Comparison in vitro permeation of curcumin from topical gels by liquid chromatography tandem mass spectrometry

Objective： To fred a more effective method of topical transdermal delivery of curcumin. Methods： We prepared curcumin carbopol （CRB） 974P and hydroxypropylcellulose （HPC） gel formulations containing menthol or Azone as permeation enhancers In this study, negative mode electrospray ionization and a triple quadruple LC/MS/MS instrument operated in multiple reaction mode was used for curcumin detection. The assay was linear over a concentration range of 10 ng/mL to 400 ng/mL for curcumin （average R2 = 0.997 2）. Excised nude mouse dorsal side skin was used in an in vitro skirt permeation study performed using the method of Franz. Results： Our results showed that all of the topical gel formulations we developed were free from skin irritation. The percutaneous flux and enhancement ratio of curcumin across nude mouse epidermis were enhanced markedly by the addition of menthol or Azone to both types of gel formulations. We found that the HPC gels containing quantities of Azone showed an enhanced permeation effect as compared to gels containing menthol. In the case of HPC gels containing Azone, the increase in permeability was significant （P〈0.05） as compared to the gels containing menthol. Conclusion： Azone shows a significantly more remarkable permeation effect than menthol. As such, this novel delivery strategy offers significant promise and is worthy of further exploration in attempts to enhance the medicinal application of curcumin     

PATHOGENESIS OF EXTRINSIC ALLERGIC ALVEOLITIS AND PULMONARY FIBTOSIS INDUCED BY STREPTOMYCES THERMOHYGROSCOPICUS

There were 48 strains of thermophilic actinomyces isolated from the specimens of mouldy hay and sputum of the patients suffering from farmer’s lung (FL). Streptomyces thermohygroscopicus (STHs),one strain of them, was used for this investigation. The microorganisms were injected into the lungs of rabbits and rats by thyrocrico – or tracheocentesis. The result showed that the pathological changes in the lungs including macrophage alveolitis, granuloma formation and diffuse interstitial were similar to that induced by other thermophilic actinomyces. IgG and C3 deposition in the lesions were also observed by immunofluorescence examination. Specific immunocomplexes in the sera of some animals were detected by ELISA with the STHs was possibly one of the pathogens responsible for FL in China’s countryside.     

Effect of Steroidal Contraceptives on Pituitary LH Storage and Release in Proestrous Rats

The pituitary response to GnRH in proestrous rats treated with steroidal contraceptives (Anordrin or Levonorgestrel) was studied.The dose of Anordrin for treated group was 3mg or 0.75mg in 0.5ml tea oil,while that of Levonorgestrel for treated group was 4mg or 1mg in 0.5ml tea oil,and the control group was treated with 0.5ml in tea oil. The rats were administered with two injections of GnRH through external jugular vein with an interval of 60 minutes.LH concentration in the plasma and its content in pituitary homogenates at 150 minutes after administration were measured by radioimmunoassay.The results indicate that LH concentration in the plasma increased remarkably while its content in the pituitary showed a slight decrease without statistical significance in Anordrin treated groups.However,in Levonorgestrel treated groups,plasma LH concentration decreased with a concurrent significant increase of LH content in the pituitary.It is suggested that different type of steroidal contraceptives may influence anterior pituitary LH release and storeage.     

Protection of salvianolate against atherosclerosis via regulating the inflammation in rats

Inflammation plays an essential role in the pathophysiology of atherosclerosis. Our study was aimed to investigate whether salvianolate, a novel water-soluble phenolic compound of Danshen, alleviates atherosclerosis via regulating the inflammation in rats. High fat diet feeding plus vitamin D3 injection was used to induce atherosclerosis in rats. Salvianolate （60, 120 or 240 mg/kg） or placebo was given to atherosclerotic rats. The plasma lipids, interleukin 6 （IL-6） and C reactive protein （CRP） were measured by ELISA. CD4＋CD25＋Foxp3＋ cells were determined by flow cytometry. Histological changes were examined by hematoxylin and eosin staining. The results showed that the levels of plasma IL-6 and CRP were elevated in the rats fed on high fat diet, and the histological analysis demonstrated the successful establishment of atherosclerosis models. Treatment with salvianolate alleviated the atherosclerotic process and decreased the levels of plasma IL-6 and CRP. Also the number of CD4＋CD25＋Foxp3＋ cells was increased in salvianolate-treated rats. It was concluded that salvianolate could treat atherosclerosis via modulating the inflammation at cytokine and cell levels.     

