贯叶金丝桃止痛酊的制备及质量控制

目的：研制贯叶金丝桃止痛酊，并建立其质量标准。方法：用70％乙醇提取贯叶金丝桃，配以丁香油、薄荷油及氮酮制成酊剂，测定pH值及鉴别，并用可见紫外分光光度法测定含量。结果：该制剂1mL含总金丝桃素0．27mg，平均加样回收率100．06％。结论：本制剂制备工艺简单，质量控制方法简便、可靠。     

不同方法除鞣质后贯叶金丝桃提取物中金丝桃素的含量变化

目的 考察用不同方法检测除鞣质后金丝桃素的含量变化和收率,建立贯叶金丝桃良好的除鞣质方法。方法采用明胶沉淀法、改良明胶沉淀法及碱性醇沉法除鞣质后,用高效液相色谱（HPLC）法测定贯叶金丝桃提取物中金丝桃素含量变化和收率;色谱柱为Phenomenex—C18柱（250mm×4．6mm,5μm）,流动相为甲醇-0．006mol／LNa214PO4（7：1V／V,H3P04调至pH=6．5）,流速为1．0mL／min,柱温30℃,检测波长590nm,外标法计算。结果金丝桃素的进样量线性范围为0．0194～0．7760μg（r=0．9999）,平均回收率为100．25％。RSD：1．29％（n=5）;明胶沉淀法、改良明胶沉淀法和碱性醇沉法除鞣质后金丝桃素含量分别为0．092％,0．098％和0．093％,收率分别为70．15％,85．21％和89．16％。结论改良明胶沉淀法具有鞣质去除完全、金丝桃素含量较高、损失较少的优点,且方法简便易行．可应用于贯叶金丝桃提取物除鞣质处理。     

HPLC法测定不同产地黄蜀葵花中金丝桃苷含量

目的分析并比较了安徽四个不同产地黄蜀葵花药材中金丝桃苷的含量。方法高效液相色谱法,色谱柱Alltima C18（5μm,4．0mm×250mm）,流动相：乙腈-水-磷酸（175：825：1）,流速1．0ml·min^-1,检测波长：360nm,进样量：20μl,柱温为40℃。结果黄山黄蜀葵花中金丝桃苷含量1．18％,RSD为2．54％;合肥黄蜀葵花中金丝桃苷含量1．07％,RSD为1．62％;六安黄蜀葵花中金丝桃苷含量1．03％,RSD为1．94％;毫州黄蜀葵花中金丝桃苷含量0．92％,RSD为1.88％。结论HPLC法测得黄山黄蜀葵花中金丝桃苷含量最高,毫州黄蜀葵花中金丝桃苷含量最低,该方法简便准确,重现性好,为各地黄蜀葵花药材的进一步开发利用提供质量控制依据。     

HPLC测定不同产地和不同部位贯叶金丝桃中的金丝桃素

目的 测定自四川、湖北、甘肃、陕西等4省48个样地的贯叶金丝桃中金丝桃素的含量,为评价药材质量提供依据.方法 采用HPLC方法,色谱柱为Agilent zorbax extend-C18柱(250 mm×4.6 mm,5μm),流动相为甲醇-6 mmol·L-1磷酸氢二钠(87.5∶12.5,用磷酸调pH6.5),检测波长为590 nm,柱温30℃,流速1 mL· min-,进样量10 μL.结果 金丝桃素4.04～252.5 ng与峰面积之间的线性良好(r2 =0.9996);回收率为99.69％,RSD=1.28％ (n =9).不同产地贯叶金丝桃中金丝桃素的含量为0.186 ～0.382 mg·g-1,不同部位中的含量为花＞叶＞茎;光照在提取过程中对金丝桃素检测结果的影响较大.结论 不同产地贯叶金丝桃的不同部位中金丝桃素的含量均不同.     

