Endotoxin test by recombinant Factor C for 0.9% sodium chloride injection

Endotoxin contamination is a threat to the safety of pharmaceutical products, especially parenteral drugs. Any sterile and/or pyrogen-free pharmaceutical product requires regulatory specifications to ensure safe patient use. This study covers the performance evaluation study of an endotoxin quantitation commercial kit by recombinant Factor C (rFC), Endozyme II® Go, for 0.9% sodium chloride injection. The samples were spiked with endotoxin solutions between 0.0005 and 10 EU/mL and tested by the rFC kit to evaluate precision, accuracy, detection and quantification limits, linearity, and robustness. Each of the six points was assayed at least five times.The relative standard deviation for precision testing ranged from 1.9 to 8.3%. The recovery accuracy values of endotoxin were between 61% and 125% for the range from 0.005 to 10 EU/mL. The results demonstrated that the rFC method allows endotoxin quantification with accuracy, precision, specificity, and linearity for the range of 0.005 and 10 EU/mL for 0.9% sodium chloride injection. (AU)

A contaminação por endotoxinas é uma ameaça à segurança dos produtos farmacêuticos, especialmente dos medicamentos parenterais. Qualquer produto farmacêutico estéril e/ou livre de pirogênios requer especificações regulatórias para garantir a segurança de uso para o paciente. Este estudo abrange o estudo de avaliação de desempenho empregando o kit comercial Endozyme II® Go para quantificação de endotoxina, por Fator C recombinante (FCr), em amostras de cloreto de sódio 0,9% para uso parenteral. As amostras foram fortificadas com cinco concentrações distintas de soluções de endotoxina na faixa entre 0,0005 e 10 UE/mL. Cada um dos cinco níveis foi testado pelo menos cinco vezes para avaliação dos critérios de precisão, exatidão, limites de detecção e quantificação, linearidade e robustez. O desvio padrão relativo para os testes de precisão variou de 1,9 a 8,3%. Os valores de recuperação de endotoxina para o parâmetro exatidão estiveram compreendidos entre 61% e 125%. Os resultados demonstraram que o método por FCr permite a quantificação de endotoxinas com exatidão, precisão, especificidade e linearidade para a faixa de 0,005 e 10 UE/mL em amostras de cloreto de sódio 0,9% para uso parenteral. (AU)

In vitro study of histocompatibility and clearance of hemoperfusion adsorbents / 中华劳动卫生职业病杂志

Objective@#To evaluate the histocompatibility and clearance of chlorpyrifos and its metabolite of activated charcoal and adsorption resin by in vitro study.@*Methods@#Venous blood from volunteers were incubation with activated charcoal or adsorbent resins, cytometry parameters and plasma components were detected for evaluation the histocompatibility of adsorbents. Venous blood from volunteers mixed with chlorpyrifos and its metabolite were incubation with activated charcoal or adsorbent resins, plasma concentration of chlorpyrifos and its metabolite were detected for evaluation the efficacy of adsorbents.@*Results@#Incubation tests show that the absorbents reduce the blood platelet (F=3.671, P<0.05) , serum glucose (F=10.564, P<0.05) , albumin (F=5.239, P<0.05) , uric acid (F=7.175, P<0.05) , creatinine (F=23.673, P<0.05) , T3 (F=11.161, P<0.05) and free T3 (F=10.256, P<0.05) . However, other cytometry parameters and plasma components were not influenced. Both activated charcoal and adsorbent resins could reduce the plasma concentration of chlorpyrifos (F=798.110, P<0.01) and its metabolite (F=1495.212, P<0.05) .@*Conclusion@#In vitro test show that both activated charcoal and adsorbent resins could clear chlorpyrifos and its metabolite, however, could not influence main cytometry parameters and plasma components, the histocompatibility of adsorbents are satisfactory.