Eiectroacupuncture Prevents Cognitive Deficits Caused by Topiramate （TPM） in Rats

Objective Recent studies have demonstrated that acupuncture is feasible to treat cognitive impairments. The objective of this study was to present behavioral evidence that electro-acupuncture （EA） could improve the learning and memory of TPM administrated rats. Methods The pattern of TPM-induced cognitive deficits in rats was made by administration of TPM intragastrically for 3 weeks. Of them the rats which showed damage in learning and memory （n=45） were randomly allocated to 3 groups： Impaired group （n=15）, EA group （n=15） and placebo-EA group （n=15）. Moreover, normal group （n=15） was set as control groups. EA stimulation was provided at acupoints located in either the midline of the back or of the head： Gv-20 （Baihui）, Gv-14 （Dazhui）. Morris water maze test was employed to assess spatial discriminative ability per group respectively and to analyze the curative effects of EA. Results Compared to the normal group, obvious cognitive deficits were found in the impaired and placebo-EA groups, and the statistic analysis showed that there were significant differences between normal and impaired groups in ANOVA. In the EA group, shortened mean escape latency was detected compared with the impaired effect on cognitive group during the same trial days; search strategy changed from random pattern adopted by impaired and placebo-EA rats to tendency or linear pattern popular in the normal group. Conclusion The present results suggested that EA exerted a protective impairment caused by TPM in rats, and EA has a specificity of cure. EA as a potential clinic method in treating TPM-induced cognitive impairment should be developed and investigated in the future.     

Biocompatibility of KLD-12 peptide hydrogel as a scaffold in tissue engineering of intervertebral discs in rabbits

KLD-12 peptide with a sequence of AcN-KLDLKLDLKLDL-CNH2 was synthesized and its biocompatibility was assessed in animals. Rabbit MSCs were cultured in the hydrogel for 2 weeks. Live cells were counted by using Calcein-AM/P1 fluorescence staining. MTT was employed to assess the viability of MSCs cultured in KLD-12 peptide solution of 0.01%, 0.03%, and 0.05%. Hemolysis test, skin irritation test and implantation test were conducted to evaluate its biocompatibility with host tissues. Our results demonstrated that the MSCs in hydrogel grew well and maintained round shape. Cell survival rate was 92.15% （mean： 92.15%±1.17%） at the 7th day and there was no difference in survival rate between day 7 and day 14. Cell proliferation test showed that the A value of the KLD-12 solutions was not significantly different from that of control groups （complete culture media） （P〉0.05） at the 24th and 48th h. The hemolysis rate of KLD-12 solution was 0.112%. Skin irritation test showed that the skin injected with KLD-12 solution remained normal and the score of skin irritation was 0. The histological examination with HE staining exhibited that the skin layers were clear and there was no infiltration with neutrophilic granulocytes and lymphocytes. It is concluded that KLD-12 peptide hydrogel bad a good biocompatibility with host rabbit and MSCs, and KLD-12 pep- tide hydrogel can provide an appropriate microenvironment for MSCs.     

A COMPARATIVE STUDY ON THE ANTIFERTILITY EFFECT OF PHENOL ALDEHYDES COMPOUNDS

The spermicidal activity of 5 phenol aldehydes compounds in vitro was compared in various species including rats, mice, rabbits, hamsters, dogs and human. In rats, mice, hamsters     

UniPron is A Fully Effective Non-hormonal Reversible Contraceptive in Baboon Model （Papio Anubis）

Objective To determine the safety and efficacy of UniPron as a reversible contraceptive.
Methods Vaginal swabs were obtained before and after UniPron administration, cultured onto appropriate culture media and bacteria identification was done based on type of media used, Gram stain reactions, colony morphology and biochemical tests. Vaginal biopsy tissues were processed using paraffin wax method, stained with hematoxylin and eosin and examined under light microscopy to determine the effect of the product on vaginal tissues. The effect of UniPron on sperm was examined by mixing the product with electroejaculated spermatozoa in vitro at different concentrations. For efficacy studies, male baboons of proven fertility were mated with UniPron treated or untreated females of proven fertility during the fertile stages.
Results All the five females （100%） that were treated with UniPron did not conceive and they regained total fertility when the treatment was stopped while all the controls conceived. At a concentration of 40%, UniPron completely immobilized spermatozoa in an in-vitro system. UniPron mechanism of action was by lowering the vaginal pH and on application in baboon, the pH was lowered for at least 3 h after which it went back to normal.
Conclusions As we plan for a study to test UniPron as a microbicide to prevent STIs including HIV, our current study has established that this novel product is effective in contraception and harmless to vaginal tissues and vaginal microbial flora in a baboon model （Papio anubis）.     

