Color Change Of Indicator Research Articles

Visual and simple determination of glucose-induced acidification by yeast cells: application to rapid cytotoxicity test

This study demonstrated that glucose-induced proton release from yeast cells was more sensitive to various inhibitors than cell proliferation. In this study the inhibition of glucose-induced proton release was determined on the basis of color change of pH indicator, methyl red, from pH 5 to pH6 at cell density of 2.5 × 107 cells/ml. When yeast cells were incubated with the inhibitors of glucose intake, glycolysis, and plasma membrane H + -ATPase for 1 h, these cytotoxic effects were observed by following the change in absorbance at 527 nm due to methyl red for 5 min. The cytotoxic effects of heavy metal ions, detergents and quinones were observed in the same manner. The above method was superior in sensitivity and measurement time to cell proliferation measurement that required 9 h. This visual cytotoxicity test (methyl red test) is expected to be useful as simple and rapid cytotoxicity test with yeast cells.

Micro-PAD card for measuring total ammonia nitrogen in saliva

This work presents a portable microfluidic paper-based analytical device (micro-PAD) card for the quantification of total ammonia nitrogen in human saliva. The amount of total ammonia nitrogen in saliva can be an indicator of the status of the oral microbiome with potential correlation to kidney health problems. The developed micro-PAD card comprises twenty units consisting of three stacked layers of circular discs: the sample layer, paper discs impregnated with sodium hydroxide solution, the PTFE membrane layer, and the detection layer, paper discs impregnated with bromothymol blue. The twenty units were aligned on transparent laminating pouches laminated to form the micro-PAD card (7.5 cm × 10.5 cm). Saliva samples can be directly dispensed onto the micro-PAD card and the detection was achieved by the BTB indicator color change, from yellow to blue, after conversion of ammonium into ammonia and diffusion of the ammonia gas through a hydrophobic layer. The determination of total ammonia nitrogen in saliva using the developed micro-PAD card intended to be very simple method and operated without the need of laboratory equipment. A quantification limit of 11.3 NH4+mg L-1 and linear application range from up to 150 NH4+mg L-1 were obtained making it suitable for the expected concentrations of total ammonia nitrogen in human saliva. It was successfully applied to saliva samples and its validation obtained by comparison against a potentiometric method. The card is stable for at least 1 month making it ideal as a portable device for point-of-care diagnosis. Graphical Abstract.

The Preparation and Characterization of Alginate–Chitosan Membranes as Solid Support for BTB and Urease Entrapment

A study of alginate-chitosan membrane synthesize was done. The membrane was prepared by mixing alginate hydrosol and chitosan hydrosol at mass ratio of 1:1 and pH of 5.28 approximately. Then it was applied for matrix immobilization of urease and bromothymol blue (BTB) by entrapment technique. The physical, chemical, thermal properties of alginate–chitosan membrane and their impact on immobilized urease activity were investigated. The polymer products were characterized by Fourier Transform Infrared Spectroscopy (FTIR), X-Ray Diffraction (XRD), Thermogravimetric Analysis (TGA), Differential Scanning Calorimetry (DSC), and Scanning Electron Microscopy (SEM). It’s showed that alginate-chitosan membrane was formed by electrostatic interaction. The obtained membran has better mechanical properties than original alginat and chitosan membranes. The immobilization urease into alginate-chitosan membrane retained the catalytic activity of the enzyme, as confirmed by color change of BTB indicator after membrane was immersed in substrate solution (urea). Therefore, alginate-chitosan membrane has good characteristics as matrix of enzyme immobilization.

