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Evaluation of Preservative Effectiveness in Antacid, Cough Syrup and Ophthalmic Solution by Microbial Challenge Test(International Journal of Pharmacognosy) (Nishant Dafale, Uttam Semwal, Piyush Agarwal, Pradip Sharma, G. N. Singh, 1(3),, 193-199,Year : 2014)
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Pharmaceutical preparations having high water content face the problem of microbial spoilage which affects consumer safety. Control on such problem is generally done by addition of specific preservative to these pharmaceutical preparations, but sometimes some physical changes and clinical hazards are observed even after addition of preservatives. The aim of present study was to evaluate and compare the effectiveness of preservatives in market samples of antacids, cough syrups and ophthalmic solutions through microbial challenge test. The samples of antacids, cough syrups and ophthalmic solutions were challenged with 3 bacterial and 2 fungal strains and results were periodically (0, 7, 14 and 28 day) investigated. The number of survive microorganisms were determined using standard microbiological dilution pour-plate technique. More than 1 log reduction of microbial counts was observed in all samples at 7 day. Moreover, the log reduction in microbial counts was significantly increased up to 28 day. Results showed that preservatives Sorbitol, Sodium Citrate, and Benzalkonium Chloride present in antacids, cough syrups and ophthalmic solutions respectively were effective against all the challenged microorganisms. Benzalkonium chloride as a preservative in ophthalmic solution was found to be most effective. Hence, from this study it is concluded that preservatives present in all tested liquid pharmaceutical preparations are effective in preventing contamination of the product during their use and storage.
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Development and validation of microbial bioassay for the quantification of potency of the?antibioticcefuroxime axetil(Analytical Methods (Royal Society of Chemistry, UK)) (Nishant Dafale, Uttam Semawl, Piyush Agarwal, G. N. Singh
,5,,690-698,Year : 2013)
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Cefuroxime is a semi-synthetic antibiotic of the cephalosporin group and has broad-spectrum antimicrobial activity. HPLC methods, detailed in various pharmacopoeias, are generally used for the quantitative determination of the potency of cefuroxime but are unable to determine its bioactivity against microorganisms. In comparison to HPLC methods bioassay methods are simple and inexpensive however, a bioassay for the determination of both the potency and bioactivity of cefuroxime has not yet been reported in any pharmacopoeia. This article is focused on the development of a bioassay method for the quantification of cefuroxime axetil in pharmaceutical preparations. The bioassay was performed with different microbial strains and Kocuria rhizophila ATCC-9341 was selected as the most susceptible microorganism against cefuroxime axetil. The percent potency of market samples were estimated by a two-level bioassay method. The potency of market samples of Biocef, Ceftech and Ceftum were found to be 100.71% (RSD 1.09%), 101.07% (RSD 1.16%) and 100.44% (RSD 1.08%), respectively. The Biocef intermediate precision RSD between days and between analysts were 1.08% and 1.01%, respectively. The specificity of the bioassay for the analysis of Biocef was also studied in parallel with the Pharmacopoeial HPLC method and the potency was calculated as 101.09%. Results show that a bioassay method can be used for the quality control of cefuroxime axetil in raw materials and pharmaceutical preparations.
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Quantification of ceftriaxone sodium in pharmaceutical preparations by a new validated microbiological bioassay(Analytical Methods (Royal Society of Chemistry, UK)) (Nishant Dafale, Uttam Semawl, Piyush Agarwal, Pradeep Sharma, G. N. Singh
,4,,2490-2498,Year : 2012)
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Ceftriaxone is a broad spectrum antibiotic which belongs to the third generation of cephalosporin group. A simple, accurate, precise, reproducible and cost effective one level agar diffusion (5 + 1) bioassay was developed and validated for the estimation of potency and bioactivity of ceftriaxone sodium in pharmaceutical preparations. Among 15 strains of bacteria and 3 strains of fungi, Kocuria rhizophila ATCC-9341 was selected as the most effective organism against ceftriaxone sodium. Optimization of the bioassay was performed by investigating several factors such as buffer pH, inoculums concentration and standard solution concentration. The developed method was successfully validated with respect to linearity, precision and accuracy. The mean potency recovery value for ceftriaxone sodium in commercial dry powder injection of Monocef was 100.44%. An evaluated validation method showed linearity (r2 ¼ 0.986), precision (interday R.S.D ¼ 1.03%, between analyst R.S.D ¼ 1.00%) and accuracy 101.17% (R.S.D ¼ 0.80%). Identification of Monocef was performed by FTIR spectroscopy. The potency of the microbial bioassay was correlated with the HPLC method using the same commercial sample (Monocef). The obtained percent potency was 101.39% by the HPLC method. Results show that the bioassay can be applicable for quantitative determination of the potency with bioactivity in commercial formulations of ceftriaxone sodium.