Estrogen Receptor α and β Expressions in Hypothalamus-pituitary-ovary Axis in Rats Exposed Lactationally to Soy Isoflavones and Bisphenol A

Objectives This paper aims to investigate the uterotrophic activities of lactational exposure to combination of soy isoflavones （SIF） and bisphenol A （BPA） and to examine estrogen receptor α （ERα） and estrogen receptor β （ERβ） expressions in hypothalamus-pituitary-ovary axis and uterus.Methods Maternal rats that were breeding about 8 litters were randomly divided into four groups with seven dams in each group.Dams in different treatment groups received corn oil （control）,150 mg/kg BW of SIF,150 mg/kg BW of BPA or combination of 150 mg/kg BW of SIF and 150 mg/kg BW of BPA,respectively,from postnatal day 5 to 11 （PND5-11） by gavage.On PND12 and PND70,10 female litters were killed and hypothalamus,pituitary,ovary and uterus were collected.ERα and ERβ expressions in these organs were detected with Western blotting assay.And vaginal opening time and estrus cycle were examined in animals fed for PND70.Results On PND12,the relative uterine weight of rats treated with ISF or BPA or their combination was significantly higher than that of untreated rats （P〈0.05）.But the relative uterine weight of rats in the co-exposure group was slightly lower than that in the group only exposed to SIF or BPA.On PND 70,however,the relative uterine weight in each treatment group was not statistically different from that in the control group （P〈0.05）.Vaginal opening time and estrus cycle in groups treated with SIF or BPA or their combination were similar to those in the control group （P〈0.05）.Exposure to SIF or BPA or their combination could up-regulate or down-regulate ERα and ERβ expressions in hypothalamus,pituitary,ovary and uterus on PND12 and PND70.These regulation patterns for ERα and ERβ were different in different organs at different time points.Conclusion Lactational exposure to ISF or BPA or their combination could induce uterotrophic responses in neonate rats,which disappeared in later life.But these data fail to suggest a possibility for synergic actions between SIF and BPA.It was also demonstrated that the uterotrophic effects of SIF and BPA exposure might,at least,involve modification of ERα or ERβ expressions in the hypothalamus-pituitary-ovary axis.     

Effects of WH-1 and WH-2 eye drops on experimental allergic uveitis in rabbits

Effects of WH-1 and WH-2 eye drops on experimental allergic uveitis(FAU)in rabbits induced by bovine serum albumin were clinically and pathologically observed in comparisonwith local drops of dexamethasone used as positive control.The results showed that WH-2 obvi-ously inhibited the inflammatory reactions while WH-1 did not.Protein concenwation in the aqueoushumor of WH-2-treated group of rabbits was 9.24±2.34mg/ml,which was notably lower(P<0.01)than both that of the untreated(14.33±3.14 mg/ml)and of the WH-1-treated(12.91±3.95mg/ml),but signifificantly higher than that of the dexame dexamethasone-treated goup(4.43±1.43mg/ml,P<0.01).The alleviation of the intraocular inflammatory reaction by therapy of WH-2eye drops was confimed pathologically.This study indicates that WH-2 has a positiveantiphlogistic effect on EAU in rabbits.     

Spermicidal activity of bivittoside D from Bohadschia vitiensis

BACKGROUND: A recent revelation about increased susceptibility to HIV by use of nonoxynol-9 (N-9) has called for identification of novel molecules with potent sperm-attenuating activity and lower side-effect profile, as suitable alternatives. The present study was designed to investigate spermicidal activity in Bohadschia vitiensis whole-body extracts followed by isolation and characterization of bioactive molecule. METHODS: Bohadschia vitiensis (Semper) was collected from the Southern Andaman coast of India. Freshly collected marine animals were extracted with methanol. A portion of the crude extract was fractionated into four fractions by macerating with hexane, chloroform, and n-butanol successively. All fractions were evaluated for spermicidal activity. Because maximum activity was localized in the n-butanol soluble fraction, it was chromatographed over a silica gel column, and elution with chloroform-methanol-water (35:10:2, v/v) yielded the major compound bivittoside D (400 mg). Bivittoside D [molecular weight (MW) 1426] is a lanostane triterpenoid with six monosaccharide units. The structure of the compound was established on the basis of physicochemical data, acid hydrolysis of saponin, identification of sugar units and aglycon, melting point, and by comparison with data reported in the literature. RESULTS: The aqueous methanol extract of the Bohadschia vitiensis caused 100% mortality of human sperm at 0.01% concentration in vitro, whereas N-9 (reference control) exhibited an equivalent activity at 0.05%. On further fractionation, activity was localized in n-butanol soluble fraction from which the major compound purified was a lanostane triterpenoid called bivittoside D. Bivittoside D was found to be a more potent spermicide ( approximately 2.3 times) than N-9 and killed 100% human sperm at the concentration of 350 muM in approximately 20 sec in vitro. Supravital staining and hypoosmotic swelling test revealed sperm membrane permeabilization by bivittoside D as the major mode of spermicidal action. However, bivittoside D was much safer than N-9 towards normal vaginal flora (Lactobacillus) in vitro, although it affected the viability of HeLa cells like other surfactants. CONCLUSION: Bivittoside D from B. vitiensis can adequately replace N-9 in vaginal contraceptives to make them more vaginally safe and ecofriendly.     

