The stability of the drug actarit was studied under different stress conditions like hydrolysis（acid,alkaline and neutral）,oxidation,photolysis and thermal degradation as recommended hy International Conference on H...The stability of the drug actarit was studied under different stress conditions like hydrolysis（acid,alkaline and neutral）,oxidation,photolysis and thermal degradation as recommended hy International Conference on Harmonization（ICH） guidelines.Drug was found to be unstable in acidic,basic and photolytic conditions and produced a common degradation product while oxidative stress condition produced three additional degradation products.Drug was impassive to neutral hydrolysis,dry thermal and accelerated stability conditions.Degradation products were identified,isolated and characterized by different spectroscopic analyses.Drug and the degradation products were synthesized by a new route using green chemistry.The chromatographic separation of the drug and its impurities was achieved in a phenomenex luna C18 column employing a step gradient elution by high performance liquid chromatography coupled to photodiode array and mass spectrometry detectors（HPLC-PDAMS）.A specilic and sensitive stability-indicating assay method for the simultaneous determination of the drug actarit.its process related impurities and degradation products was developed and validated.展开更多
A simple, rapid, precise, accurate, rugged and robust stability-indicating ultra-fast high performance liquid chromatographic (UHPLC) method has been developed for the estimation of related compounds (imp-A, imp-B, im...A simple, rapid, precise, accurate, rugged and robust stability-indicating ultra-fast high performance liquid chromatographic (UHPLC) method has been developed for the estimation of related compounds (imp-A, imp-B, imp-C, imp-D and imp-E) in Naproxen and also the assay of Naproxen from bulk drug samples. The stability indicating capability of the method was proven by subjecting the samples to stress conditions such as acid, base, oxidation, photolysis and thermal degradation. The efficient chromatographic separation was achieved using mobile phase solution A prepared as buffer solution 10 mM monobasic potassium phosphate pH 4.0 ± 0.05 adjusted with diluted ortho phosphoric acid solution and solution B acetonitrile with linear gradient elution on poroshell 120 EC-C18 shot column (50 mm × 4.6 mm, 2.7 μm) and UV detection at 235 nm at a flow rate 1.0 mL/min, column oven temperature was set to 25?C. The above are all known impurities and degradation impurities are well resolved with Naproxen peak and these are eluted within a 10 min runtime of HPLC. The photo diode array detector was used for peak homogeneity testing during stress study experiments and the overall mass balance was found to be 99.2% to 100.2% in all stress conditions. The linear calibration range was found to be 0.05 μg/mL to 0.75 μg/mL for related compounds and 50 μg/mL to 150 μg/mL for Naproxen and the accuracy of the method was found to be 91.5% to 98.5% recovery for the related substance method and 95.4% to 97.4% recovery for the assay method. The Naproxen and related compounds were found to be stable up to 48 hours and the method validation data show excellent results for precision, linearity, specificity, limit of detection, limit of quantitation and robustness. The present method can be successfully used for routine QC and stability studies and it will help to reduce the analysis cost, time and effluent load compared to conventional HPLC methods.展开更多
A patented kinetic uricase method was evaluated for serum uric acid assay. Initial absorbance of the reaction mixture before uricase action （A0） was obtained by correcting the absorbance at 293 nm measured before th...A patented kinetic uricase method was evaluated for serum uric acid assay. Initial absorbance of the reaction mixture before uricase action （A0） was obtained by correcting the absorbance at 293 nm measured before the addition of uricase solution, and background absorbance （Ab） was predicted by an integrated method. Uric acid concentration in reaction solution was calculated from AA, the difference between A0 and Ab, using the absorptivity preset for uric acid. This kinetic uricase method exhibited CV〈4.3% and recovery of 100%. Lipids, bilirubin, hemoglobin, ascorbic acid, reduced glutathione and xanthine 〈0.32 mmol/L in serum had no significant effects. △A linearly responded to 1.2 to 37.5 μmol/L uric acid in reaction solution containing 15 μl serum. The slope of linear response was consistent with the absorptivity preset for uric acid while the intercept was consistent with that for serum alone. Uric acid concentrations in clinic sera by different uricase methods positively correlated to each other. By Bland-Altman analysis, this kinetic uricase method accorded with that by quantifying the total change of UV absorbance on the completion of uricase reaction. These results demonstrated that this kinetic uricase method is reliable for serum uric acid assay with enhanced resistance to both xanthine and other common errors, wider range of linear response and much lower cost.展开更多
A new and senseitive Co (II)-biuret method for determination of protein has for the first time been established. In visible wavelength region, the Co (II) -protein complexes had anabsorption peak at 360 nm. -Under the...A new and senseitive Co (II)-biuret method for determination of protein has for the first time been established. In visible wavelength region, the Co (II) -protein complexes had anabsorption peak at 360 nm. -Under the present conditions. the assay had a linear range of 0-120 μg/ml of protein with a detection limit of about 2 μg/ml, and similar calibration curves were obtained for BSA. and BγG. Compared with Cu (II) -biuret method, the present Co (II) -biuret method was more simple, sensitive, and tolerant of interferences from non-protein molecules.展开更多
A simple, rapid and precise method has been developed for determination of lipase activity in microbial media. The method is based on using phenyl acetate as substrate for lipase and determination of liberated phenol ...A simple, rapid and precise method has been developed for determination of lipase activity in microbial media. The method is based on using phenyl acetate as substrate for lipase and determination of liberated phenol by Folin Ciocalteu reagent. Reaction mixture containing substrate 2.4 ml of phenyl acetate 165 μM in Tris HCl buffer, 0.1 M and pH 7, with 1% (v/v) Triton X-100) and 0.1 ml lipase is incubated at 40?C during 10 minutes and the absorbance was measured at 750 nm. Linearity was observed in the concentration range 0-0.8 g/L lipase.展开更多