秦岭三种金丝桃属植物药中金丝桃甙的含量测定

用薄层一紫外分光光度法测定了秦岭地区三种金丝桃属植物药贯叶连翘、红旱莲、大对经草中金丝桃甙的含量。结果以红旱莲含金丝桃甙为最高，达１．１７２％。     

正交试验优选贯叶连翘中金丝桃素超声波提取工艺

目的：优选贯叶连翘中金丝桃素的超声波提取工艺。方法：采用超声波提取技术,以正交试验设计,考察溶剂种类、溶剂浓度、料液倍量比、超声波功率、单次辐射时间和提取时间对提取效果的影响。结果：优选出的工艺条件为80%乙醇,料液比1∶10（w/v）,提取功率400 W,单次辐射3 s,每次提取20 min。在最优条件下,金丝桃素提取率为（96.04±0.25）%,得到浸膏中金丝桃素含量为0.31%。结论：本工艺简单、经济、提取率高,浸膏中金丝桃素含量符合要求。     

紫外分光光度法测定贯叶金丝桃胶囊中金丝桃素的含量

目的：建立制剂贯叶金丝桃胶囊中金丝桃素的含量测定方法。方法：采用紫外分光光度法，在590nm波长处检测。结果：金丝桃素在3.29—16.49mg.L^-1(r＝0．9999)范围内吸收值与其浓度呈良好的线性关系，方法平均回收率99．54％，RSD为0.44％(n＝6)。结论：该方法简便、快速、准确，可作为本制剂的质量控制方法。     

贯叶连翘提取物中金丝桃苷的HPLC测定

建立了HPLC法测定贯叶连翘提取物中金丝桃苷的含量。采用C18柱，以甲醇-0．025mol／L磷酸溶液（50：50）为流动相，检测波长363nm。金丝桃苷在0．25～2．5μg／ml范围内线性关系良好。平均同收率为101．4％，RSD为0．66％。     

HPLC法测定雷公藤粗制品中雷公藤甲素的含量

目的：建立高效液相色谱法测定雷公藤粗制品中雷公藤甲素的含量。方法：雷公藤粗制品经洗脱提取后，采用十八烷基键合硅胶柱，以乙腈-水（30：70）为流动相，以218nm为检测波长，检测雷公藤粗制品中雷公藤甲素的含量。结果：雷公藤甲素在1．92～11．52μg／ml范围内线性良好，r=0．9999，平均回收率为95．32％，RSD为1．16％。结论：本法操作简便、结果准确，可用于雷公藤粗制品中雷公藤甲素的含量测定。     

不同野菊花提取物中蒙花苷和木犀草素的含量测定比较

目的：通过比较不同的提取方法对野菊花提取物中蒙花苷和木犀草素含量测定的影响，研究中药提取新工艺。方法：采用加热同流提取法、超声波提取法和组织破碎提取法3种提取工艺，同时提取野菊花中蒙花苷和木犀草素；利用RP—HPLC法，以水-乙腈-4％醋酸溶液为流动相（流速1．0mL·min^-1）同时测定蒙花苷和木犀草素含量。结果：蒙花苷和木犀草素分别在0．048～0．240mg·mL^-1（r〉0．9995）、0．0059～0．0294mg·mL^-1（r〉0．9999）浓度范围内线性关系良好，加热回流法、组织破碎法提取的蒙花苷和木犀草素含量均高于超声法，组织破碎法较回流法操作简便、节能节时，且可避免高温对热敏性物质的破坏性，表现出较高优越性。结论：组织破碎提取法有着高效经济的优势，是一种值得推广的高新提取工艺。     

Clinic at the Health Food Store? Employee Recommendations and Product Analysis

STUDY OBJECTIVES: To determine what products health food store employees recommend for depression, to analyze the content of these products based on label claims, and to evaluate employee statements or recommendations for accuracy and safety. METHODS: Twelve health food stores were selected for the study. One investigator approached an employee in each store and asked what they recommended for depression plus five additional questions regarding product use. Thirteen products containing St. John's wort were purchased and analyzed for hypericin and pseudohypericin content using high-performance liquid chromatography (HPLC). Total hypericin content was calculated by adding the values for hypericin and pseudohypericin. RESULTS: All 12 health food store employees recommended a St. John's wort supplement for treatment of depression. Furthermore, numerous comments made by employees regarding St. John's wort and the treatment of depression were unsafe and inaccurate. The HPLC analysis revealed that no product contained +/- 10% of the stated label claim for hypericin content, and two products contained 0% hypericin. The total hypericin content (hypericin plus pseudohypericin) of only two products was within +/- 10% of the label claim for hypericin. CONCLUSIONS: Health food store employees offer health care advice regarding treatment of depression with dietary supplements without proper scientific and medical training. Their comments could cause significant harm to customers. In addition, the inconsistencies of dietary supplement content continue to raise concern for individuals who use these agents as medical treatment.     