Spermicidal Effect of Jieze No.1 in Combination with Nonoxynol-9 In Vitro

Presently, topical spermicides are character ized by their convenience and negligible detrimentalinfluence on the human body compared with othercontraception methods. Nonoxynol 9 (N 9) is themost common spermicide contraception in theworld market for its ability to prevent transmissionof some STD pathogens[1, 2]. But the drawbacks ofN 9 are that it disrupts lipid membranes[3], causesvaginal irritation, and influences the normal bacte rial clot of vagina[4] in high doses. Th…     

0.5%鬼臼毒素纳米脂质载体在阴道黏膜的毒性试验

目的考察0.5%鬼臼毒素纳米脂质载体在阴道黏膜的安全性。方法（1）刺激性试验：将12只新西兰兔均分为3组,分别予 生理盐水（NS）、0.5%鬼臼毒素酊剂（POD-T）和0.5%鬼臼毒素纳米脂质载体（POD-NLC）,阴道给药10 d后取阴道组织行病理检 查,根据Eckstein评分标准进行评估。（2）急性毒性试验：将20只SD雌性大鼠均分2 组,分别予赋形剂和POD-NLC,以300倍临 床剂量在24 h内分3次阴道给药,连续观察14 d后解剖并计算脏器系数。（3）长期毒性试验：80只SD雌性大鼠均分4组,分别予 赋形剂、低剂量POD-NLC（15倍临床剂量）、中剂量POD-NLC（30倍临床剂量）和高剂量POD-NLC（60倍临床剂量）。连续13 周每日阴道给药1次,每周记录体质量,给药结束后对每组2/3的大鼠进行抽血化验、解剖并计算脏器系数和肝脏组织病理学检 查。为观察药物延迟毒性,每组剩余1/3的大鼠停药观察2周后做同前处理。结果（1）刺激性试验：0.5% POD-NLC对阴道黏膜 为极轻刺激性,0.5% POD-T为中度刺激性;（2）急性毒性试验：赋形剂组与POD-NLC组大鼠相比,各器官的脏器系数的差异无 统计学意义（P>0.05）;（3）长期毒性试验：赋形剂组与POD-NLC各剂量组大鼠的一般表现、行为活动、体质量变化、血常规、血生 化、重要器官脏器系数及肝脏病理未见明显异常,且组间差异无统计学意义（P>0.05）。结论POD-NLC在阴道黏膜用药刺激性 较小,对机体无明显的急性及长期毒性,有较高的安全性。     

Toxicological studies on safety of rare earths used in agriculture

Studies on the toxicity and safety of a mixture of rare earth metal nitrates (Ce, La, Nd, Pr, and Sm) used in agricultural operations are reported. In mice, rats, and guinea pigs, the oral LD50 ranged from 1397 to 1876 mg/kg; absorption in the gastrointestinal tract was low. The accumulation coefficient was greater than 5. In rabbits, a topical application of a suspension of 500 mg/ml produced mild irritation of the skin and eye mucosa. Subchronic and chronic toxicity studies were done at different dose levels in monkeys (100 mg/kg) and rats (200 and 1800 mg/kg); biochemical and histopathological examination of tissues showed no abnormal or specific pathological changes. In chronic feeding studies with rats, the incidence of tumors and malignant tumors in test groups was lower than that in the control. Rat fetuses did not show any teratogenicity when the dams were orally fed up to 330 mg/kg of this nitrate mixture. Ames mutagenicity tests were negative at a 50 mg/kg level. The results indicate that an oral dose of 60 mg/kg should be considered as a no-effect level with an ADI of 0.6 mg/kg and that the level of rare earth nitrates used in Chinese agriculture is within acceptable risk or safety limits.     