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Exploration of genetic information from dynamic microbial population for efficiency enhancement of azo dye degrading system (Nishant Dafale
,19,,310-323,Year : 2011)
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Dye degradation is presently an important area of scientific activity. Today, most wastewater treatment makes use of the conventional processes in the presence and action of a native microbial population. However, the potential natural microbial population and genetically engineered microorganisms (GEMs) could successfully bioaugment dye biotreatment systems to enhance efficiency. Consequently, treatment facilities are designed to maintain a high density of the desired microbial population to satisfy the bioremediation demand. Nevertheless, malefactions resulting in a decrease of activity are frequent. To better understand the function of the bacterial community, a full description of the microbial population is required. The prominent task of the microbiologist is to compare the structure, dynamics, and function of the existing microbial populations. Even though the last decade has seen a revolution in microbiology, microbial population monitoring still relies on the tools that were available at the beginning of this century. It is the goal of this review to explain the potential and importance of the newly available molecular tools for analyzing microbial populations. Molecular techniques over the last few decades have revealed an enormous reservoir of unexplained microbes. This large genetic diversity has an immense potential to be used as a resource for the development of novel biotransformations, bioremediation processes, and bioenergy generation. This paper will review bioremediation and the exploration of genetic information from microbial populations for efficiency enhancement.
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Selection of indicator bacteria based on screening of 16S rDNA metagenomic library from a two-stage anoxic-oxic bioreactor system for azo dyes decolorization(Bioresource Technology) (Nishant Dafale, Leena Agrawal, A. Kapley, Sudhir Meshram, H. Purohit & Satish Wate
,101,,476–484,Year : 2010)
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Dye degradation has gained attention of late due to indiscriminate disposal from user industries. Enhancing efficiency of biological treatment provides a cheaper alternative vis-à-vis other advanced technologies. Dye molecules are metabolized biologically via anoxic and oxic treatments. In this study, bacterial community surviving on dye effluent working in anoxic–oxic bioreactor was analyzed using 16S rDNA approach. Azo-dye decolorizing and degrading bacterial community was enriched in lab-scale two-stage anoxic–oxic bioreactor. 16S rDNA metagenomic libraries of enriched population were constructed, screened and phylogenetically analyzed separately. Removal of ∼35% COD with complete decolorization was observed in anoxic bioreactor. Process was carried out by uncultured gamma proteobacterium constituting 48% of the total population and 12% clones having homology to Klebsiella. Aromatic amines generated during partial treatment under anoxic bioreactor were treated by aerobic population having 72% unculturable unidentified bacterium and rest of the population consisting of Thauera sp., Pseudoxanthomonassp., Desulfomicrobium sp., Ottowia sp., Acidovorax sp., and Bacteriodetes bacteriumsp.
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Transformation of azo dyes during wet heat sterilization-A source of error in typical microbial decolorization experiments(International Journal of Environmental Pollution) (Nishant Dafale, Satish Wate, Sudhir Meshram & Nageshwara Rao
,431-3,,264-273,Year : 2010)
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Dye degradation has gained attention of late due to indiscriminate disposal from user industries. Enhancing efficiency of biological treatment provides a cheaper alternative vis-à-vis other advanced technologies. Dye molecules are metabolized biologically via anoxic and oxic treatments. In this study, bacterial community surviving on dye effluent working in anoxic–oxic bioreactor was analyzed using 16S rDNA approach. Azo-dye decolorizing and degrading bacterial community was enriched in lab-scale two-stage anoxic–oxic bioreactor. 16S rDNA metagenomic libraries of enriched population were constructed, screened and phylogenetically analyzed separately. Removal of ∼35% COD with complete decolorization was observed in anoxic bioreactor. Process was carried out by uncultured gamma proteobacterium constituting 48% of the total population and 12% clones having homology to Klebsiella. Aromatic amines generated during partial treatment under anoxic bioreactor were treated by aerobic population having 72% unculturable unidentified bacterium and rest of the population consisting of Thauera sp., Pseudoxanthomonassp., Desulfomicrobium sp., Ottowia sp., Acidovorax sp., and Bacteriodetes bacteriumsp. This paper presents the role of moist heat sterilisation in decolourisation of azo dyes during autoclaving itself, thereby leading to over estimation of actual decolourisation through subsequent microbial process. This surprise phenomenon was probed in detail by studying the effects of temperature, pressure and role of electron donor/carbon sources on decolourisation. In the presence of 10 g/litre glucose, 75% decolourisation of Reactive Black 5 (RB-5) dye was observed after autoclaving of medium at 121°C for 15 min at 15 psi. Studies repeated with other azo dyes revealed that Reactive Orange 16 (RO-16) was affected by autoclaving whereas Reactive Red 11 (RR-11) and Reactive Red 141 (RR-141) did not show significant decolourisation. The reduction of dye was dependent on concentration of electron donor/carbon source and autoclave conditions. The results indicate that investigators must screen the dyes for decolourisation during autoclaving and choose the appropriate means of sterilisation to remove the artifice or incorporate correction factor for dye concentration at the start of experiment.