PKC-Mediated Potentiation of Morphine Analgesia by St. John’s Wort in Rodents and Humans

Our purpose was to combine the use of morphine with clinically available inhibitors of protein kinase C (PKC), finally potentiating morphine analgesia in humans. Thermal tests were performed in rodents and humans previously administered with acute or chronic morphine combined or not with increasing doses of the PKC-blocker St. John’s Wort (SJW) or its main component hypericin. Phosphorylation of the γ subunit of PKC enzyme was assayed by western blotting in the periaqueductal grey matter (PAG) from rodents co-administered with morphine and hypericin and was prevented in rodent PAG by SJW or hypericin co-administration with morphine, inducing a potentiation of morphine analgesia in thermal pain. The score of pain assessment in healthy volunteers were decreased by 40% when morphine was co-administered with SJW at a dose largely below those used to obtain an antidepressant or analgesic effect in both rodents and humans. The SJW/hypericin potentiating effect lasted in time and preserved morphine analgesia in tolerant mice. Our findings indicate that, in clinical practice, SJW could reduce the dose of morphine obtaining the same analgesic effect. Therefore, SJW and one of its main components, hypericin, appear ideal to potentiate morphine-induced analgesia.     

黑龙江省两种金丝桃属植物理化鉴别特征

目的：为了探讨黑龙江省两种金丝桃属植物长柱金丝桃(Hypericum ascyron L．)、乌腺金丝桃(H．attenuatum Choisy．)理化鉴别特征。方法：分别采用黄酮、皂苷、鞣质及金丝桃素的鉴别试验。结果：黄酮定性反应均为阳性；皂苷定性反应表明均含三萜皂苷；鞣质的定性反应表明它们所含鞣质为缩合鞣质；金丝桃素的定性反应表明乌腺金丝桃含金丝桃素。结论：二者理化鉴别特征有一定差别．能通过化学鉴别方法予以鉴别。     

Single pre-treatment with hypericin,a St. John’s wort secondary metabolite,attenuates cisplatin- and mitoxantrone-induced cell death in A2780, A2780cis and HL-60 cells

St. John’s wort (SJW, Hypericum perforatum L.) is a commonly used natural antidepressant responsible for the altered toxicity of some anticancer agents. These interactions have been primarily attributed to the hyperforin-mediated induction of some pharmacokinetic mechanisms. However, as previously demonstrated by our group, hypericin induces the expression of two ABC transporters: multidrug resistance-associated protein 1 (MRP1) and breast cancer resistance protein (BCRP). Because cisplatin (CDDP) and mitoxantrone (MTX) are potential substrates of ABC transporters, we investigated the effect of 24 h hypericin pre-treatment on the cytotoxicity of CDDP and MTX in human cancer cell lines. CDDP-sensitive and -resistant ovarian adenocarcinoma cell lines A2780/A2780cis, together with HL-60 promyelocytic leukemia cells and ABCG2-over-expressing cBCRP subclone, were used in our experiments. We present CDDP cytotoxicity attenuated by hypericin pre-treatment in both A2780 and A2780cis cells and MTX cytotoxicity in HL-60 cells. In contrast, hypericin potentiated MTX-induced death in cBCRP cells. Interestingly, hypericin did not restore cell proliferation in rescued cells. Nevertheless, hypericin did increase the expression of MRP1 transporter in A2780 and A2780cis cells indicating the impact of hypericin on certain resistance mechanisms. Additionally, our results indicate that hypericin may be the potential substrate of BCRP transporter. In conclusion, for the first time, we report the ability of hypericin to affect the onset and/or progress of CDDP- and MTX-induced cell death, despite strong cell cycle arrest. Thus, hypericin represents another SJW metabolite that might be able to affect the effectiveness of anti-cancer drugs and that could interact with ABC transporters, particularly with BCRP.     

Radiopharmaceutical evaluation of 131I-protohypericin as a necrosis avid compound

Hypericin is a necrosis avid agent useful for nuclear imaging and tumor therapy. Protohypericin, with a similar structure to hypericin except poorer planarity, is the precursor of hypericin. In this study, we aimed to investigate the impact of this structural difference on self-assembly, and evaluate the necrosis affinity and metabolism in the rat model of reperfused hepatic infarction. Protohypericin appeared less aggregative in solution compared with hypericin by fluorescence analysis. Biodistribution data of ¹³¹I-protohypericin showed the percentage of injected dose per gram of tissues (%ID/g) increased with time and reached to the maximum of 7.03 at 24?h in necrotic liver by gamma counting. The maximum ratio of target/non-target tissues was 11.7-fold in necrotic liver at 72?h. Pharmacokinetic parameters revealed that the half-life of ¹³¹I-protohypericin was 14.9?h, enabling a long blood circulation and constant retention in necrotic regions. SPECT-CT, autoradiography, and histological staining showed high uptake of ¹³¹I-protohypericin in necrotic tissues. These results suggest that ¹³¹I-protohypericin is a promising necrosis avid compound with a weaker aggregation tendency compared with hypericin and it may have a broad application in imaging and oncotherapy.     