乙酰甲胺磷·三唑磷乳油的毒性研究

目的研究以乙酰甲胺磷和三唑磷为主要成分的乳油的急性毒性,为其毒理安全性评价提供依据。方法按国家标准GB15670—1995进行急性经口、经皮、吸入毒性试验,急性皮肤及眼刺激试验、皮肤致敏试验。结果20％乙酰甲胺磷·三唑磷乳油对雌雄性sD大鼠经口LD50均为369mg／kg．BW;急性经皮LD50〉2000mg／kg．BW;急性吸入LC50〉2000mg／m3;对家兔皮肤刺激反应积分均值为0;急性眼刺激积分指数（I．A．O．I．）=6．0,（M．I．0．I．）48h为0;对豚鼠皮肤致敏率为0。结论20％乙酰甲胺磷·三唑磷乳油经口毒性属于中等毒,经皮毒性属于低毒,吸入毒性属于低毒,时眼睛有轻度刺激性。对皮肤无刺激性,对豚鼠致敏强度分类为弱致敏。     

Spermicidal Effect of Jieze No. 1 in Combination with Nonoxynol-9 In Vitro

Summary Spermicidal effect of Jieze No. 1 (JZ1) in combination with nonoxynol-9 (N-9) was examinedin vitro. The minimum spermicidal concentration of JZ1 decoction, N-9 and their mixture solution in 20 s and 3 min were examined by improved spermicidal test of Sander-cramerin vitro. The percentages of progressively moving spermatozoa, moving spermatozoa and viable spermatozoa were also observed 20 s, 3 min and 30 min after the addition of the liquid medicine. Our results showed that sperms did not recover their activities in a revival test when the minimum spermicidal concentration of either JZ1 decoction, or N-9, or the mixed solution of the two agents, was used. N-9 (JZ1 in the mixed group) showed significant differences in the percentages of progressively moving spermatozoa, moving spermatozoa, and visible spermatozoa in 20 s, 3 min, and 30 min, when compared with N-9 alone (P<0.01). We are led to conclude that JZ1 decoction can improve N-9 spermicidal actionin vitro, and when used in combination with N-9, it has synergic effect. XU Ping, female, born in 1979, Graduate Student This project was supported by the National “10th Five-Year” Key Technologies R & D (No. 2004BA 709 B13-02).     

白速消消毒液刺激性和皮肤变态反应综合研究

目的检测白速消皮肤黏膜消毒液的皮肤刺激性、阴道粘膜刺激性和产生皮肤变态反应的可能性及其强度。方法依照《消毒技术规范》(2002年版)中的皮肤刺激试验、阴道粘膜刺激试验和皮肤变态反应试验方法进行。结果一次皮肤刺激试验受试皮肤的最高皮肤刺激指数为0.3;阴道粘膜刺激试验染毒组和对照组的阴道粘膜未见明显白细胞浸润、充血和水肿,阴道粘膜刺激指数为1.6;皮肤变态反应试验组动物皮肤未见红斑与水肿,各时间段的皮肤刺激反应积分为零。结论白速消皮肤黏膜消毒液对家兔一次皮肤刺激强度为无刺激性,对家兔一次阴道粘膜刺激强度为极轻刺激性,致敏强度为未见皮肤变态反应。     

96%禾草敌原药的急性毒性研究

目的：观察96%禾草敌原药的急性毒性,为其安全生产和使用提供科学依据。方法：通过急性经口毒性试验、急性经皮毒性试验、皮肤和眼睛刺激试验及皮肤致敏试验对96%禾草敌原药的急性毒性进行综合评价。结果：以雌雄性大鼠为试验动物,96%禾草敌原药急性经口、经皮LD50（95%可信限）分别为681（562～1 210）mg/kg和1 470（1 210～2 610）mg/kg;家兔皮肤刺激平均分值为0;眼刺激积分指数（I.A.O.I）最高为2,眼刺激的平均指数（M.I.O.I）48 h后为0;豚鼠皮肤致敏率为0%。结论：在本实验条件下,96%禾草敌原药急性经口、经皮毒性分别属低毒类和中等毒类,对皮肤和眼睛均为无刺激性,属弱致敏性农药。     

辣椒碱纳米乳注射剂的溶血性与血管刺激性试验

目的对辣椒碱纳米乳注射剂进行溶血性与血管刺激性评价。方法采用家兔体外溶血性试验与家兔体内血管刺激性试验。结果0.004%辣椒碱纳米乳注射剂3 h内无溶血以及无红细胞凝集反应,按20μg/kg家兔耳缘静脉注射无血管刺激性。结论在有效剂量下辣椒碱纳米乳注射剂无溶血及血管刺激性反应,为进一步研究提供参考。