antiemetic drugs on glucose 6-phosphate dehydrogenase and some antioxidant enzymes.

The aim of this study was to investigate effect of antiemetics on G6PD and antioxidant enzymes. Antiemetics are currently being used to reduce or prevent nausea and vomiting in patients. This is the first study to show effect of antiemetics on glucose-6-phosphate dehydrogenase (G6PD) and antioxidant enzyme activities. For in vitro studies, G6PD was purified from human erythrocyte, 10, 26-fold in a yield of 51.3% by using ammonium sulphate precipitation and 2',5'-ADP-Sepharose 4B affinity gel. The purified enzyme showed a single band on sodium dodecyl sulfate-polyacrilamide gel electrophoresis (SDS-PAGE). The effects of four different antiemetics (granisetron hydrochloride, ondansetron hydrochloride, metoclopramide hydrochloride, trimethobenzamide hydrochloride) were investigated on the purified enzyme. Granisetron hydrochloride and ondansetron hydrochloride inhibited the enzyme activity (Ki values; 5.05 mM and 0.034 mM, I50 values; 3.9 mM and 0.036 mM, respectively). Metoclopramide hydrochloride, trimethobenzamide hydrochloride showed no inhibition effects. In addition, in vivo studies, effects of ondansetron hydrochloride on the G6PD, glutathione reductase (GR), glutathione peroxidase (GPx) and catalase (CAT) were examined in the rat erythrocytes. G6PD (49% of control), GR (55% of control), CAT (60% of control) activities in erythrocytes were significantly decreased whereas GPx (183% of control) was significantly increased. A marked alteration in these enzymes may be result of oxidative stress in the rats receiving ondansetron hydrochloride.     

Effects of melatonin on enzyme activities of glucose-6-phosphate dehydrogenase from human erythrocytes in vitro and from rat erythrocytes in vivo.

The in vivo and in vitro effects of melatonin on glucose-6-phosphate dehydrogenase (G6PD) from human erythrocytes have been investigated. For this purpose, human erythrocyte glucose-6-phosphate dehydrogenase was purified, at the beginning, 13.654 times in a yield of 28% by using ammonium sulphate precipitation and 2',5'-ADP Sepharose 4B affinity gel. A temperature of +4 degrees C was maintained during the purification process. Enzyme activity was determined by the Beutler method using a spectrophotometer at 340 nm. This method was utilized for all kinetic studies. For in vitro experiments, the enzyme activity increased below 0.08 mM melatonin concentration and reached a plateau above 0.1 mM. Ten mg kg(-1)melatonin was administered intraperitonally and indicated the stimulatory effect on the enzyme. Time-dependent in vivo studies were executed for melatonin in Sprague-Dawley type rats. It was found that G6PD activity in the erythrocytes was increased by the melatonin in 1.5 and 3.5 h. These results show that both in vitro(below 0.08 mM) and in vivo pharmacological levels of melatonin increased enzyme activity in erythrocytes. The findings also indicate that melatonin may be pharmacologically useful in patients where a deficiency of the enzyme in red blood cells (RBC) causes haemolytic anaemia.