Study on the Viability Inhibitory Effects and Mechanism of Dictamnine on Mice Spleen Lymphocyte in vitro / 中国药房

OBJECTIVE： To study the inhibitory effect of dictamnine on the viability of mice spleen lymphocyte in vitro and explore its potential mechanism. METHODS： The primary spleen lymphocytes of mice were isolated and cultured. The cells were treated with 0 （blank control）， 50， 100， 150 μmol/L dictamnine for 24 h. MTT assay was used to determine the cell viability； Lactic dehydrogenase （LDH） assay was used to determine release rate of LDH. Early apoptosis rate was detected by flow cytometry. Necrosis rate was detected by Hoechst 33342 and PI double staining； Western blot assay was used to detect the protein expressions of Caspase 3 and Cleaved-Caspase 3 in cells. Comet assay was used to detect DNA damage in cells （reflected in the proportion of DNA tail area）. RESULTS： Compared with blank control， 100， 150 μmol/L dictamnine could significantly inhibit the viability of lymphocytes （P＜0.01）. 150 μmol/L dictamnine could significantly increase the release of LDH （P＜0.05）， and release rate reached 79.37％. 50， 100， 150 μmol/L dictamnine could improve the early apoptotic rate of lymphocyte， but there was no statistical significance （P＞0.05）. 150 μmol/L dictamnine could significantly increase the necrosis rate （P＜0.05）， and necrosis rate reached 78.64％. 50， 100， 150 μmol/L dictamnine could increase the protein expression of Caspase 3， but there was no statistical significance （P＞0.05）， while 50， 100 μmol/L dictamnine could improve the protein expression of Cleaved-Caspase 3 significantly （P＜0.05）. DNA damage was induced in a dose-dependent manner by dictamnine， in which 100 and 150 μmol/L dictamnine could significantly increase DNA tail area （P＜0.01）. CONCLUSIONS： Dictamnine can inhibit spleen lymphocyte viability， and the mechanism may be related to inducing spleen lymphocyte necrosis and DNA damage.

Obtaining, characterization and evaluation of two candidate preparations for probiotics developed with agroindustrial waste / Obtención, caracterización y evaluación de dos preparados candidatos a probióticos desarrollados con residuos agroindustriales

ABSTRACT Objective. Obtain, characterize and evaluate two bio-prepares developed from the sugar cane molasses - orange vinasse fermented with yeast and/or lactic acid bacteria. Materials and methods. A completely randomized design was used, with five repeats per treatment. The evaluated treatments were: T1, Lactobacillus acidophilus, Lactobacillus bulgaricus, Streptococcus thermophilus y T2, the previous bacteria plus Saccharomyces cerevisiae and Kluyveromyces fragilis (L-4 UCLV). The previous mentioned microorganisms were inoculated in a substratum compounded by molasses - vinasse and these were incubated at 37°C for 24 hours. To the bioprepares, physiochemical, microbiological and in vitro tests was made to evaluate the probiotic capacity. Results. Both bioprepares presented a dark brown color, sweet and a pH lesser than 4. The bromatological and microbiologic development were higher (p>0.05) in T2. Both bioprepares the viability was higher than 92%. in vitro tests two bioprepares were resistant to an acid pH, bile salts, broad spectrum of microbial activity and inhibitory effect to E. coli, Salmonella spp. and S. aureus. Conclusions. The bioprepares obtained from sugar cane molasses - orange vinasse fermented with yeast and lactic acid bacteria manifested physiochemical and microbiologic properties appropriated to probiotic products. In in vitro tests, their potential was demonstrated as a probiotic.

RESUMEN Objetivo. Obtener, caracterizar y evaluar dos biopreparados desarrollados a partir de melaza de caña de azúcar - vinaza de naranja fermentados con levaduras y/o bacterias ácido lácticas. Materiales y métodos. Se utilizó un diseño completamente aleatorizado con cinco repeticiones por tratamiento. Los tratamientos evaluados fueron: T1, Lactobacillus acidophilus, Lactobacillus bulgaricus, Streptococcus thermophilus y T2, las bacterias anteriores más Saccharomyces cerevisiae y Kluyveromyces fragilis (L-4 UCLV). En un sustrato compuesto por melaza- vinaza se inocularon los microorganismos anteriormente mencionados y estos fueron incubados a 37ºC por 24 h. Se les determinaron a los biopreparados los parámetros fisicoquímicos, microbiológico y se realizaron las pruebas in vitro para evaluar la capacidad probiótica. Resultados. Ambos biopreparados presentaron un color marrón oscuro, dulzón y con pH inferior a 4. El comportamiento bromatológicos y microbiológicos fueron mayores (p>0.05) en el T2. En ambos biopreparados la viabilidad fue superior a 92%. En pruebas in vitro, ambos biopreparados fueron resistentes a pH ácido, sales biliares, amplio espectro de actividad antimicrobiana y efecto inhibitorio a la E. coli, Salmonella spp. y S. aureus. Conclusiones. Los biopreparados obtenidos a partir de melaza de caña de azúcar-vinaza de naranja fermentados con levaduras y/o bacterias ácido lácticas demostraron propiedades físicoquímicas, microbiológicas apropiadas para productos probióticos. En las pruebas in vitro, se demostró su efecto potencial como probiótico.