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Bioremediation of wastewater containing azo dyes using sequential anaerobic-aerobic bioreactor system and its biodiversity(Environmental Reviews) (Nishant Dafale, Satish Wate, Sudhir Meshram & Nageshwara Rao
,18,,21-36,Year : 2010)
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Wide range of dyes and dyestuffs used in textile manufacturing are xenobiotic compounds and attract stricter to strict environmental regulations. The ability of microbial consortia to decolorize and metabolize dyes has long been known, and the use of bioremediation based technologies for treating textile wastewater has attracted interest. These dyes are decolorized by microbial consortia but technologies for their complete mineralization are still not developed. The most logical concept for the removal of azo dyes in biological wastewater treatment systems is based on anaerobic treatment, for the reductive decolorization, in combination with aerobic treatment, for the degradation of the by-products (aromatic amines) generated in the anaerobic bioreactor. Several research and review articles were published on anaerobic decolorization; however, research on complete mineralization of dyes through sequential anaerobic–aerobic bioreactors has received greater attention recently. Bioremediation through sequential anaerobic–aerobic bioreactor system has been reviewed in this article with critical appraisal using data generated through our experiments. While reviewing this work, we realized the importance of microbial diversity in a treatment unit to better understand the functional status to enhance the mineralization activity of the bioreactor.
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Concentration and recovery of coliphages from water with bituminous coal(Water Environmental Research) (Nishant Dafale, Sukumar Lakhe, K. Yadav, Hemant Purohit & Tapan Chakraborty
,80 3,,282-288,Year : 2008)
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Coliphages represent a process indicator for fecal pollution. The coal bed concentration method prepared for enterovirus was refined for a concentration of coliphages. A bed made from 1.5 g of 120-mesh coal powder was used for concentrating coliphage from 200 mL of a water sample with or without the addition of aluminum chloride at different pH values. The isolated E. coli strain EC-R8 was found to be more susceptible to the desired coliphage and showed significant coliphage-coliform response, with clear plaque used for further studies. The complete coliphage adsorption was achieved with the addition of 0.0005 M AICI3 at pH 6.0. Adsorbed coliphages were eluted with 3% beef extract in Mcllvaine buffer at pH 7.1, with an average recovery of 78.74%. This concentration technique was applied for the detection of coliphages from the well water of Nagpur city (India) and found to contain coliphages in the range of 2 to 28 plaque-forming units per liter (PFU/L). Water Environ. Res.,
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Decolorization of azo dyes and simulated dye bath wastewater using acclimatized microbial consortium?biostimulation and halotolerance(Bioresource Technology) (Nishant Dafale, Nageshwara Rao, Sudhir Meshram & Satish Wate
,99,,2552-2558,Year : 2008)
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Anaerobic acclimatization of activated sludge from a textile effluent treatment plant to high concentration of RB5 could effectively decolorize RB5 dye solution. The strains viz. Pseudomonas aeruginosa and Bacillus circulans and other unidentified laboratory isolates (NAD1 and NAD6) were predominantly present in the microbial consortium. The conditions for efficient decolorization, biostimulation to increase effectiveness of microbial consortium, its tolerance to high salt concentration and non-specific ability towards decolorization of eight azo dyes, are reported. The optimum inoculums concentration for maximum decolorization were found to be 1–5 ml of 1800 ± 50 mg l1 MLSS and 37 C, respectively. The decolorization efficiency was 70–90% during 48 h. The biomass showed efficient decolorization even in the presence of 10% NaCl, as tested with RB5. In the presence of flavin adenine dinucleotide (FAD) more than 99% decolorization occurred in 8 h. The decolorization of RB5 was traced to extracellular enzymes. The effectiveness of acclimatized biomass under optimized conditions towards decolorization of two types of synthetic dye bath wastewaters that were prepared using chosen azo dyes is reported.
