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International Journal of Environmental... Dec 2017The key to obtaining an optimum performance of an enzyme is often a question of devising a suitable enzyme and optimisation of conditions for its immobilization. In this...
Preparation and Optimisation of Cross-Linked Enzyme Aggregates Using Native Isolate White Rot Fungi Trametes versicolor and Fomes fomentarius for the Decolourisation of Synthetic Dyes.
The key to obtaining an optimum performance of an enzyme is often a question of devising a suitable enzyme and optimisation of conditions for its immobilization. In this study, laccases from the native isolates of white rot fungi and/or , obtained from Czech forests, were used. From these, cross-linked enzyme aggregates (CLEA) were prepared and characterised when the experimental conditions were optimized. Based on the optimization steps, saturated ammonium sulphate solution (75 wt.%) was used as the precipitating agent, and different concentrations of glutaraldehyde as a cross-linking agent were investigated. CLEA aggregates formed under the optimal conditions showed higher catalytic efficiency and stabilities (thermal, pH, and storage, against denaturation) as well as high reusability compared to free laccase for both fungal strains. The best concentration of glutaraldehyde seemed to be 50 mM and higher efficiency of cross-linking was observed at a low temperature 4 °C. An insignificant increase in optimum pH for CLEA laccases with respect to free laccases for both fungi was observed. The results show that the optimum temperature for both free laccase and CLEA laccase was 35 °C for and 30 °C for . The CLEAs retained 80% of their initial activity for and 74% for after 70 days of cultivation. Prepared cross-linked enzyme aggregates were also investigated for their decolourisation activity on malachite green, bromothymol blue, and methyl red dyes. Immobilised CLEA laccase from showed 95% decolourisation potential and CLEA from demonstrated 90% decolourisation efficiency within 10 h for all dyes used. These results suggest that these CLEAs have promising potential in dye decolourisation.
Topics: Ammonium Sulfate; Azo Compounds; Bromthymol Blue; Catalysis; Color; Coloring Agents; Cross-Linking Reagents; Enzymes, Immobilized; Glutaral; Laccase; Polyporales; Rosaniline Dyes; Temperature; Trametes
PubMed: 29295505
DOI: 10.3390/ijerph15010023 -
Food Chemistry Jan 2023Intelligent packaging that provides real-time information on food quality is useful for consumers. We developed a kimchi ripening indicator that can determine the...
Intelligent packaging that provides real-time information on food quality is useful for consumers. We developed a kimchi ripening indicator that can determine the ripeness of kimchi inside packaging and evaluated its applicability and limitations. The indicator was made using calcium hydroxide, which captures CO, and four pH-sensitive dyes (cresol red, bromothymol blue, bromocresol purple, and methyl red). Fourier-transform infrared spectra of the prepared powders showed shapes similar to that of calcium hydroxide, and the dyes were evenly distributed on the calcium hydroxide surfaces. When the developed indicators were evaluated for kimchi packaging application, the indicator made from synthesized calcium hydroxide and bromothymol blue was the most reliable and clearly reflected useful kimchi ripening information. The indicator developed in this study is judged to be practically usable at temperatures of 4-15 °C. However, its usefulness is limited in that the seller cannot change the packaging capacity or kimchi capacity.
Topics: Bromcresol Purple; Bromthymol Blue; Calcium Hydroxide; Carbon Dioxide; Coloring Agents; Fermentation; Fermented Foods; Food Packaging; Hydrogen-Ion Concentration
PubMed: 36055147
DOI: 10.1016/j.foodchem.2022.134039 -
A Convenient, Rapid, Sensitive, and Reliable Spectrophotometric Assay for Adenylate Kinase Activity.Molecules (Basel, Switzerland) Feb 2019Enzymatic activity assays are essential and critical for the study of enzyme kinetics. Adenylate kinase (Adk) plays a fundamental role in cellular energy and nucleotide...
Enzymatic activity assays are essential and critical for the study of enzyme kinetics. Adenylate kinase (Adk) plays a fundamental role in cellular energy and nucleotide homeostasis. To date, assays based on different principles have been used for the determination of Adk activity. Here, we show a spectrophotometric analysis technique to determine Adk activity with bromothymol blue as a pH indicator. We analyzed the effects of substrates and the pH indicator on the assay using orthogonal design and then established the most optimal assay for Adk activity. Subsequently, we evaluated the thermostability of Adk and the inhibitory effect of KCl on Adk activity with this assay. Our results show that this assay is simple, rapid, and precise. It shows great potential as an alternative to the conventional Adk activity assay. Our results also suggest that orthogonal design is an effective approach, which is very suitable for the optimization of complex enzyme reaction conditions.