Accelerated aging by microwave heating and methods to distinguish aging of rice

AbstractAimTo study the impact of microwave heating (950 and 1,400 W for 72 s) in accelerating the aging process in selected rice varieties, namely IR‐50, Sona masuri, and RNR‐15048 (Rajendranagar variety) stored as brown and polished form.MethodsThe impact was investigated by comparing the efficiencies of two existing chemical methods, namely bromothymol blue indicator (Method‐1) based on the indicator color change with respect to changes in rice pH followed by peroxidase enzyme color reaction (Method‐2) using test tube (spectrophotometer at 470 nm) and single‐grain tests (96‐well plate microplate reader at 386 nm) paralleled with in vitro enzyme assessment for fresh‐aged indices calculation during 6 months of storage.ResultsThe results revealed the maximum R2‐value of 0.993 (brown rice) and 0.963 (polished rice) at 1,400 W for 72 s in RNR‐15048 in single grain test (Method‐1) followed by a maximum R2‐value of 0.976 in brown rice of Sona masuri at 950 W for 72 s and 0.941 in polished rice of RNR‐15048 at 1,400 W for 72 s in single‐grain test (Method‐2).ConclusionAmong the above methods, Method‐1 was found to be more profound in obtaining the consistent results and can be highly recommended for both stake holders and rice processing agencies due to low cost, ease in chemical preparation, and color changes can be easily distinguishable during storage.Practical ApplicationsAging is one of the important and prudent steps in rice grain. Aged rice fetches more price in the market than fresh rice due to improved aroma, texture, and taste after cooking. Accelerated aging requires less storage space and time compared to natural aging. Identification of the aging process in rice grains is a challenging task. Bromothymol blue indicator based on pH change in rice grains due to increase in free fatty acids and peroxidase enzyme color change due to peroxidase enzyme (EC.1.11.1.7) depletion during storage helps to identify the rice aging through fresh‐aged index calculation. Among both the methods, Bromothymol blue indicator is one of the cost‐effective methods for quick identification of rice aging based on indicator color change. This method helps the farmers and other rice processing sectors to distinguish the aged rice from fresh grains as it is cheap, ease in chemical preparation, and affordable.

Application Nano Zeolite-Molybdate For Avocado Ripeness Indicator

Zeolite is aluminosilicate containing pores and cavities that are usually used as absorbent materials. Nano zeolite - molybdate was used as a ripeness indicator of avocado. The ripeness indicator was made by dissolving molybdate in hydrogen peroxide (H2O2) and mixed with nano zeolite. This study was aimed to determine the colour change of indicator, and the relationship between the changing colour of indicator and the ripening of avocado. The ripeness indicator was prepared used activated nano zeolite with HCl 1 N for 24 h, then mixed with 4 mL peroxomolybdate. Nano zeolite-molybdate tablets was packed using 0,04 mm thick of LDPE plastic, and it was inserted inside avocado packaging. The ripeness indicator would identified by the ethylene production of avocados, and resulted changing the indicator colour from yellow to blue. The yellow colour would represent unripe avocado, but the blue colour would represent ripe avocados. The colour changing was related with the decrease of avocado quality during 10 days storage with the rate of weight loss 0.0113%, the rate of sugar level 0.7182% and the rate of avocado hardness 2.1132% by day.

Expression of an Environmentally Friendly Enzyme, Engineered Carbonic Anhydrase, in Escherichia coli

Carbon dioxide (CO2) is one of the main causes for global warming. The most important enzyme that converts CO2 into bicarbonate and prevents CO2 emissions in the environment is carbonic anhydrase (CA). The aim of this study was to clone the heat-stable CA gene in Escherichia coli host. The CA gene coding frame of Caminibacter mediatlanticus was optimized based on the more frequently used codons by E. coli. Accordingly, nine codons were inserted in this gene fragment. After CA gene insertion in cloning vector (pGH), it was sub-cloned in pBI121 expression vector and transformed into E. coli XL1-Blue. The accuracy of gene cloning was confirmed by PCR, restriction enzyme digestion and sequencing. Cell extract was prepared from recombinant bacteria. Culture supernatants were assayed for CA activity by the color change of bromothymol blue as indicator. The time for the indicator color changes were recorded and the assays were repeated five times. The average of color change time for cell extract of recombinant, non-recombinant E. coli and control reaction were 1.5, 16, and 48 min, respectively. The amount of active enzyme was calculated to be 92.8 units/ml and the enzyme retained its activity after of incubation at 60 °C, 65 °C, and 70 °C. Due to the positive properties of CA from C. mediatlanticus, codon optimization, modification, and expression of this gene, using a simple and inexpensive method, may be used to obtain CA with enhanced properties.