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Biodegradation enhancement of purified terephthalic acid wastewater by coagulation-flocculation process as pretreatment(Journal of Hazardous Material ) (M. Karthik, Nishant Dafale, Praduamya Pathe & Tapas Nandy
,154 1-3,,721-730,Year : 2008)
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In this work, the coagulation–flocculation process was used as pretreatment for purified terephthalic acid (PTA) wastewater with the objective of improving its overall biodegradability. PTA production generates wastewaters with toxicants p-xylene [1,4-dimethyl-benzene (C8H10)], a major raw material used in the production process, along with some of the intermediates, viz., p-toluic acid, benzoic acid, 4-carboxybenzaldehyde, phthalic acid and terephthalic acid. These compounds affect the bio-oxidation process of wastewater treatment; hence removal of these constituents is necessary, prior to conventional aerobic treatment. This paper addresses the application of coagulation–flocculation process using chemical coagulants, viz., aluminium sulphate (alum), polyaluminium chloride (PAC), ferrous sulphate and ferric chloride in combination with anionic polyelectrolyte. Polyaluminium chloride (PAC) in conjunction with lime and polyelectrolyte removed about 63.1% chemical oxygen demand (COD) and 45.2% biochemical oxygen demand (BOD) from PTA wastewater. Coagulation–flocculation process coupled with aerobic bio-oxidation treatment of PTA wastewater achieved, COD & BOD removals of 97.4% and 99.4%, respectively. The biodegradability enhancement evaluated in terms of the BOD5/COD ratio, increased from 0.45 to 0.67 at the optimum conditions. The results obtained from these studies indicate that the coagulation–flocculation process could be a suitable pretreatment method in reducing toxicity of PTA wastewater whilst enhancing biodegradability for aerobic biological treatment scheme.
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Kinetic study approach of remazol black-B use for the development of two-stage anoxic-oxic reactor for decolorization/biodegradation of azo dyes by activated bacterial consortium(Journal of Hazardous Material ) (Nishant Dafale, Satish Wate, Sudhir Meshram & Tapas Nandy
,159 ,,319-328,Year : 2008)
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The laboratory-isolated strains Pseudomonas aeruginosa, Rhodobacter sphaeroides, Proteus mirabilis, Bacillus circulance, NAD 1 and NAD 6 were observed to be predominant in the bacterial consortium responsible for effective decolorization of the azo dyes. The kinetic characteristics of azo dye decolorization by bacterial consortium were determined quantitatively using reactive vinyl sulfonated diazo dye, remazol black-B (RB-B) as a model substrate. Effects of substrate (RB-B) concentration as well as different substrates (azo dyes), environmental parameters (temperature and pH), glucose and other electron donor/co-substrate on the rate of decolorization were investigated to reveal the key factor that determines the performance of dye decolorization. The activation energy (Ea) and frequency factor (K0) based on the Arrhenius equation was calculated as 11.67 kcal mol−1 and 1.57 × 107 mg l g MLSS−1 h−1, respectively. The Double-reciprocal or Lineweaver–Burk plot was used to evaluate Vmax, 15.97 h−1 and Km, 85.66 mg l−1. The two-stage anoxic–oxic reactor system has proved to be successful in achieving significant decolorization and degradation of azo dyes by specific developed bacterial consortium with a removal of 84% color and 80% COD for real textile effluents vis-a-vis ` ≥90% color and COD removal for synthetic dye solution
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Microbial decolorization of reactive black-5 in a two stage anaerobic-aerobic reactor using acclimatized activated textile sludge (Sagrika Mohanty, Nishant Dafale & Nageshwara Rao
,17,,403-413,Year : 2006)
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A two-stage anaerobic–aerobic treatment process based on mixed culture of bacteria isolated from textile dye effluent was used to degrade reactive black 5 dye (RB-5). The anaerobic step was studied in more detail by varying the dye concentration from 100 to 3000 mg l)1 . The results showed that major decolorization was achieved during the anaerobic process. The time required for decolorization by >90% increased as the concentration of the dye increased. It was also found that maintaining dissolved oxygen (DO) concentration below 0.5 mg l)1 and addition of a co-substrate viz., glucose, facilitates anaerobic decolorization reaction remarkably. An attempt was made to identify the metabolites formed in anaerobic process by using high performance liquid chromatography (HPLC) and UV–VIS spectrophotometry. A plate assay was performed for the detection of dominant decolorizing bacteria. Only a few bacterial colonies with high clearing zones (decolorization zones) were found. The results showed that under anaerobic condition RB-5 molecules were reduced and aromatic amines were generated. The aromatic amine metabolite was partly removed in subsequent aerobic bio-treatment. It was possible to achieve more than 90% decolorization and approximately 46% reduction in amine metabolite concentration through two-stage anaerobic–aerobic treatment after a reaction period of 2 days.
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