In vitro evaluation of ivermectin, moxidectin, albendazole and pyrantel against cyathostomins of horses / Avaliação in vitro da ivermectina, moxidectina, albendazole e pirantel contra ciatostomíneos de equinos

Abstract Cyathostomins are the most prevalent nematodes of horses, and multidrug resistance has been reported worldwide. There is a need to implement alternative drug monitoring analytical tests. The objective of this study was to determine the consistency (5 repetitions) of the larval migration on agar test (LMAT) using ivermectin, moxidectin, pyrantel or albendazole against cyathostomin infective-stage larvae in eight different concentrations. LMAT showed a strong coefficient of determination (R2 > 0.91), between the test repetitions (n=5). The average 50% effective concentration (EC50) for ivermectin, moxidectin, pyrantel and albendazole were 0.0404, 0.0558, 0.0864 and 0.0988 nMol, respectively. The results of the EC50 for albendazole showed the greatest range of concentration. Ivermectin and moxidectin had the lowest in between-test variation. In the future, internationally certified susceptible isolates could be used for screening new drug candidates, or to follow up the pattern of drug efficacy from field populations.

Resumo Ciatostomíneos são os nematodas mais prevalentes em equinos e a resistência múltipla foi relatada em todo o mundo. Existe a necessidade de implementar o monitoramento dos produtos com testes analíticos alternativos. O objetivo deste estudo foi determinar a consistência (5 repetições) do teste de migração larval em ágar (TMLA) usando ivermectina, moxidectina, pirantel e albendazole contra larvas infectantes de ciatostomíneos em oito concentrações diferentes. O TMLA demonstrou um coeficiente de determinação (R2) acima de 0,91 entre as repetições do teste. A concentração efetiva para 50% (CE50) para ivermectina, moxidectina, pirantel e albendazole foi de 0,0404; 0,0558; 0,0864 e 0,0988 nMol, respectivamente. A CE50 do albendazole demonstrou a maior amplitude entre os testes. A ivermectina e a moxidectina tiveram as menores variações das doses entre as repetições. No futuro, isolados certificados susceptíveis poderão ser testados com o TMLA para indicação de novos produtos e mesmo para acompanhar o perfil de eficácia de populações do campo.

Predicting the sensitizing potency of trichloroethylene by in vitro test / 中国职业医学

OBJECTIVE: To predict the sensitizing potency and optimal sensitization dose of trichloroethylene( TCE) by an in vitro skin sensitization test on a human acute monocytosis cell line( THP-1).METHODS: THP-1 cells were cultured in vitro and exposed to 2,4-dinitrochlorobenzene( DNCB),sodium dodecyl sulfate( SDS),tert-butylhydroquinone( tBHQ)and TCE for 24 hours.Flow cytometry was used to detect the expression of cell surface marker such as cluster of differentiation( CD) 86 and CD54,and the optimal dose range for sensitization detection was determined.With the relative fluorescence intensity( RFI),CD86 ≥ 150 and CD54 ≥ 200 as the standard,the sensitizing potency and optimal sensitization dose of TCE were predicted.RESULTS: The concentration range of reagents for sensitization test on THP-1 cells was the dose range at which the relative cell survival rate reached 75.0%-100.0%.DNCB at the doses of 20.83,25.00 and 30.00 μmol/L,tBHQ at the dose of 5.80 μmol/L,TCE at the doses of 8.33,10.00 and 12.00 mmol/L,can cause sensitivity.SDS was recognized as a negative sensitizer.The expression of CD86 and CD54 was the highest when the concentration of TCE was 8.33 mmol/L,which was considered as the best sensitization dose.CONCLUSION: The optimum sensitization dose of TCE is 8.33 mmol/L,which can provide the basis for dose design in future study of TCE sensitization pathways.