Topics: Adenylate Kinase; Bromthymol Blue; Homeostasis; Hydrogen-Ion Concentration; Kinetics; Phosphorylation; Potassium Chloride; Spectrophotometry
PubMed: 30781833
DOI: 10.3390/molecules24040663 -
International Journal of Biological... May 2023Continuous wound monitoring is one strategy to minimise infection severity and inform prompt variations in therapeutic care following infection diagnosis. However,...
Continuous wound monitoring is one strategy to minimise infection severity and inform prompt variations in therapeutic care following infection diagnosis. However, integration of this functionality in therapeutic wound dressings is still challenging. We hypothesised that a theranostic dressing could be realised by integrating a collagen-based wound contact layer with previously demonstrated wound healing capability, and a halochromic dye, i.e. bromothymol blue (BTB), undergoing colour change following infection-associated pH changes (pH: 5-6 ➔ >7). Two different BTB integration strategies, i.e. electrospinning and drop-casting, were pursued to introduce long-lasting visual infection detection capability through retention of BTB within the dressing. Both systems had an average BTB loading efficiency of 99 wt% and displayed a colour change within 1 min of contact with simulated wound fluid. Drop-cast samples retained up to 85 wt% of BTB after 96 h in a near-infected wound environment, in contrast to the fibre-bearing prototypes, which released over 80 wt% of BTB over the same time period. An increase in collagen denaturation temperature (DSC) and red shifts (ATR-FTIR) suggest the formation of secondary interactions between the collagen-based hydrogel and the BTB, which are attributed to count for the long-lasting dye confinement and durable dressing colour change. Given the high L929 fibroblast viability in drop-cast sample extracts (92 %, 7 days), the presented multiscale design is simple, cell- and regulatory-friendly, and compliant with industrial scale-up. This design, therefore, offers a new platform for the development of theranostic dressings enabling accelerated wound healing and prompt infection diagnosis.
Topics: Humans; Precision Medicine; Bandages; Collagen; Wound Healing; Surgical Wound Infection; Bromthymol Blue; Hydrogels
PubMed: 36870632
DOI: 10.1016/j.ijbiomac.2023.123866 -
Journal of Agricultural and Food... Nov 2020This work presents a colorimetric dye-based array for naked-eye detection of chicken meat spoilage. The array is obtained by fixing five acid-base indicators, -cresol...
This work presents a colorimetric dye-based array for naked-eye detection of chicken meat spoilage. The array is obtained by fixing five acid-base indicators, -cresol purple (), -cresol red (), bromothymol blue (), thymol blue (), and chlorophenol red (), and a sensing molecule specific for thiols, 5,5'-dithiobis(2-nitrodibenzoic acid), called Ellman's reagent (), on a cellulose-based support. The dyes, being permanently charged, are fixed on the support via ion-exchange. The entire degradation process of beast poultry meat, at ambient temperature and in a domestic fridge, is followed by the change of the color of the array, placed in the headspace over the meat samples. The device is set after selection of the most suitable starting form, which could be the acidic or the basic color of indicators, being the proper dye concentration and the dimension of the spots already established. Basing on sensors colors, we identified three levels of the degradation process of chicken meat, named SAFE, WARNING, and HAZARD. By instrumental analysis, we demonstrated that sensors response was correlated to volatile organic compounds (VOCs) composition in the headspace and, thus, to meat spoilage progress. We demonstrated that biogenic amines (BAs), commonly considered a critical spoilage marker, are indeed produced into the samples but never present in the headspace, even in traces, during the investigated time-lapse. The VOC evolution nevertheless allows one to assign the sample as WARNING and further HAZARD. Some indicators turned out to be more informative than others, and the best candidates for a future industrial application resulted in a bromothymol blue ()-, chlorophenol red ()-, and Ellman's reagent ()-based array.
Topics: Animals; Bromthymol Blue; Chickens; Color; Colorimetry; Coloring Agents; Food Analysis; Food Safety; Meat; Phenolsulfonphthalein; Thymolphthalein; Volatile Organic Compounds
PubMed: 33118801
DOI: 10.1021/acs.jafc.0c03771 -
Journal of Food Protection Mar 2017A modified polymyxin-egg yolk-mannitol-bromothymol blue agar (mPEMBA) was developed by supplementing polymyxin-egg yolk-mannitol-bromothymol blue agar (PEMBA) with...