Synthesis and preliminary biological evaluation of a 99mTc‐labeled hypericin derivative as a necrosis avid imaging agent

Mono‐[¹²³I]iodohypericin and mono‐[¹²³I]iodohypericin monocarboxylic acid are iodine‐123‐labeled hypericin derivatives which have shown great promise in preclinical studies as necrosis avid imaging agents in animal models of infarction. In view of the more attractive properties of a ^99mTc‐labeled hypericin derivative, we have synthesized a conjugate of protohypericin monocarboxylic acid with S‐benzoylmercaptoacetyldiglycyl‐diaminopentane in an overall yield of 15%. The conjugate was labeled with technetium‐99m by exchange labeling at pH 10 in a labeling yield of 95% followed by photocyclization to yield ^99mTc‐mercaptoacetyldiglycyl‐1,5‐diaminopentylene hypericincarboxamide (^99mTc‐13). The negatively charged ^99mTc‐13 complex was purified by reversed phase high‐pressure liquid chromatography and the log P_7.4 was determined to be 2.36. In normal NMRI mice, the complex showed slow hepatobiliary clearance while plasma clearance was rapid. The tracer was evaluated in rats with reperfused hepatic infarction by ex vivo autoradiography, gamma counting and histochemical techniques. Unlike the radioiodinated hypericin derivatives, the new tracer agent did not show preferential uptake in necrotic tissue on autoradiography and gamma counting techniques. Conjugation of hypericin with a ^99mTc‐chelate, resulting in a change in size, charge and lipophilicity, had a profound effect on the necrosis avidity of the tracer agent. The results show that ^99mTc‐13 is not suitable for imaging necrosis. Copyright © 2008 John Wiley & Sons, Ltd.     

How to make hypericin water-soluble

Hypericin, isolated from Hypericum perforatum, is an effective photodynamic substance as demonstrated by various studies. Practical forms of applications of hypericin solutions for systemic use and introduction into body cavities are, however, lacking. We developed an aqueous solution of hypericin non-covalently bound to polyvinylpyrrolidone (PVP). PVP is a poly-N-vinylamide of various degrees of polymerization and forms of intermolecular crosslinks suitable for diagnostic and therapeutic applications. We used PVP (molecular weights of PVP between 10 kD and 40 kD) as a complex forming agent to prepare hypericin for photodynamic therapy and diagnostics. In pure water, hypericin forms aggregates which are non-soluble and non-fluorescent. The hypericin-PVP complex binds more than 1000 mg of hypericin in presence of 100 g PVP or less and is soluble in 1 liter of pure water. Aqueous complex solutions of hypericin-PVP display a characteristic absorption spectrum and fluorescence emission band around 600 nm wavelength. Varying concentrations of hypericin do not cause a blue- or red-shift in the absorption maximum at 595 nm. Excitation at 200 nm to 500 nm leads to emission at 590 nm; a property conducive to diagnostic investigations both in vitro and in vivo. Furthermore, hypericin-PVP exhibits high photostability in the presence of oxygen and broad band light which ensures reproducible photodynamic therapy and diagnosis. CONCLUSION: Hypericin forms liquid molecular chromophore complexes in water when bound to PVP thus allowing investigations in biological media.     

Hypericin in the dark inhibits key steps of angiogenesis in vitro

Photoactivated hypericin has a potent cytotoxic effect over a wide range of cells. However, very recently hypericin has been shown to have antitumoral and antimetastatic effects in the dark. The aim of this study was to test whether hypericin in the dark affects angiogenesis. Different in vitro assays were used to study the potential effects of this compound on key steps of angiogenesis, namely, a colorimetric assay of cell proliferation/viability, a tubular formation on Matrigel assay, zymographic assays for gelatinases and urokinase, a wound assay for migration and a fluorometric assay for invasion through Matrigel. In this report, we show for the first time that hypericin kept in the dark inhibits several key steps of the angiogenic process, namely, bovine endothelial cell proliferation, formation of tubular-like structures on Matrigel, migration and invasion, as well as extracellular matrix degrading urokinase.