An unusual dual hypersensitivity reaction to moxifloxacin in a patient

Both immediate and nonimmediate type hypersensitivity reactions (HRs) with a single dose of quinolone in the same patient have not been previously reported. A 47-year-old female patient referred to us because of the history of a nonimmediate type HR to radio contrast agent and immediate type HR to clarithromycin. She experienced anaphylaxis in minutes after the second dose of 50 mg when she was provocated with moxifloxacin. She was treated immediately with epinephrine, fluid replacement and methylprednisole and pheniramine. On the following day she came with macular eruptions, and she was treated with methylprednisolone. The positive patch test performed with moxifloxacin as well as the lymphocyte transformation test proved the T-cell mediated HR. In order to prove the immediate type HR, basophil activation test was performed but was found negative. This case report presents for the first time the 2 different types of HRs in a patient with a test dose of quinolone.

Verification of small intestinal submucosa graft stent in vitro / 国际生物医学工程杂志

Objective To verify the feasibility of using small intestine submucosa for graft stents.Methods A Z-type Nitinol wire stent was used as the metal stent material,and porcine small intestine submucosa was used as the biofilm material to prepare the bio-coated stent.In vitro implantation,extracorporeal pulsation and extracorporeal flexion of the stents were designed based on the requirements for the small intestine submucosa graft stent implantation during the endovascular aneurysm repair and of the mechanical properties of the graft stent during human body activity.The feasibility of using small intestine submucosa for the graft stents was evaluated according to the patency of the stent lumen after the experiment,the presence or absence of cracks in the membrane,and the presence or absence of fracture of the metal stent.Results In the in vitro experiments,all the samples were able to maintain complete tunica and luminal patency except the 8th sample that showed suture rupture.Besides,all the samples showed good rebounding and adhering properties in both in vitro implantations and extracorporeal pulsation pressure experiments,and no dangerous problems were found such as film rupture,suture loss and stent fracture.Conclusion Small intestine submucosa can meet the durability requirements of graft stents,adapt to the conditions of graft stents such as implantation,bending and blood pulsation,and can be used as the covering material of graft stents.

In vitro evaluation of ivermectin, moxidectin, albendazole and pyrantel against cyathostomins of horses

Abstract Cyathostomins are the most prevalent nematodes of horses, and multidrug resistance has been reported worldwide. There is a need to implement alternative drug monitoring analytical tests. The objective of this study was to determine the consistency (5 repetitions) of the larval migration on agar test (LMAT) using ivermectin, moxidectin, pyrantel or albendazole against cyathostomin infective-stage larvae in eight different concentrations. LMAT showed a strong coefficient of determination (R2 > 0.91), between the test repetitions (n=5). The average 50% effective concentration (EC50) for ivermectin, moxidectin, pyrantel and albendazole were 0.0404, 0.0558, 0.0864 and 0.0988 nMol, respectively. The results of the EC50 for albendazole showed the greatest range of concentration. Ivermectin and moxidectin had the lowest in between-test variation. In the future, internationally certified susceptible isolates could be used for screening new drug candidates, or to follow up the pattern of drug efficacy from field populations.

Resumo Ciatostomíneos são os nematodas mais prevalentes em equinos e a resistência múltipla foi relatada em todo o mundo. Existe a necessidade de implementar o monitoramento dos produtos com testes analíticos alternativos. O objetivo deste estudo foi determinar a consistência (5 repetições) do teste de migração larval em ágar (TMLA) usando ivermectina, moxidectina, pirantel e albendazole contra larvas infectantes de ciatostomíneos em oito concentrações diferentes. O TMLA demonstrou um coeficiente de determinação (R2) acima de 0,91 entre as repetições do teste. A concentração efetiva para 50% (CE50) para ivermectina, moxidectina, pirantel e albendazole foi de 0,0404; 0,0558; 0,0864 e 0,0988 nMol, respectivamente. A CE50 do albendazole demonstrou a maior amplitude entre os testes. A ivermectina e a moxidectina tiveram as menores variações das doses entre as repetições. No futuro, isolados certificados susceptíveis poderão ser testados com o TMLA para indicação de novos produtos e mesmo para acompanhar o perfil de eficácia de populações do campo.