Three sensitive,selective and reproducible stability-indicating methods are presented for determination of nitazoxanide (NTZ),a new anti-protozoal drug,in presence of its degradation products.Method A utilizes the fir...Three sensitive,selective and reproducible stability-indicating methods are presented for determination of nitazoxanide (NTZ),a new anti-protozoal drug,in presence of its degradation products.Method A utilizes the first derivative of ratio spectra spectrophotometry by measurement of the amplitude at 364.4 nm using one of the degradation products as a divisor.Method B is a chemometric-assisted spectrophotometry,where principal component regression (PCR) and partial least squares (PLS) were applied.These two approaches were successfully applied to quantify NTZ in presence of degradation products using the information included in the absorption spectra in the range 260-360 nm.Method C is based on the separation of NTZ from its degradation products followed by densitometric measurement of the bands at 254 nm.The separation was carried out on silica gel 60F 254,using chloroform-methanol-ammonia solution-glacial acetic acid (95:5:1:1 by volume,pH=5.80) as a developing system.These methods are suitable as stability-indicating methods for the determination of NTZ in presence of its degradation products either in bulk powder or in pharmaceutical formulations.Statistical analysis of the results has been carried out revealing high accuracy and good precision.展开更多
The present work reports a simple, fast and sensitive microbiological assay applying the turbidimetric method for the determination of ciprofloxacin hydrochloride （CIPRO HC1） in ophthalmic solutions. The validation ...The present work reports a simple, fast and sensitive microbiological assay applying the turbidimetric method for the determination of ciprofloxacin hydrochloride （CIPRO HC1） in ophthalmic solutions. The validation method yielded good results and included excellent linearity, precision, accuracy and specificity. The bioassay is based on the inhibitory effect of CIPRO HC1 upon the strain of Staphylococcus epMermidis ATCC 12228 used as the test microorganism. The results were ar, ated statistically by analysis of variance （ANOVA） and were found to be linear （r=0.9994, in the range of 14.0-56.0 lag/mL）, precise （intraday RSD %=2.06; interday RSD%=2.30） and accurate （recovery = 99.7%）. The turbidimeaic assay was compared to the UV spectrophotometric and I-IPIX2 methods for the same drug. The tuIbidimetric bioassay described on this paper for determination of ciprofioxacin hydrochloride in ophthalmic solution is an alternative to the physicochemical methods disclosed in the literature and can be used in quality control routine.展开更多
Potency is one of the most important indexes of inactivated vaccines.A number of methods have been established to assay the potency,of which the NIH test and single-dose mouse protection test are the "prescribed ...Potency is one of the most important indexes of inactivated vaccines.A number of methods have been established to assay the potency,of which the NIH test and single-dose mouse protection test are the "prescribed methods".Here,we report a method to semi-quantitatively assay the potency of an inactivated rabies vaccine,which uses fewer animals and takes less time to complete.Depending on the quality requirements of a vaccine(e.g.minimum potency),a rabies reference vaccine is,for example,diluted to the minimum potency,and 50 μL of the dilution is taken to inoculate 10 mice.The same amount of the test rabies vaccine is inoculated into another 10 mice.After two weeks,all mice are bled and serum samples are assayed for viral neutralizing antibody by the fluorescent antibody virus neutralization(FAVN) test.By comparing the median and interquartile range of antibody titers of the reference vaccine with those of the test vaccine,the test vaccine potency can be semi-quantitatively judged as to whether it is in accord with the required quality.The reliability of this method was also confirmed in dogs.The procedure can be recommended for batch potency testing during inactivated rabies vaccine production.展开更多
Para-phenylenediamine （PPD）, tile main toxic ingredients of hair dyes, have been used by millions of consumers to improve their appearance. Stone Hair Dye （SHD） mainly contain PPD. SHD and PPD were evaluated by 3-...Para-phenylenediamine （PPD）, tile main toxic ingredients of hair dyes, have been used by millions of consumers to improve their appearance. Stone Hair Dye （SHD） mainly contain PPD. SHD and PPD were evaluated by 3-（4, 5-dimethylthiazol-2-yl）-2, 5-diphenyltetrazdium bromide or MTT assay, which measures mitochondria metabolism in the entire cell culture and provides information about the percentage of cell survival. Utilizing the MTT assay, the cytotoxicity of SHD and PPD was determining on SH Sy5y culture from nervous system of rat. The short term exposure SH Sy5y culture were incubated with various aqueous solutions of different concentrations of SHD and PPD, and the LC50 of SHD and para-phenylenediamine was found to be 9.15 and 12.4 mg/ml. With increasing the concentration, cytotoxicity effect increase in PPD and SHD. There is a significant differences （P〈0.01） of cytotoxicity among concentrations used. It concluded that in vitro assay could give important information of the mechanism of toxicity at low dosages.展开更多
Pepino mosaic virus(PepMV)causes severe disease in tomato and other Solanaceous crops around globe.To effectively study and manage this viral disease,researchers need new,sensitive,and high-throughput approaches for v...Pepino mosaic virus(PepMV)causes severe disease in tomato and other Solanaceous crops around globe.To effectively study and manage this viral disease,researchers need new,sensitive,and high-throughput approaches for viral detection.In this study,we purified PepMV particles from the infected Nicotiana benthamiana plants and used virions to immunize BALB/c mice to prepare hybridomas secreting anti-PepMV monoclonal antibodies(mAbs).A panel of highly specific and sensitive murine mAbs(15B2,8H6,23D11,20D9,3A6,and 8E3)could be produced through cell fusion,antibody selection,and cell cloning.Using the mAbs as the detection antibodies,we established double antibody sandwich enzyme-linked immunosorbent assay(DAS-ELISA),Dot-ELISA,and Tissue print-ELISA for detecting PepMV infection in tomato plants.Resulting data on sensitivity analysis assays showed that both DAS-ELISA and Dot-ELISA can efficiently monitor the virus in PepMV-infected tissue crude extracts when diluted at 1:1310720 and 1:20480(weight/volume ratio(w/v),g/mL),respectively.Among the three methods developed,the Tissue print-ELISA was found to be the most practical detection technique.Survey results from field samples by the established serological approaches were verified by reverse transcription polymerase chain reaction(RT-PCR)and DNA sequencing,dem on strati ng all three serological methods are reliable and effective for monitoring PepMV.An ti-PepMV mAbs and the newly developed DAS-ELISA,Dot-ELISA,and Tissue print-ELISA can benefit PepMV detection and field epidemiological study,and management of this viral disease,which is already widespread in tomato plants in Yunnan Province of China.展开更多