Синтез и исследование противомикробной активности производных нифуроксазида

В связи с постоянно растущим числом инфекций, вызываемых устойчивыми к синтетическим антибактериальным препаратам и антибиотикам микроорганизмами, основные усилия исследователей всего мира направлены на решение данной проблемы. Одним из самых эффективных среди антибактериальных препаратов является класс замещенных нитрофуранов. Синтезированы 4 аналога нифуроксазида (представителя нитрофуранового ряда), структура которых подтверждена с помощью ЯМР, ИК спектроскопии и масс-спектрометрии. Все полученные соединения изучены на противомикробную и противогрибковую активность. Активность по отношению к бактериям Escherichia coli, Staphylococcus aureus, Staphylococcus haemolyticus и Pseudomonas aeruginosa оценивали методом диффузии в агар. Полученные вещества подавляли рост всех исследованных штаммов бактерий, кроме Escherichia coli; диаметр зон ингибирования находился в пределах от 13.5 до 28 мм в зависимости от концентрации исследуемого вещества и штамма бактерий. Одно из изученных в этой работе соединений – пиридиновый аналог нифуроксазида – превзошло по активности стандарт (нифуроксазид) по отношению к Staphylococcus aureus и поэтому может считаться перспективным соединением для разработки новых антибактериальных средств. Определение ингибирующей активности синтезированных препаратов по отношению к дрожжевым грибам Candida albicans и Cryptococcus neoformans проводили с помощью метода последовательных разведений с оценкой эффекта по изменению окраски индикатора. Ни одно из полученных соединений не проявило активности в отношении данных культур. Полученные результаты подтверждают, что замещенные аналоги нифуроксазида обладают противомикробной активностью и являются перспективными кандидатами для разработки новых антибактериальных средств.

Synthesis and study of the antimicrobial activity of nifuroxazide derivatives

ABSTRACT The number of infections caused by microorganisms that are resistant to antibiotics and synthetic antibacterial drugs is growing fast worldwide. This is one of the most important and urgent problems in health care. The main efforts of researchers around the world are focused on solving this issue. Nitrofurans represent one of the most effective classes of antibacterial drugs. We have synthesized 4 analogues of nifuroxazide – a well known nitrofuran antibiotic – and confirmed their structures by NMR, IR spectroscopy, and mass-spectrometry. All of the obtained compounds were studied for antimicrobial and antifungal activity. Activity against Escherichia coli, Staphylococcus aureus, Staphylococcus haemolyticus, and Pseudomonas aeruginosa was evaluated by the agar diffusion method. The synthesized compounds suppressed the growth of all the studied bacterial strains except Escherichia coli; the diameter of the inhibition zones ranged from 13.5 to 28 mm depending on the concentration of the tested compound and bacterial strain. One of the compounds studied in this project – the pyridine analogue of nifuroxazide – exceeded the activity of the standard (nifuroxazide) against the Staphylococcus aureus. The inhibitory activity of the synthesized compounds against the Candida albicans and Cryptococcus neoformans yeasts was determined using the microdilution method. The results were assessed according to the indicator color change. None of the studied compounds showed activity against these cultures. The obtained results confirm that substituted nifuroxazides have significant antimicrobial activity and, therefore, can be considered as promising candidates for developing new antibacterial drugs.

High-resolution optical pH imaging of concrete exposed to chemically corrosive environments

Major types of chemical concrete degradation such as carbonation, leaching and acid attacks are strongly associated with decreasing internal, surface and/or external pH. Thus, a precise pH determination is crucial for the assessment regarding the degree of corrosion and corresponding development of prevention strategies. Conventional pH measurement methods for concrete, such as pH visualization via color change of phenolphthalein indicator, pH analyses of extracted pore solutions and surface pH characterizations using flat surface electrodes have proven to hold significant limitations and inadequacies. This contribution presents the application of sensor foils based on luminescent, pH sensitive dyes for quantification and imaging of the spatial distribution of surface pH of concrete within a pH range between 9.35 and 12.35. For this purpose, an imaging technique called time-domain dual lifetime referencing (t-DLR) was used. High-resolution pH images of concrete samples exposed to carbonation and biogenic acid corrosion were successfully recorded and consequently compared to the state-of-the-art methods for pH assessment on concrete-based construction materials.