In vitro simulation experimental study of a fully magnetically levitated ventricular assist device based on mock circulatory system / 医用生物力学

Objective To investigate the circulatory supporting effect of the third generation fully magnetically levitated China Heart ventricular assist device (CH-VAD) under heart failure (HF) condition.Methods An in vitro mock circulatory system (MCS) was developed.This system could simulate a healthy adult under resting state and a patient with heart failure,and incorporate the CH-VAD to evaluate the assisting performance under continuous flow mode.Furthermore,CH-VAD was equipped with a pulsatile flow controller and its initial performance was accessed.The pulsatile mode was obtained by using sinusoidal velocity waveform of the pump which synchronized the CH-VAD with the ventricle simulator of the MCS.Results CH-VAD under continuous flow mode could recover the hemodynamic parameters (arterial pressure and cardiac output) under HF condition to normal range.Preliminary pulsatile test results showed that amplitude of current pulse speed had a minor influence on the hemodynamic performance.CH-VAD under continuous flow and pulsatile flow mode could obtain comparable mean arterial pressure,systolic arterial pressure,diastolic arterial pressure and mean flow.Conclusions CH-VAD can generate a certain degree of speed pulse via appropriate pulsatility control,so as to provide sufficient support on ventricular function.Further optimization on pulsatile controller of CH-VAD is required to conform to natural physiology.The developed MCS can be utilized as an effective and controllable in vitro platform for design,optimization and verification of VADs or other mechanical circulatory support devices.

In vitro simulation experimental study of a fully magnetically levitated ventricular assist device based on mock circulatory system / 医用生物力学

Objective To investigate the circulatory supporting effect of the third generation fully magnetically levitated China Heart ventricular assist device (CH-VAD) under heart failure (HF) condition.Methods An in vitro mock circulatory system (MCS) was developed.This system could simulate a healthy adult under resting state and a patient with heart failure,and incorporate the CH-VAD to evaluate the assisting performance under continuous flow mode.Furthermore,CH-VAD was equipped with a pulsatile flow controller and its initial performance was accessed.The pulsatile mode was obtained by using sinusoidal velocity waveform of the pump which synchronized the CH-VAD with the ventricle simulator of the MCS.Results CH-VAD under continuous flow mode could recover the hemodynamic parameters (arterial pressure and cardiac output) under HF condition to normal range.Preliminary pulsatile test results showed that amplitude of current pulse speed had a minor influence on the hemodynamic performance.CH-VAD under continuous flow and pulsatile flow mode could obtain comparable mean arterial pressure,systolic arterial pressure,diastolic arterial pressure and mean flow.Conclusions CH-VAD can generate a certain degree of speed pulse via appropriate pulsatility control,so as to provide sufficient support on ventricular function.Further optimization on pulsatile controller of CH-VAD is required to conform to natural physiology.The developed MCS can be utilized as an effective and controllable in vitro platform for design,optimization and verification of VADs or other mechanical circulatory support devices.

In vitro simulation experimental study of a fully magnetically levitated ventricular assist device based on mock circulatory system / 医用生物力学

Objective To investigate the circulatory supporting effect of the third generation fully magnetically levitated China Heart ventricular assist device (CH-VAD) under heart failure (HF) condition. Methods An in vitro mock circulatory system (MCS) was developed. This system could simulate a healthy adult under resting state and a patient with heart failure, and incorporate the CH-VAD to evaluate the assisting performance under continuous flow mode. Furthermore, CH-VAD was equipped with a pulsatile flow controller and its initial performance was accessed. The pulsatile mode was obtained by using sinusoidal velocity waveform of the pump which synchronized the CH-VAD with the ventricle simulator of the MCS. Results CH-VAD under continuous flow mode could recover the hemodynamic parameters (arterial pressure and cardiac output) under HF condition to normal range. Preliminary pulsatile test results showed that amplitude of current pulse speed had a minor influence on the hemodynamic performance. CH-VAD under continuous flow and pulsatile flow mode could obtain comparable mean arterial pressure, systolic arterial pressure, diastolic arterial pressure and mean flow. Conclusions CH-VAD can generate a certain degree of speed pulse via appropriate pulsatility control, so as to provide sufficient support on ventricular function. Further optimization on pulsatile controller of CH-VAD is required to conform to natural physiology. The developed MCS can be utilized as an effective and controllable in vitro platform for design, optimization and verification of VADs or other mechanical circulatory support devices.