A modified polymyxin-egg yolk-mannitol-bromothymol blue agar (mPEMBA) was developed by supplementing polymyxin-egg yolk-mannitol-bromothymol blue agar (PEMBA) with trimethoprim to improve the selectivity for and recoverability of Bacillus cereus from naturally and artificially contaminated food samples. The number of B. cereus in mPEMBA was significantly higher than in PEMBA, indicating better recoverability (P < 0.05) in red pepper powder (PEMBA 0.80 ± 0.22 log CFU/g versus mPEMBA 1.95 ± 0.17 log CFU/g) and soybean paste (PEMBA 2.19 ± 0.18 log CFU/g versus mPEMBA 3.09 ± 0.13 log CFU/g). In addition, mPEMBA provided better visual differentiation of B. cereus colonies than PEMBA, which is attributable to the reduced number of competing microflora. We conclude that the addition of trimethoprim to PEMBA could generate a synergistic effect to improve selectivity for B. cereus .
Topics: Agar; Bacillus cereus; Bromthymol Blue; Culture Media; Egg Yolk; Food Microbiology; Mannitol; Polymyxins
PubMed: 28207304
DOI: 10.4315/0362-028X.JFP-16-206 -
Acta Poloniae Pharmaceutica 2015Two simple, rapid and sensitive spectrophotometric methods have been developed for the determination of naproxen in pure, pharmaceutical preparation and human serum...
Two simple, rapid and sensitive spectrophotometric methods have been developed for the determination of naproxen in pure, pharmaceutical preparation and human serum samples. These methods are based on the formation of yellow ion-pair complexes between naproxen and two sulfophthalein acid dyes, namely bromocresol green (BCG method) and bromothymol blue (BTB method). The resulting complexes were measured at 424 nm (BCG method) and at 422 nm (BTB method). The effects of variables such as reagent concentration and reaction time were investigated to optimize the procedure. Beer's law was obeyed in the concentration range of 10-105 µg/mL and 5-85 µg/mL and the detection limits were found to be 0.347 and 0.31 µg/mL for BCG and BTB methods, respectively. The developed methods have been successfully applied for the determination of naproxen in bulk drugs, pharmaceutical formulations and human serum samples with good accuracy and precision. The results are comparable to those of reference methods, and hence are recommended for quality control and routine analysis.
Topics: Bromcresol Green; Bromthymol Blue; Humans; Hydrogen-Ion Concentration; Naproxen; Spectrophotometry; Tablets
PubMed: 26665392
DOI: No ID Found -
PloS One 2015Over the last few decades the establishment of nanoparticles as suitable drug carriers with the transport of drugs across biological barriers such as the...
Over the last few decades the establishment of nanoparticles as suitable drug carriers with the transport of drugs across biological barriers such as the gastrointestinal barrier moved into the focus of many research groups. Besides drug transport such carrier systems are well suited for the protection of drugs against enzymatic and chemical degradation. The preparation of biocompatible and biodegradable nanoparticles based on poly(lactic-co-glycolic acid) (PLGA) is intensively described in literature, while especially nanoparticles with cationic properties show a promising increased cellular uptake. This is due to the electrostatic interaction between the cationic surface and the negatively charged lipid membrane of the cells. Even though several studies achieved the successful preparation of nanoparticles stabilized with the cationic surfactants such as didodecyldimethylammonium bromide (DMAB), in most cases insufficient attention was paid to a precise analytical characterization of the nanoparticle system. The aim of the present work was to overcome this deficit by presenting a new perspective in the formulation and characterization of DMAB-stabilized PLGA nanoparticles. Therefore these nanoparticles were carefully examined with regard to particle diameter, zeta potential, the effect of variation in stabilizer concentration, residual DMAB content, and electrolyte stability. Without any steric stabilization, the DMAB-modified nanoparticles were sensitive to typical electrolyte concentrations of biological environments due to compression of the electrical double layer in conjunction with a decrease in zeta potential. To handle this problem, the present study proposed two modifications to enable electrolyte stability. Both polyvinyl alcohol (PVA) and polyethylene glycol (PEG) modified DMAB-PLGA-nanoparticles were stable during electrolyte addition. Furthermore, in contrast to unmodified DMAB-PLGA-nanoparticles and free DMAB, such modifications led to a lower cytotoxic activity against Caco-2 cells. In conclusion this study offers a closer and critical point of view on preparation, in vitro and analytical evaluation of DMAB-stabilized PLGA nanoparticles for the physiological use.