A lock solution composed of gentamicin sulfate(5 mg/mL) and ethylenediaminetetraacetic acid disodium salt(EDTA-Na2, 30 mg/mL) could fully eradicate in vivo bacterial biofilms in totally implantable venous access ports...A lock solution composed of gentamicin sulfate(5 mg/mL) and ethylenediaminetetraacetic acid disodium salt(EDTA-Na2, 30 mg/mL) could fully eradicate in vivo bacterial biofilms in totally implantable venous access ports(TIVAP). In this study, fabrication, conditioning and sterilization processes of antimicrobial lock solution(ALS) were detailed and completed by a stability study. Stability of ALS was conducted for12 months in vial(25 °C 7 2 °C, 60% 7 5% relative humidity(RH), and at 40 °C 7 2 °C, RH 75% 7 5%)and for 24 h and 72 h in TIVAP(40 °C 7 2 °C, RH 75% 7 5%). A stability indicating HPLC assay with UV detection for simultaneous quantification of gentamicin sulfate and EDTA-Na2 was developed. ALS was assayed by ion-pairing high performance liquid chromatography(HPLC) needing gentamicin derivatization, EDTA-Na2 metallocomplexation of samples and gradient mobile phase. HPLC methods to separate four gentamicin components and EDTA-Na2 were validated. Efficiency of sterility procedure and conditioning of ALS was confirmed by bacterial endotoxins and sterility tests. Physicochemical stability of ALS was determined by visual inspection, osmolality, pH, and sub-visible particle counting. Results confirmed that the stability of ALS in vials was maintained for 12 months and 24 h and 72 h in TIVAP.展开更多
A new institutional clinical trial assessed the improvement of sleep disorders in 40 children with autism treated by immediate-release melatonin formulation in different regimens(0.5 mg, 2 mg, and 6 mg daily) for one ...A new institutional clinical trial assessed the improvement of sleep disorders in 40 children with autism treated by immediate-release melatonin formulation in different regimens(0.5 mg, 2 mg, and 6 mg daily) for one month. The objectives of present study were to(i) prepare low-dose melatonin hard capsules for pediatric use controlled by two complementary methods and(ii) carry out a stability study in order to determine a use-bydate. Validation of preparation process was claimed as ascertained by mass uniformity of hard capsules.Multicomponent analysis by attenuated total reflectance Fourier transformed infrared(ATR-FTIR) of melatonin/microcrystalline cellulose mixture allowed to identify and quantify relative content of active pharmaceutical ingredients and excipients. Absolute melatonin content analysis by high performance liquid chromatography in 0.5 mg and 6 mg melatonin capsules was 93.6% ± 4.1% and 98.7% ± 6.9% of theoretical value, respectively. Forced degradation study showed a good separation of melatonin and its degradation products. The capability of the method was 15, confirming a risk of false negative < 0.01%. Stability test and dissolution test were compliant over 18 months of storage with European Pharmacopoeia. Preparation of melatonin hard capsules was completed manually and melatonin in hard capsules was stable for 18 months, in spite of low doses of active ingredient. ATR-FTIR offers a real alternative to HPLC for quality control of highdose melatonin hard capsules before the release of clinical batches.展开更多
Ten actinomycete strains isolated from the Yellow Sea off China＇s coasts were identified as belonging to two genera by 16S rDNA phylogenetic analysis： Streptomyces and Nocardiopsis. Six Streptomyces strains （MA10, ...Ten actinomycete strains isolated from the Yellow Sea off China＇s coasts were identified as belonging to two genera by 16S rDNA phylogenetic analysis： Streptomyces and Nocardiopsis. Six Streptomyces strains （MA10, 2SHXF01-3, MA35, MA05-2, MA05-2-1 and MA08-1） and one Nocardiopsis strain （MA03） were predicted to have the potential to produce aromatic polyketides based on the analysis of the KSa （ketoacyl-synthase） gene in the type II PKS （polyketides synthase） gene cluster. Four strains （MA03, MA01, MA10 and MA05-2） exhibited significant inhibitory effects on mycelia growth （inhibition rate 〉50%） and subsequent aria- toxin production （inhibition rate 〉75%） of the mutant aflatoxigenic Aspergillus parasiticus NFRI-95. The ethyl acetate extracts of the broth of these four strains displayed significant inhibitory effects on mycelia growth, and the IC50 values were calculated （MA03： 0.275 mg mL-1, MA01：0.106 mg mL-1, MA10：1.345 mg mL-1 and MA05-2：1.362 mg mL-1）. Five strains （2SHXF01-3, MA03, MA05-2, MA01 and MA08-1） were selected based on their high cytotoxic activities. The ethyl acetate extract of the Nocardiopsis strain MA03 was particularly noted for its high antitumor activity against human carcinomas of the cervix （HeLa）, lung （A549）, kidney （Caki-1） and liver （HepG2） （IC50： 2.890, 1.981, 3.032 and 2.603 μgmL-1, respectively）. The extract also remarkably inhibited colony formation of HeLa cells at an extremely low concentration （0.5μgmL 1）. This study highlights that marine-derived actinomycetes are a huge resource of compounds for the biological control of aflatoxin contamination and the development of novel drugs for human carcinomas.展开更多
A simple, precise, accurate stability-indicating gradient reversed-phase high-performance liquid chromatographic （RP-HPLC） method was developed for the quantitative determination of zotepine （ZTP） in bulk and phar...A simple, precise, accurate stability-indicating gradient reversed-phase high-performance liquid chromatographic （RP-HPLC） method was developed for the quantitative determination of zotepine （ZTP） in bulk and pharmaceutical dosage forms in the presence of its degradation products （DPs）. The method was developed using Phenomenex C18 column （250 mm ~ 4.6 mm i.d., 5 mm） with a mobile phase containing a gradient mixture of solvents, A （0.05%trifluoroacetic acid （TFA）, pH ？ 3.0） and B （acetonitrile）. The eluted compounds were monitored at 254 nm;the run time was within 20.0 min, in which ZTP and its DPs were well separated, with a resolution of 41.5. The stress testing of ZTP was carried out under acidic, alkaline, neutral hydrolysis, oxidative, photolytic and thermal stress conditions. ZTP was found to degrade significantly in acidic, photolytic, thermal and oxidative stress conditions and remain stable in basic and neutral conditions. The developed method was validated with respect to specificity, linearity, limit of detection, limit of quantification, accuracy, precision and robustness as per ICH guidelines. This method was also suitable for the assay determination of ZTP in pharmaceutical dosage forms. The DPs were characterized by LC-MS/MS and their fragmentation pathways were proposed.展开更多
Standard parenteral nutrition solutions are mixtures comprising interacting components that may degrade themselves over time.The objective of this study was to investigate the physicochemical and microbiological stabi...Standard parenteral nutrition solutions are mixtures comprising interacting components that may degrade themselves over time.The objective of this study was to investigate the physicochemical and microbiological stability of a hospital preparation for parenteral nutrition in neonatology.The analyses were performed throughout the storage of the preparations at 2–8°C(up to 4 months).The extent of stability was based on the determination of amino acids dosage,visual and physicochemical properties(glucose and electrolytes concentrations,pH and osmolality measurements,particle counting)and microbiological analysis(sterility test).A thermal degradation of ascorbic acid was conducted to evaluate the antioxidant properties of the parenteral mixture.Physicochemical and microbiological controls were found to comply with the specifications.Amino acids showed a good stability throughout the 4 months storage except for cysteine,which was progressively degraded to cystine,conferring a yellow coloration to parenteral solutions.Parenteral nutrition standards solutions remain stable for 4 months at 2–8°C,ensuring safe administration in preterm infants.展开更多