Design and fabrication of starch‐nano clay composite films loaded with methyl orange and bromocresol green for determination of spoilage in milk package

In this work, new composite film based on starch and nano clay particles was provided. The nano composite film was loaded by methyl orange (MO) and bromocresol green (BG) indicators. Two type films, including starch‐nano clay‐methyl orange (SCMO) and starch‐nano clay‐bromocresol green (SCBG) were provided. Some properties of provided films, including solubility in water, moisture uptake, water vapor permeability, FT‐IR, and scanning electron microscopy were studied. According to the results, MO and BG were loaded between starch and nano clay particles physically and clay particle size was between 40 and 100 nm. Finally, the SCMO and SCBG simultaneously were improvised in the milk bottle. According to the color change of indicators in the different pH (MO [3.1‐4.4] and BG [3.8‐5.4]), the fabricated films were used for determination of milk spoilage in a milk bottle. The best advantage of fabricated SCMO and SCBG films is that the indicators do not release into the milk by exposing with the milk. The milk pH is changed during the storage time (from 6.7 to the 4), and in the low pH, the milk is spoiled so the fabricated film smartly could show the spoilage of milk by color changing.

Feasibility of Using an Electrolysis Cell for Quantification of the Electrolytic Products of Water from Gravimetric Measurement

A gravimetric method for the quantitative assessment of the products of electrolysis of water is presented. In this approach, the electrolysis cell was directly powered by 9 V batteries. Prior to electrolysis, a known amount of potassium hydrogen phthalate (KHP) was added to the cathode compartment, and an excess amount of KHCO3 was added to the anode compartment electrolyte. During electrolysis, cathode and anode compartments produced OH−(aq) and H+(aq) ions, respectively. Electrolytically produced OH−(aq) neutralized the KHP, and the completion of this neutralization was detected by a visual indicator color change. Electrolytically produced H+(aq) reacted with HCO3 −(aq) liberating CO2(g) from the anode compartment. Concurrent liberation of H2(g) and O2(g) at the cathode and anode, respectively, resulted in a decrease in the mass of the cell. Mass of the electrolysis cell was monitored. Liberation of CO2(g) resulted in a pronounced effect of a decrease in mass. Experimentally determined decrease in mass (53.7 g/Faraday) agreed with that predicted from Faraday's laws of electrolysis (53.0 g/Faraday). The efficacy of the cell was tested to quantify the acid content in household vinegar samples. Accurate results were obtained for vinegar analysis with a precision better than 5% in most cases. The cell offers the advantages of coulometric method and additionally simplifies the circuitry by eliminating the use of a constant current power source or a coulometer.

吸収スペクトル測定による酸-塩基指示薬の変色域決定 : 気体を溶液に吹き込んで pH を変化させる方法

吸収スペクトル測定による酸-塩基指示薬の変色域決定 : 気体を溶液に吹き込んで pH を変化させる方法

Development of a robust system for high-throughput colorimetric assay of diverse amino acid decarboxylases

Abstract Amino acid decarboxylases catalyze decarboxylation of amino acids into amines that possess wide industrial applications. As key enzymes in biobased production of industrially important amines such as cadaverine, putrescine and β-alanine, lysine decarboxylase, ornithine decarboxylase and aspartic acid decarboxylase have attracted increasing attention. To develop enzyme variants with superior catalytic properties, there is a great need for high-throughput assay of these decarboxylases. Here we report the development of assays based on the color change of pH indicator – chlorophenol red (CPR) or bromothymol blue (BTB) – in decarboxylation reactions, in which one proton was consumed per carboxylic group decarboxylated resulting in an increase in pH. First, two buffer-indicator pairs, 4-morpholineethanesulfonic acid (MES)-CPR and 3-morpholinopropanesulfonic acid (MOPS)-BTB, were chosen on the basis of their similar pKa values at approximately pH 6.0 and 7.0, both of which are physiologically relevant. Next, the effects of buffer strength and indicator concentration on absorbance changes were examined in assay mixtures with NaOH titration, which mimicked proton consumption in decarboxylation reactions. Finally, high-throughput quantification of lysine decarboxylase, ornithine decarboxylase and aspartic acid decarboxylase was achieved using a microplate format. These results suggest that our indicator assay system may have potential applications for screening diverse decarboxylases.