In vitro simulation experimental study of a fully magnetically levitated ventricular assist device based on mock circulatory system / 医用生物力学

Objective To investigate the circulatory supporting effect of the third generation fully magnetically levitated China Heart ventricular assist device (CH-VAD) under heart failure (HF) condition.Methods An in vitro mock circulatory system (MCS) was developed.This system could simulate a healthy adult under resting state and a patient with heart failure,and incorporate the CH-VAD to evaluate the assisting performance under continuous flow mode.Furthermore,CH-VAD was equipped with a pulsatile flow controller and its initial performance was accessed.The pulsatile mode was obtained by using sinusoidal velocity waveform of the pump which synchronized the CH-VAD with the ventricle simulator of the MCS.Results CH-VAD under continuous flow mode could recover the hemodynamic parameters (arterial pressure and cardiac output) under HF condition to normal range.Preliminary pulsatile test results showed that amplitude of current pulse speed had a minor influence on the hemodynamic performance.CH-VAD under continuous flow and pulsatile flow mode could obtain comparable mean arterial pressure,systolic arterial pressure,diastolic arterial pressure and mean flow.Conclusions CH-VAD can generate a certain degree of speed pulse via appropriate pulsatility control,so as to provide sufficient support on ventricular function.Further optimization on pulsatile controller of CH-VAD is required to conform to natural physiology.The developed MCS can be utilized as an effective and controllable in vitro platform for design,optimization and verification of VADs or other mechanical circulatory support devices.

In Vitro Diagnostic Testing for Antibiotic Allergy

Allergy to antibiotics is an important worldwide problem, with an estimated prevalence of up to 10% of the population. Reaction patterns for different antibiotics have changed in accordance with consumption trends. Most of the allergic reactions to antibiotics have been reported for betalactams, followed by quinolones and macrolides and, to a lesser extent, to others, such as metronidazole clindamycin and sulfonamides. The diagnostic procedure includes a detailed clinical history, which is not always possible and can be unreliable. This is usually followed by in vivo, skin, and drug provocation tests. These are not recommended for severe, potentially lifethreaten reactions or for drugs that are known to produce a high rate of false positive results. Given the limitations of in vivo tests, in vitro test can be helpful for diagnosis, despite having suboptimal sensitivity. The most highly employed techniques for diagnosing immediate reactions to antibiotics are immunoassays and basophil activation tests, while lymphocyte transformation tests are more commonly used to diagnose non-immediate reactions. In this review, we describe different in vitro techniques employed to diagnose antibiotic allergy.

Study on the Function Change of Caco-2 Cell CYP3A4 and P-gp under Diabetic Conditions and Its Mecha-nism / 中国药房

OBJECTIVE:To investigate the function change of intestinal CYP3A4 and P-glycoprotein (P-gp) under diabetic conditions and its mechanism. METHODS:Caco-2 cells were respectively added with 25, 50 and 100 μmol/L midazolam (CYP3A4 probe substrate) for incubation for 15,30,60,120,180 min;with 0.1,0.2,0.4 μg/ml rhodamine 123 (P-gp sub-strate)for incubation for 15,30,60,90,120 min to determine the substrate concentrations and incubation time. Caco-2 cells were added with insulin,glucose and fatty acid (palmitic acid and oleic acid) at different concentrations,and then the production of 1′-OH-midazolam,the metabolite of midazolam,and the intake of rhodamine 123 were determined,in order to investigated the ef-fect on the function of CYP3A4 and P-gp. RESULTS:The optimal substrate concentrations and incubation time were as follows as 50 μmol/L midazolam,0.1 μg/ml rhodamine 123 and incubation for 2 h. With the increase in the concentration of insulin,glucose and palmitic acid,the production of 1′-OH-midazolam and the activity of CYP3A4 reduced;the intake of rhodamine 123 in-creased,and the efflux transport function of P-gp decreased. Oleic acid had no significant effect on the production of 1′-OH-mid-azolam or the intake of rhodamine 123. CONCLUSIONS:Under diabetes condition,the increase of insulin,glucose and palmitic acid may reduce the function of CYP3A4 and P-gp,while oleic acid has no effect on the function.