Objective:Angiogenesis is the development of new blood vessels.The ion channels on endothelium play a vital action in cell proliferation and so in the related angiogenesis.We aimed to investigate the anti-angiogenic e...Objective:Angiogenesis is the development of new blood vessels.The ion channels on endothelium play a vital action in cell proliferation and so in the related angiogenesis.We aimed to investigate the anti-angiogenic effects of Mefloquine(Cl-channel blocker) and4-Aminopyridine(K+ channel blocker).Methods:The anti-angiogenic activities of Mefloquine and 4-Aminopyridine(4-AP)were investigated by in-vivo(sponge implantation method),in-vitro(aortic ring assay)and in-ovo(CAM,Chick Chorioallantoic membrane) methods.The standard antiangiogenic drug used was Bevacizumab.Results:In the CAM assay,both the ion channel blockers exhibited noticeable antiangiogenic activity at the concentrations of 10-5M and 10-4M where they significantly exhibited ant proliferative activity by inhibiting the new blood vessel formation.For the further confirmation anti-angiogenic activity was evaluated in vitro and in vivo.In Rat aortic ring assay reduction in the area of sprouts were observed with 40 m M of 4-AP and7 m M of Mefloquine.A significant reduction in weight of sponges,number of blood vessels formed and hemoglobin content were observed at 4.2 mg/kg of 4-AP and 20 mg/kg and 30 mg/kg of Mefloquine.Conclusions:These scientific findings indicate the use of Mefloquine and 4-Aminopyridine in pathological situations involving excessive angiogenesis.Negative regulation of cell volume,cell migration and proliferation of blood vessels may be the underlying molecular mechanisms.展开更多
Macitentan (MAC) is a pulmonary arterial hypertension (PAH) drug marketed as a tablet and often has stability issues in the final dosage form. Quantitative determination of MAC and its associated impurities in tablet ...Macitentan (MAC) is a pulmonary arterial hypertension (PAH) drug marketed as a tablet and often has stability issues in the final dosage form. Quantitative determination of MAC and its associated impurities in tablet dosage form has not been previously reported. This study quantified impurities present in Macitentan tablets using a binary solvent-based gradient elution method using reversed phase-high performance liquid chromatography.The developed method w as validated per International Conference on Harmonization (ICH) guidelines and the drug product w as subjected to forced degradation studies to evaluate stability. The developed method efficiently separated the drug and impurities (48 min) w ithout interference from solvents,excipients,or other impurities. The developed method met all guidelines in all characteristics w ith recoveries ranging from 85%-115%,linearity w ith r 2≥0. 996 6,and substantial robustness. The stability-indicating nature of the method w as evaluated using stressed conditions (hydrolysis:1 N HCl at 80℃/15 min; 1 N NaOH at 25℃/45 min; humidity stress (90%relative humidity) at 25℃for 24 h,oxidation:at 6%(v/v) H2O2,80℃/15 min,thermolysis:at105℃/16 h and photolysis:UV light at 200 Wh/m2; Fluorescent light at 1. 2 million luxh). Forced degradation experiments show ed that the developed method w as effective for impurity profiling. All stressed samples w ere assayed and mass balance w as> 96%. Forced degradation results indicated that MAC tablets w ere sensitive to hydrolysis (acid and alkali) and thermal conditions. The developed method is suitable for both assay and impurity determination,w hich is applicable to the pharmaceutical industry.展开更多
SDS-PAGE was applied to determine trypsin activity and inhibition. After the hydrolysis by trypsin to substrate bovine serum albulnin (BSA) under different temperatures and pH, the hydrolysis degree of BSA was conduct...SDS-PAGE was applied to determine trypsin activity and inhibition. After the hydrolysis by trypsin to substrate bovine serum albulnin (BSA) under different temperatures and pH, the hydrolysis degree of BSA was conducted using SDS-PAGE. From the quantitative analysis to the electrophoresis bands of BSA and its hydrolysis products in SDS-PAGE pattern, the change of trypsin activity was determined, and then the optimum temperature at 40°C and the optimum pH at pH 8.5 - 8.7 for trypsin activity were obtained. All the target bonds in BSA molecule could be hydrolyzed at the same time by trypsin. The inhibition was due to the binding of inhibitor to trypsin, which made it impossible for trypsin to touch the substrate protein. SDS-PAGE was demonstrated to be also an effect method for assaying the characteristics of trypsin activity and its inhibition.展开更多
The pathogenic effect of Staphylococci is due to extra-cellular factors and properties such as adherence and biofilm production. The nature of the biofilm and the physiological properties of biofilm-producing bacteria...The pathogenic effect of Staphylococci is due to extra-cellular factors and properties such as adherence and biofilm production. The nature of the biofilm and the physiological properties of biofilm-producing bacteria result in an inherent antibiotic resistance and require further investigation. Two hundred and sixty Staphylococcal strains were cultured from 600 clinical specimens obtained from hospitalized patients. Among these, 155 were identified as coagulase-positive (CPS) and 105 as coagulase-negative (CNS) staphylococci. Staphylococcal strains were tested for biofilm production using the tissue culture plate (TCP) method. TCP detection showed that of the 155 CPS, 124 (80%) were biofilm producers, while 63 (60%) of the 105 CNS were biofilm producers. Biofilm-producing strains were scanned by scanning electron microscope (SEM) to confirm biofilm formation, study biofilm production, and examine antibiotic effects on biofilm formation. Disc diffusion method was used to study resistance of planktonic and biofilm-forming cells to antibiotics. Planktonic cells were less resistant to antibiotics than biofilm-forming cells. Microbroth dilution method and a new BioTimer assay were used to determine antibiotic MICs affecting planktonic and biofilm cells. Both methods showed that the MICs for planktonic cells were less than that for biofilm cells. The BioTimer assay was therefore found to be sensitive, accurate, and reliable, with results in agreement with those from the broth dilution method and SEM.展开更多