Topics: Bromthymol Blue; Caco-2 Cells; Colloids; Electrolytes; Humans; Lactic Acid; Microscopy, Fluorescence; Nanoparticles; Polyglycolic Acid; Polylactic Acid-Polyglycolic Acid Copolymer; Quaternary Ammonium Compounds
PubMed: 26147338
DOI: 10.1371/journal.pone.0127532 -
Annals of Laboratory Medicine Sep 2016Rapid detection of carbapenemase-producing gram-negative bacilli (GNB) is required for optimal treatment of infected patients. We developed and assessed a new disk...
BACKGROUND
Rapid detection of carbapenemase-producing gram-negative bacilli (GNB) is required for optimal treatment of infected patients. We developed and assessed a new disk carbapenemase test (DCT).
METHODS
Paper disks containing 0.3 mg of imipenem and bromothymol blue indicator were developed, and the performance of the DCT were evaluated by using 742 strains of GNB with or without carbapenemases.
RESULTS
The paper disks were simple to prepare, and the dried disks were stable at -20°C and at 4°C. The DCT detected 212 of 215 strains (98.6% sensitivity with 95% confidence interval [CI] 96.0-99.5%) of GNB with known class A (KPC and Sme) and class B (NDM, IMP, VIM, and SIM) carbapenemases within 60 min, but failed to detect GES-5 carbapenemase. The DCT also detected all two Escherichia coli isolates with OXA-48, but failed to detect GNB with OXA-232, and other OXA carbapenemases. The DCT showed 100% specificity (95% CI, 99.2-100%) in the test of 448 imipenem-nonsusceptible, but carbapenemase genes not tested, clinical isolates of GNB.
CONCLUSIONS
The DCT is simple and can be easily performed, even in small laboratories, for the rapid detection of GNB with KPC, NDM and the majority of IMP, VIM, and SIM carbapenemases.
Topics: Anti-Bacterial Agents; Bacterial Proteins; Bromthymol Blue; Drug Resistance, Bacterial; Gram-Negative Bacteria; Imipenem; Microbial Sensitivity Tests; Paper; beta-Lactamases
PubMed: 27374708
DOI: 10.3343/alm.2016.36.5.434 -
Revista Do Instituto de Medicina... Sep 2016Cryptococcosis is a leading invasive fungal infection in immunocompromised patients. Considering the high prevalence and severity of these infections in...
ANTIFUNGAL SUSCEPTIBILITY TESTING AND GENOTYPING CHARACTERIZATION OF Cryptococcus neoformans AND gattii ISOLATES FROM HIV-INFECTED PATIENTS OF RIBEIRÃO PRETO, SÃO PAULO, BRAZIL.
Cryptococcosis is a leading invasive fungal infection in immunocompromised patients. Considering the high prevalence and severity of these infections in immunocompromised patients attended at HC-FMRP-USP, the present research aimed to characterize the clinical isolates of Cryptococcus strains by biochemical and molecular methods and evaluate antifungal susceptibility of clinical isolates. Fifty isolates from 32 HIV-positive patients were obtained at HC-FMRP-USP. Most of the isolates (78.1%) were identified as C. neoformans, and 100% of C. neoformans and C. gattii strains were susceptible to amphotericin B, ketoconazole and fluconazole. All isolates were classified as serotype A (grubbii variety) by PCR and most of them were characterized in mating type MATa. PCR analysis of specific M13 microsatellite sequence revealed that VNI type was predominant among C. neoformans, while VGII was predominant among C. gattii. The strains did not show a significant resistance to the antifungals tested, and Canavanine-Glycine-Bromthymol Blue Agar (CGB) proved to be a reliable test presenting a good correlation with the molecular characterization. C. neoformans isolated from disseminated infections in the same patient showed molecular identity when different anatomical sites were compared; besides, the studied strains did not present a significant increase in resistance to antifungal agents. In addition, the homogeneity of the molecular types and detection of the mating types suggested a low possibility of crossing among the strains.
PubMed: 27680174
DOI: 10.1590/S1678-9946201658069