Optimization and Preliminary Methodology Study of in vitro Fresh Human Whole Blood Detection Method for Pyrogen-related Factor IL-1β / 中国药房

OBJECTIVE:To establish the detection method for pyrogen-related factor interleukin 1β(IL-1β)through optimiz-ing detection condition,and to conduct preliminary methodology study. METHODS:The in vitro fresh human whole blood detec-tion method was used. The bacterial endotoxin standard solution(5,2,0.8,0.32 EU/ml)were added into diluted blood;using di-luted RPMI 1640 as blank control,the content of IL-1β in blood sample was determined by ELISA after incubation. The relation-ship of the addition of different attenuants(RPMI 1640 culture,sterilized normal saline)and fetal bovine serum,final dilution vol-ume fraction(40%,20%,10%and 8.3%)and storage duration(2,5,6,8,26 h)with the contents of endotoxin IL-1βwere in-vestigated,and related coefficient and detection limits were calculated. Different dilution times of Qingkailing injection and Ginaton injection samples and interference solutions were added into diluted blood to detect their recovery. RESULTS:The results indicated that RPMI 1640 media and 40% diluted blood was more sensitive(detection limit was 0.128 EU/ml,r=0.993);while the addition of fetal bovine serum didn’t influence the results. The detection limits of blood sample storied at 4 ℃ for 26 h were 0.128 EU/ml (r>0.990). When Qingkailing injection and Ginaton injection were diluted 10,32 and more times,the detection method was not interfered and the recovery ranged 68%-118%. CONCLUSIONS:Established in vitro fresh human whole blood detection method can be used for the detection of pyrogen,and provides trial evidence for the pyrogen detection of TCM injection.

Homeopathic drug standardization through biological evaluations: An untrodden avenue

There is a dearth of chemico-analytical or instrumental methods for standardization and quality control of higher dilutions of homeopathic drugs. Aim: This review highlights the challenges in standardization of anti-inflammatory homeopathic drugs and suggests a battery of biological assays for their standardization. Methods: We retrieved a total 57 scientific reports from the experimental studies and scientific reviews published between January 1999 and June 2014 related to anti-inflammatory homeopathic drugs and their high dilutions. These comprised of 18 reports on preclinical evaluation, 15 on source materials, 9 on isolated constituents and 15 studies on in-vitro experiments. Few recent citations which supported the initial studies were added later during the compilation of the manuscript. Conclusion: Standardization and quality control of homeopathic mother tinctures and high dilutions warrants an urgent attention. As biological activities are observed to be attributed to the high dilutions which are practically devoid of active ingredients, their standardization may be done through the suggested battery of biological investigations. It is suggested that the current methods of standardization of homeopathic drugs need to be upgraded to include sensitive, reproducible and relevant biological assays so that the end users are assured of the quality, efficacy, and safety of homeopathic dilutions...

Study of Mechanism on in vitro Permeation of Phillyrin through Blood-brain Barrier / 中国药房

OBJECTIVE:To research the mechanism of in vitro permeation of phillyrin through blood-brain barrier. METH-ODS:After Madin-Darby canine kidney epithelial cells transfected with colorectal cancer MDR1 gene (MDCK-MDR1) were cul-tured with phillyrin solution of 0(negative control),10,25,50,75 and 100μg/ml for 24 h,cell viability was determined by resa-zurin method and cell survival rate was calculated. After MDCK-MDR1 cells were cultured with phillyrin solution of 10,25,50, 75 and 100 μg/ml for 10 min,the content of phillyrin in the cells was determined,and concentration-uptake rate curve was drawn. Following 3 h culture of MDCK-MDR1 cells with phillyrin solution of 0 (negative control),50 and 100 μg/ml,the structure of cell tight junction protein was observed under the inverted microscope. RESULTS:Compared to the negative control group,after 24 h cell culture with phillyrin solution of 10-100 μg/ml,no obvious change in cell survival rate occurred. MDCK-MDR1 cells cultured with the phillyrin at a mass concentration of 10-100 μg/ml demonstrated a nonlinear relationship with concentration of phillyrin and a gradual saturation trend. After the cells were cultured with phillyrin of 50 and 100 μg/ml for 3 h,cell tight junction protein was intact. CONCLUSIONS:The absorption of phillyrin through the simulated blood-brain barrier may be in the form of passive transportation combined with active transportation,the concertration has effect on cell tight junction protein.