Commelina diffusa Burm. F. is a herbaceous tropical plant with different traditional medicinal uses. Present study is aimed to isolate the bioactive compounds from DCM-Methanol extract of the powdered whole plant of C...Commelina diffusa Burm. F. is a herbaceous tropical plant with different traditional medicinal uses. Present study is aimed to isolate the bioactive compounds from DCM-Methanol extract of the powdered whole plant of Commelina diffusa and to investigate the cytotoxic, antimicrobial and antioxidant activities of the crude extract and its twelve vacuum liquid chromatographic fraction (CD1-12). Vacuum Liquid Chromatography (VLC) was employed to isolate bioactive metabolites. The physicochemical properties of the isolated compound were examined and the molecular structure was elucidated by NMR spectroscopy.?Cytotoxic, antibacterial, antioxidant activities were evaluated following brine shrimp lethality bioassay, disc diffusion method and DPPH free radical scavenging assay respectively. The isolated compound was identified as steroid (stigmasterol) which had significant cytotoxic effect on vero cell line. The crude extract and its fractions (CD8-CD12) exhibited strong cytotoxicity in brine shrimp lethality bioassay having LC50 values 3.79, 9.19, 29.49, 16.60, 19.36, 44.58 μg/mL respectively. The crude extract showed mild to strong antibacterial activity. Fractions (CD5-CD12) showed mild to strong antibacterial activity against Escherichia coli in comparison with Kanamycin (standard). Strong antioxidant activities were found in crude extract (IC50 value 30.52 μg/ml), CD11 (IC50 value 39.27 μg/ml) and CD12 (IC50 value 19.50 μg/ml). The study suggests Commelina diffusa plant extract to have strong antioxidant and cytotoxic activity which is indicative of presence of compounds with broad spectrum of curative applications. One compound namely stigmasterol has been isolated from the plant.展开更多
文摘The stability of the drug actarit was studied under different stress conditions like hydrolysis（acid,alkaline and neutral）,oxidation,photolysis and thermal degradation as recommended hy International Conference on Harmonization（ICH） guidelines.Drug was found to be unstable in acidic,basic and photolytic conditions and produced a common degradation product while oxidative stress condition produced three additional degradation products.Drug was impassive to neutral hydrolysis,dry thermal and accelerated stability conditions.Degradation products were identified,isolated and characterized by different spectroscopic analyses.Drug and the degradation products were synthesized by a new route using green chemistry.The chromatographic separation of the drug and its impurities was achieved in a phenomenex luna C18 column employing a step gradient elution by high performance liquid chromatography coupled to photodiode array and mass spectrometry detectors（HPLC-PDAMS）.A specilic and sensitive stability-indicating assay method for the simultaneous determination of the drug actarit.its process related impurities and degradation products was developed and validated.
文摘A simple, rapid, precise, accurate, rugged and robust stability-indicating ultra-fast high performance liquid chromatographic (UHPLC) method has been developed for the estimation of related compounds (imp-A, imp-B, imp-C, imp-D and imp-E) in Naproxen and also the assay of Naproxen from bulk drug samples. The stability indicating capability of the method was proven by subjecting the samples to stress conditions such as acid, base, oxidation, photolysis and thermal degradation. The efficient chromatographic separation was achieved using mobile phase solution A prepared as buffer solution 10 mM monobasic potassium phosphate pH 4.0 ± 0.05 adjusted with diluted ortho phosphoric acid solution and solution B acetonitrile with linear gradient elution on poroshell 120 EC-C18 shot column (50 mm × 4.6 mm, 2.7 μm) and UV detection at 235 nm at a flow rate 1.0 mL/min, column oven temperature was set to 25?C. The above are all known impurities and degradation impurities are well resolved with Naproxen peak and these are eluted within a 10 min runtime of HPLC. The photo diode array detector was used for peak homogeneity testing during stress study experiments and the overall mass balance was found to be 99.2% to 100.2% in all stress conditions. The linear calibration range was found to be 0.05 μg/mL to 0.75 μg/mL for related compounds and 50 μg/mL to 150 μg/mL for Naproxen and the accuracy of the method was found to be 91.5% to 98.5% recovery for the related substance method and 95.4% to 97.4% recovery for the assay method. The Naproxen and related compounds were found to be stable up to 48 hours and the method validation data show excellent results for precision, linearity, specificity, limit of detection, limit of quantitation and robustness. The present method can be successfully used for routine QC and stability studies and it will help to reduce the analysis cost, time and effluent load compared to conventional HPLC methods.
基金Project (No. 30200266) supported by the National Natural Science Foundation of China
文摘A patented kinetic uricase method was evaluated for serum uric acid assay. Initial absorbance of the reaction mixture before uricase action （A0） was obtained by correcting the absorbance at 293 nm measured before the addition of uricase solution, and background absorbance （Ab） was predicted by an integrated method. Uric acid concentration in reaction solution was calculated from AA, the difference between A0 and Ab, using the absorptivity preset for uric acid. This kinetic uricase method exhibited CV〈4.3% and recovery of 100%. Lipids, bilirubin, hemoglobin, ascorbic acid, reduced glutathione and xanthine 〈0.32 mmol/L in serum had no significant effects. △A linearly responded to 1.2 to 37.5 μmol/L uric acid in reaction solution containing 15 μl serum. The slope of linear response was consistent with the absorptivity preset for uric acid while the intercept was consistent with that for serum alone. Uric acid concentrations in clinic sera by different uricase methods positively correlated to each other. By Bland-Altman analysis, this kinetic uricase method accorded with that by quantifying the total change of UV absorbance on the completion of uricase reaction. These results demonstrated that this kinetic uricase method is reliable for serum uric acid assay with enhanced resistance to both xanthine and other common errors, wider range of linear response and much lower cost.
文摘A new and senseitive Co (II)-biuret method for determination of protein has for the first time been established. In visible wavelength region, the Co (II) -protein complexes had anabsorption peak at 360 nm. -Under the present conditions. the assay had a linear range of 0-120 μg/ml of protein with a detection limit of about 2 μg/ml, and similar calibration curves were obtained for BSA. and BγG. Compared with Cu (II) -biuret method, the present Co (II) -biuret method was more simple, sensitive, and tolerant of interferences from non-protein molecules.
文摘A simple, rapid and precise method has been developed for determination of lipase activity in microbial media. The method is based on using phenyl acetate as substrate for lipase and determination of liberated phenol by Folin Ciocalteu reagent. Reaction mixture containing substrate 2.4 ml of phenyl acetate 165 μM in Tris HCl buffer, 0.1 M and pH 7, with 1% (v/v) Triton X-100) and 0.1 ml lipase is incubated at 40?C during 10 minutes and the absorbance was measured at 750 nm. Linearity was observed in the concentration range 0-0.8 g/L lipase.
文摘Three sensitive,selective and reproducible stability-indicating methods are presented for determination of nitazoxanide (NTZ),a new anti-protozoal drug,in presence of its degradation products.Method A utilizes the first derivative of ratio spectra spectrophotometry by measurement of the amplitude at 364.4 nm using one of the degradation products as a divisor.Method B is a chemometric-assisted spectrophotometry,where principal component regression (PCR) and partial least squares (PLS) were applied.These two approaches were successfully applied to quantify NTZ in presence of degradation products using the information included in the absorption spectra in the range 260-360 nm.Method C is based on the separation of NTZ from its degradation products followed by densitometric measurement of the bands at 254 nm.The separation was carried out on silica gel 60F 254,using chloroform-methanol-ammonia solution-glacial acetic acid (95:5:1:1 by volume,pH=5.80) as a developing system.These methods are suitable as stability-indicating methods for the determination of NTZ in presence of its degradation products either in bulk powder or in pharmaceutical formulations.Statistical analysis of the results has been carried out revealing high accuracy and good precision.
基金supported by PACD-FCFAr-UNESP(AraraquaraBrazil)+9 种基金FAPESP(Sao PauloBrazil)FUNDUNEP(Sao PauloBrazil)CNPq(BrasíliaBrazil)E.C.L.Cazedey was funded by CAPES(BrasíliaBrazil)H.R.N.Salgado was funded by CNPq(BrasíliaBrazil)
文摘The present work reports a simple, fast and sensitive microbiological assay applying the turbidimetric method for the determination of ciprofloxacin hydrochloride （CIPRO HC1） in ophthalmic solutions. The validation method yielded good results and included excellent linearity, precision, accuracy and specificity. The bioassay is based on the inhibitory effect of CIPRO HC1 upon the strain of Staphylococcus epMermidis ATCC 12228 used as the test microorganism. The results were ar, ated statistically by analysis of variance （ANOVA） and were found to be linear （r=0.9994, in the range of 14.0-56.0 lag/mL）, precise （intraday RSD %=2.06; interday RSD%=2.30） and accurate （recovery = 99.7%）. The turbidimeaic assay was compared to the UV spectrophotometric and I-IPIX2 methods for the same drug. The tuIbidimetric bioassay described on this paper for determination of ciprofioxacin hydrochloride in ophthalmic solution is an alternative to the physicochemical methods disclosed in the literature and can be used in quality control routine.
基金the China National"863"Program(Approval No.2011AA10A212)Special Fund for Agro-Scientific Research in the Public Interest(ApprovalNo.201203056)
文摘Potency is one of the most important indexes of inactivated vaccines.A number of methods have been established to assay the potency,of which the NIH test and single-dose mouse protection test are the "prescribed methods".Here,we report a method to semi-quantitatively assay the potency of an inactivated rabies vaccine,which uses fewer animals and takes less time to complete.Depending on the quality requirements of a vaccine(e.g.minimum potency),a rabies reference vaccine is,for example,diluted to the minimum potency,and 50 μL of the dilution is taken to inoculate 10 mice.The same amount of the test rabies vaccine is inoculated into another 10 mice.After two weeks,all mice are bled and serum samples are assayed for viral neutralizing antibody by the fluorescent antibody virus neutralization(FAVN) test.By comparing the median and interquartile range of antibody titers of the reference vaccine with those of the test vaccine,the test vaccine potency can be semi-quantitatively judged as to whether it is in accord with the required quality.The reliability of this method was also confirmed in dogs.The procedure can be recommended for batch potency testing during inactivated rabies vaccine production.
文摘Para-phenylenediamine （PPD）, tile main toxic ingredients of hair dyes, have been used by millions of consumers to improve their appearance. Stone Hair Dye （SHD） mainly contain PPD. SHD and PPD were evaluated by 3-（4, 5-dimethylthiazol-2-yl）-2, 5-diphenyltetrazdium bromide or MTT assay, which measures mitochondria metabolism in the entire cell culture and provides information about the percentage of cell survival. Utilizing the MTT assay, the cytotoxicity of SHD and PPD was determining on SH Sy5y culture from nervous system of rat. The short term exposure SH Sy5y culture were incubated with various aqueous solutions of different concentrations of SHD and PPD, and the LC50 of SHD and para-phenylenediamine was found to be 9.15 and 12.4 mg/ml. With increasing the concentration, cytotoxicity effect increase in PPD and SHD. There is a significant differences （P〈0.01） of cytotoxicity among concentrations used. It concluded that in vitro assay could give important information of the mechanism of toxicity at low dosages.
基金National Key R&D Program of China(Nos.2019YFD1001800 and 2017YFD0201604)the National Natural Science Foundation of China(Nos.31772125 and 31972234)。
文摘Pepino mosaic virus(PepMV)causes severe disease in tomato and other Solanaceous crops around globe.To effectively study and manage this viral disease,researchers need new,sensitive,and high-throughput approaches for viral detection.In this study,we purified PepMV particles from the infected Nicotiana benthamiana plants and used virions to immunize BALB/c mice to prepare hybridomas secreting anti-PepMV monoclonal antibodies(mAbs).A panel of highly specific and sensitive murine mAbs(15B2,8H6,23D11,20D9,3A6,and 8E3)could be produced through cell fusion,antibody selection,and cell cloning.Using the mAbs as the detection antibodies,we established double antibody sandwich enzyme-linked immunosorbent assay(DAS-ELISA),Dot-ELISA,and Tissue print-ELISA for detecting PepMV infection in tomato plants.Resulting data on sensitivity analysis assays showed that both DAS-ELISA and Dot-ELISA can efficiently monitor the virus in PepMV-infected tissue crude extracts when diluted at 1:1310720 and 1:20480(weight/volume ratio(w/v),g/mL),respectively.Among the three methods developed,the Tissue print-ELISA was found to be the most practical detection technique.Survey results from field samples by the established serological approaches were verified by reverse transcription polymerase chain reaction(RT-PCR)and DNA sequencing,dem on strati ng all three serological methods are reliable and effective for monitoring PepMV.An ti-PepMV mAbs and the newly developed DAS-ELISA,Dot-ELISA,and Tissue print-ELISA can benefit PepMV detection and field epidemiological study,and management of this viral disease,which is already widespread in tomato plants in Yunnan Province of China.