Sensitivity of Plasmodium falciparum to Antimalarial Drugs in Hainan Island, China

Pyronaridine and artesunate have been shown to be effective in falciparum malaria treatment. However, pyronaridine is rarely used in Hainan Island clinically, and artesunate is not widely used as a therapeutic agent. Instead, conventional antimalarial drugs, chloroquine and piperaquine, are used, explaining the emergence of chloroquine-resistant Plasmodium falciparum. In this article, we investigated the sensitivity of P. falciparum to antimalarial drugs used in Hainan Island for rational drug therapy. We performed in vivo (28 days) and in vitro tests to determine the sensitivity of P. falciparum to antimalarial drugs. Total 46 patients with falciparum malaria were treated with dihydroartemisinin/piperaquine phosphate (DUO-COTECXIN) and followed up for 28 day. The cure rate was 97.8%. The mean fever clearance time (22.5+/-10.6 hr) and the mean parasite clearance time (27.3+/-12.2 hr) showed no statistical significance with different genders, ages, temperatures, or parasite density (P>0.05). The resistance rates of chloroquine, piperaquine, pyronarididine, and artesunate detected in vitro were 71.9%, 40.6%, 12.5%, and 0%, respectively (Ppiperaquine>pyronarididine>artesunate. The inhibitory dose 50 (IC50) was 3.77x10(-6) mol/L, 2.09x10(-6) mol/L, 0.09x10(-6) mol/L, and 0.05x10(-6) mol/L, and the mean concentrations for complete inhibition (CIMC) of schizont formation were 5.60x10(-6) mol/L, 9.26x10(-6) mol/L, 0.55x10(-6) mol/L, and 0.07x10(-6) mol/L, respectively. Dihydroartemisinin showed a strong therapeutic effect against falciparum malaria with a low toxicity.

Sensitivity of Plasmodium falciparum to Antimalarial Drugs in Hainan Island, China

Pyronaridine and artesunate have been shown to be effective in falciparum malaria treatment. However, pyronaridine is rarely used in Hainan Island clinically, and artesunate is not widely used as a therapeutic agent. Instead, conventional antimalarial drugs, chloroquine and piperaquine, are used, explaining the emergence of chloroquine-resistant Plasmodium falciparum. In this article, we investigated the sensitivity of P. falciparum to antimalarial drugs used in Hainan Island for rational drug therapy. We performed in vivo (28 days) and in vitro tests to determine the sensitivity of P. falciparum to antimalarial drugs. Total 46 patients with falciparum malaria were treated with dihydroartemisinin/piperaquine phosphate (DUO-COTECXIN) and followed up for 28 day. The cure rate was 97.8%. The mean fever clearance time (22.5+/-10.6 hr) and the mean parasite clearance time (27.3+/-12.2 hr) showed no statistical significance with different genders, ages, temperatures, or parasite density (P>0.05). The resistance rates of chloroquine, piperaquine, pyronarididine, and artesunate detected in vitro were 71.9%, 40.6%, 12.5%, and 0%, respectively (Ppiperaquine>pyronarididine>artesunate. The inhibitory dose 50 (IC50) was 3.77x10(-6) mol/L, 2.09x10(-6) mol/L, 0.09x10(-6) mol/L, and 0.05x10(-6) mol/L, and the mean concentrations for complete inhibition (CIMC) of schizont formation were 5.60x10(-6) mol/L, 9.26x10(-6) mol/L, 0.55x10(-6) mol/L, and 0.07x10(-6) mol/L, respectively. Dihydroartemisinin showed a strong therapeutic effect against falciparum malaria with a low toxicity.