基金supported by Centre de Recherche Translationnelle de I'Institut Pasteur, grant Number S- PI15007-02Asupported by the French Government's Investissement d'Avenir program:Laboratoire d'Excellence ‘Integrative Biology of Emerging Infectious Diseases’ (grant no. ANR-10-LABX62-IBEID.)+1 种基金the Fondation pour la Recherche Médicale (grant no. DEQ. 20140329508)the Center for Translational Science of the Institut Pasteur (S-PI15007-02A)
文摘A lock solution composed of gentamicin sulfate(5 mg/mL) and ethylenediaminetetraacetic acid disodium salt(EDTA-Na2, 30 mg/mL) could fully eradicate in vivo bacterial biofilms in totally implantable venous access ports(TIVAP). In this study, fabrication, conditioning and sterilization processes of antimicrobial lock solution(ALS) were detailed and completed by a stability study. Stability of ALS was conducted for12 months in vial(25 °C 7 2 °C, 60% 7 5% relative humidity(RH), and at 40 °C 7 2 °C, RH 75% 7 5%)and for 24 h and 72 h in TIVAP(40 °C 7 2 °C, RH 75% 7 5%). A stability indicating HPLC assay with UV detection for simultaneous quantification of gentamicin sulfate and EDTA-Na2 was developed. ALS was assayed by ion-pairing high performance liquid chromatography(HPLC) needing gentamicin derivatization, EDTA-Na2 metallocomplexation of samples and gradient mobile phase. HPLC methods to separate four gentamicin components and EDTA-Na2 were validated. Efficiency of sterility procedure and conditioning of ALS was confirmed by bacterial endotoxins and sterility tests. Physicochemical stability of ALS was determined by visual inspection, osmolality, pH, and sub-visible particle counting. Results confirmed that the stability of ALS in vials was maintained for 12 months and 24 h and 72 h in TIVAP.
文摘A new institutional clinical trial assessed the improvement of sleep disorders in 40 children with autism treated by immediate-release melatonin formulation in different regimens(0.5 mg, 2 mg, and 6 mg daily) for one month. The objectives of present study were to(i) prepare low-dose melatonin hard capsules for pediatric use controlled by two complementary methods and(ii) carry out a stability study in order to determine a use-bydate. Validation of preparation process was claimed as ascertained by mass uniformity of hard capsules.Multicomponent analysis by attenuated total reflectance Fourier transformed infrared(ATR-FTIR) of melatonin/microcrystalline cellulose mixture allowed to identify and quantify relative content of active pharmaceutical ingredients and excipients. Absolute melatonin content analysis by high performance liquid chromatography in 0.5 mg and 6 mg melatonin capsules was 93.6% ± 4.1% and 98.7% ± 6.9% of theoretical value, respectively. Forced degradation study showed a good separation of melatonin and its degradation products. The capability of the method was 15, confirming a risk of false negative < 0.01%. Stability test and dissolution test were compliant over 18 months of storage with European Pharmacopoeia. Preparation of melatonin hard capsules was completed manually and melatonin in hard capsules was stable for 18 months, in spite of low doses of active ingredient. ATR-FTIR offers a real alternative to HPLC for quality control of highdose melatonin hard capsules before the release of clinical batches.
基金supported by the COMRA project (No. DY125-15-R-01)
文摘Ten actinomycete strains isolated from the Yellow Sea off China＇s coasts were identified as belonging to two genera by 16S rDNA phylogenetic analysis： Streptomyces and Nocardiopsis. Six Streptomyces strains （MA10, 2SHXF01-3, MA35, MA05-2, MA05-2-1 and MA08-1） and one Nocardiopsis strain （MA03） were predicted to have the potential to produce aromatic polyketides based on the analysis of the KSa （ketoacyl-synthase） gene in the type II PKS （polyketides synthase） gene cluster. Four strains （MA03, MA01, MA10 and MA05-2） exhibited significant inhibitory effects on mycelia growth （inhibition rate 〉50%） and subsequent aria- toxin production （inhibition rate 〉75%） of the mutant aflatoxigenic Aspergillus parasiticus NFRI-95. The ethyl acetate extracts of the broth of these four strains displayed significant inhibitory effects on mycelia growth, and the IC50 values were calculated （MA03： 0.275 mg mL-1, MA01：0.106 mg mL-1, MA10：1.345 mg mL-1 and MA05-2：1.362 mg mL-1）. Five strains （2SHXF01-3, MA03, MA05-2, MA01 and MA08-1） were selected based on their high cytotoxic activities. The ethyl acetate extract of the Nocardiopsis strain MA03 was particularly noted for its high antitumor activity against human carcinomas of the cervix （HeLa）, lung （A549）, kidney （Caki-1） and liver （HepG2） （IC50： 2.890, 1.981, 3.032 and 2.603 μgmL-1, respectively）. The extract also remarkably inhibited colony formation of HeLa cells at an extremely low concentration （0.5μgmL 1）. This study highlights that marine-derived actinomycetes are a huge resource of compounds for the biological control of aflatoxin contamination and the development of novel drugs for human carcinomas.
文摘A simple, precise, accurate stability-indicating gradient reversed-phase high-performance liquid chromatographic （RP-HPLC） method was developed for the quantitative determination of zotepine （ZTP） in bulk and pharmaceutical dosage forms in the presence of its degradation products （DPs）. The method was developed using Phenomenex C18 column （250 mm ~ 4.6 mm i.d., 5 mm） with a mobile phase containing a gradient mixture of solvents, A （0.05%trifluoroacetic acid （TFA）, pH ？ 3.0） and B （acetonitrile）. The eluted compounds were monitored at 254 nm;the run time was within 20.0 min, in which ZTP and its DPs were well separated, with a resolution of 41.5. The stress testing of ZTP was carried out under acidic, alkaline, neutral hydrolysis, oxidative, photolytic and thermal stress conditions. ZTP was found to degrade significantly in acidic, photolytic, thermal and oxidative stress conditions and remain stable in basic and neutral conditions. The developed method was validated with respect to specificity, linearity, limit of detection, limit of quantification, accuracy, precision and robustness as per ICH guidelines. This method was also suitable for the assay determination of ZTP in pharmaceutical dosage forms. The DPs were characterized by LC-MS/MS and their fragmentation pathways were proposed.
文摘Standard parenteral nutrition solutions are mixtures comprising interacting components that may degrade themselves over time.The objective of this study was to investigate the physicochemical and microbiological stability of a hospital preparation for parenteral nutrition in neonatology.The analyses were performed throughout the storage of the preparations at 2–8°C(up to 4 months).The extent of stability was based on the determination of amino acids dosage,visual and physicochemical properties(glucose and electrolytes concentrations,pH and osmolality measurements,particle counting)and microbiological analysis(sterility test).A thermal degradation of ascorbic acid was conducted to evaluate the antioxidant properties of the parenteral mixture.Physicochemical and microbiological controls were found to comply with the specifications.Amino acids showed a good stability throughout the 4 months storage except for cysteine,which was progressively degraded to cystine,conferring a yellow coloration to parenteral solutions.Parenteral nutrition standards solutions remain stable for 4 months at 2–8°C,ensuring safe administration in preterm infants.
文摘Objective:Angiogenesis is the development of new blood vessels.The ion channels on endothelium play a vital action in cell proliferation and so in the related angiogenesis.We aimed to investigate the anti-angiogenic effects of Mefloquine(Cl-channel blocker) and4-Aminopyridine(K+ channel blocker).Methods:The anti-angiogenic activities of Mefloquine and 4-Aminopyridine(4-AP)were investigated by in-vivo(sponge implantation method),in-vitro(aortic ring assay)and in-ovo(CAM,Chick Chorioallantoic membrane) methods.The standard antiangiogenic drug used was Bevacizumab.Results:In the CAM assay,both the ion channel blockers exhibited noticeable antiangiogenic activity at the concentrations of 10-5M and 10-4M where they significantly exhibited ant proliferative activity by inhibiting the new blood vessel formation.For the further confirmation anti-angiogenic activity was evaluated in vitro and in vivo.In Rat aortic ring assay reduction in the area of sprouts were observed with 40 m M of 4-AP and7 m M of Mefloquine.A significant reduction in weight of sponges,number of blood vessels formed and hemoglobin content were observed at 4.2 mg/kg of 4-AP and 20 mg/kg and 30 mg/kg of Mefloquine.Conclusions:These scientific findings indicate the use of Mefloquine and 4-Aminopyridine in pathological situations involving excessive angiogenesis.Negative regulation of cell volume,cell migration and proliferation of blood vessels may be the underlying molecular mechanisms.
基金the management of Sinotherapeutics Inc. for supporting this study
文摘Macitentan (MAC) is a pulmonary arterial hypertension (PAH) drug marketed as a tablet and often has stability issues in the final dosage form. Quantitative determination of MAC and its associated impurities in tablet dosage form has not been previously reported. This study quantified impurities present in Macitentan tablets using a binary solvent-based gradient elution method using reversed phase-high performance liquid chromatography.The developed method w as validated per International Conference on Harmonization (ICH) guidelines and the drug product w as subjected to forced degradation studies to evaluate stability. The developed method efficiently separated the drug and impurities (48 min) w ithout interference from solvents,excipients,or other impurities. The developed method met all guidelines in all characteristics w ith recoveries ranging from 85%-115%,linearity w ith r 2≥0. 996 6,and substantial robustness. The stability-indicating nature of the method w as evaluated using stressed conditions (hydrolysis:1 N HCl at 80℃/15 min; 1 N NaOH at 25℃/45 min; humidity stress (90%relative humidity) at 25℃for 24 h,oxidation:at 6%(v/v) H2O2,80℃/15 min,thermolysis:at105℃/16 h and photolysis:UV light at 200 Wh/m2; Fluorescent light at 1. 2 million luxh). Forced degradation experiments show ed that the developed method w as effective for impurity profiling. All stressed samples w ere assayed and mass balance w as> 96%. Forced degradation results indicated that MAC tablets w ere sensitive to hydrolysis (acid and alkali) and thermal conditions. The developed method is suitable for both assay and impurity determination,w hich is applicable to the pharmaceutical industry.
文摘SDS-PAGE was applied to determine trypsin activity and inhibition. After the hydrolysis by trypsin to substrate bovine serum albulnin (BSA) under different temperatures and pH, the hydrolysis degree of BSA was conducted using SDS-PAGE. From the quantitative analysis to the electrophoresis bands of BSA and its hydrolysis products in SDS-PAGE pattern, the change of trypsin activity was determined, and then the optimum temperature at 40°C and the optimum pH at pH 8.5 - 8.7 for trypsin activity were obtained. All the target bonds in BSA molecule could be hydrolyzed at the same time by trypsin. The inhibition was due to the binding of inhibitor to trypsin, which made it impossible for trypsin to touch the substrate protein. SDS-PAGE was demonstrated to be also an effect method for assaying the characteristics of trypsin activity and its inhibition.
文摘The pathogenic effect of Staphylococci is due to extra-cellular factors and properties such as adherence and biofilm production. The nature of the biofilm and the physiological properties of biofilm-producing bacteria result in an inherent antibiotic resistance and require further investigation. Two hundred and sixty Staphylococcal strains were cultured from 600 clinical specimens obtained from hospitalized patients. Among these, 155 were identified as coagulase-positive (CPS) and 105 as coagulase-negative (CNS) staphylococci. Staphylococcal strains were tested for biofilm production using the tissue culture plate (TCP) method. TCP detection showed that of the 155 CPS, 124 (80%) were biofilm producers, while 63 (60%) of the 105 CNS were biofilm producers. Biofilm-producing strains were scanned by scanning electron microscope (SEM) to confirm biofilm formation, study biofilm production, and examine antibiotic effects on biofilm formation. Disc diffusion method was used to study resistance of planktonic and biofilm-forming cells to antibiotics. Planktonic cells were less resistant to antibiotics than biofilm-forming cells. Microbroth dilution method and a new BioTimer assay were used to determine antibiotic MICs affecting planktonic and biofilm cells. Both methods showed that the MICs for planktonic cells were less than that for biofilm cells. The BioTimer assay was therefore found to be sensitive, accurate, and reliable, with results in agreement with those from the broth dilution method and SEM.
文摘Commelina diffusa Burm. F. is a herbaceous tropical plant with different traditional medicinal uses. Present study is aimed to isolate the bioactive compounds from DCM-Methanol extract of the powdered whole plant of Commelina diffusa and to investigate the cytotoxic, antimicrobial and antioxidant activities of the crude extract and its twelve vacuum liquid chromatographic fraction (CD1-12). Vacuum Liquid Chromatography (VLC) was employed to isolate bioactive metabolites. The physicochemical properties of the isolated compound were examined and the molecular structure was elucidated by NMR spectroscopy.?Cytotoxic, antibacterial, antioxidant activities were evaluated following brine shrimp lethality bioassay, disc diffusion method and DPPH free radical scavenging assay respectively. The isolated compound was identified as steroid (stigmasterol) which had significant cytotoxic effect on vero cell line. The crude extract and its fractions (CD8-CD12) exhibited strong cytotoxicity in brine shrimp lethality bioassay having LC50 values 3.79, 9.19, 29.49, 16.60, 19.36, 44.58 μg/mL respectively. The crude extract showed mild to strong antibacterial activity. Fractions (CD5-CD12) showed mild to strong antibacterial activity against Escherichia coli in comparison with Kanamycin (standard). Strong antioxidant activities were found in crude extract (IC50 value 30.52 μg/ml), CD11 (IC50 value 39.27 μg/ml) and CD12 (IC50 value 19.50 μg/ml). The study suggests Commelina diffusa plant extract to have strong antioxidant and cytotoxic activity which is indicative of presence of compounds with broad spectrum of curative applications. One compound namely stigmasterol has been isolated from the plant.