MODEL KINETIKA PERUBAHAN WARNA LABEL INDIKATOR DARI KLOROFIL DAUN SINGKONG (MANIHOT ESCULENTA CRANTZ)

Time Temperature Indicators (TTI) is a smart packaging that important for consumers to know the appropriateness of product through an indicator colour change. Currently, there is no development of the kinetics models at TTI to give information for users about the relationship between colour change indicator and time storage. The objective of this research was to develop a kinetic model of smart label. This label was made from chitosan and PVA were added by chlorophyll extracted from cassava leaves. Indicator label stored at five storage temperatures to check the label performance at temperatures of freezer (-10±2°C), refrigerator (2±3oC), air conditioning (17±2oC), room (25±2oC), and hot temperature (50oC). Parameter was observed in the colour change at the label is the value of ohue which show the real colour.The results showed that label indicator changed colour from green to brown. The changes of colour indicator label wouldldarker when stored at high temperatures.The kinetic model of chlorophyll discolouration was developed by Arrhenius equation using ordo 0 and ordo 1. The plot of ln k value and 1/T has resulted on the equation for kinetic model discolouration label the values of k = 9.22.1010 e -15294,7/T with activation energy of 15.2947 kkal/mol for ordo 0 and k = 1.81.1012 e -19891,9/T with activation energy of 19.8919 kkal/mol for ordo 1. The model could predict the quality change of some product accordance with the kinetic model.Keywords : kinetic model, smart label, chlorophyll

Rapid Aminoglycoside NP Test for Rapid Detection of Multiple Aminoglycoside Resistance in Enterobacteriaceae.

The rapid aminoglycoside NP (Nordmann/Poirel) test was developed to rapidly identify multiple aminoglycoside (AG) resistance in Enterobacteriaceae It is based on the detection of the glucose metabolism related to enterobacterial growth in the presence of a defined concentration of amikacin plus gentamicin. Formation of acid metabolites was evidenced by a color change (orange to yellow) of the red phenol pH indicator. The rapid aminoglycoside NP test was evaluated by using bacterial colonies of 18 AG-resistant isolates producing 16S rRNA methylases, 20 AG-resistant isolates expressing AG-modifying enzymes (acetyl-, adenyl-, and phosphotransferases), and 10 isolates susceptible to AG. Its sensitivity and specificity were 100% and 97%, respectively, compared to the broth dilution method, which was taken as the gold standard for determining aminoglycoside resistance. The test is inexpensive, rapid (<2 h), and implementable worldwide.

Acetylcholinesterase Inhibitors Assay Using Colorimetric pH Sensitive Strips and Image Analysis by a Smartphone

Smartphones are widely spread and their usage does not require any trained personnel. Recently, smartphones were successfully used in analytical chemistry as a simple detection tool in some applications. This paper focuses on immobilization of acetylcholinesterase (AChE) onto commercially available pH strips with stabilization in the gelatin membrane. AChE degrades acetylcholine into choline and acetic acid which causes color change of acid-base indicator. Smartphone served as a tool for measurement of indicator color change from red to orange while inhibitors blocked this process. AChE inhibitors were measured with limits of detection, 149 nM and 22.3 nM for galanthamine and donepezil, respectively. Organic solvents were measured for method interferences. Measurement procedure was performed on 3D printed holder and digital photography was evaluated using red-green-blue (RGB) channels. The invented assay was validated to the standard Ellman's test and verified on murine plasma samples spiked with inhibitors. We consider that the assay is fully suitable for practical performance.

TEST SYSTEM BASED ON ROOT EXUDATES FOR HIGH-YIELDING COMMON BUCKWHEAT ( FAGOPYRUM ESCULENTUM MOENCH.) FORM SCREENING

A new effective non-invasive method of screening of highly productive forms of buckwheat sowing (Fagopyrum esculentum Moench.) based on rapid testing of buckwheat seedling intensity exudation of organic acids root system in the laboratory is offered. Buckwheat seeds were germinated on agar gel layer which contains in its composition acid-base indicator followed by visual assessment of the indicator color changes around primary root and plants with the largest area of color change were selected. The effectiveness of the method was confirmed in the field conditions by phenotyping of plants and significant differences in determining the structure and yield performance of selected plants were found. Statistical analysis of indicators grain number and grain weight showed that these indicators in selected plants were over 6 times higher than in the control variant with the degree of reliability of 99%.

Alginate cryogel based glucose biosensor

Cryogel is macroporous structure provides a large surface area for biomolecule immobilization. In this work, an alginate cryogel based biosensor was developed to detect glucose. The cryogel was prepared using alginate cross-linked by calcium chloride under sub-zero temperature. This porous structure was growth in a 100 μL micropipette tip with a glucose oxidase enzyme entrapped inside the cryogel. The glucose detection was based on the colour change of redox indicator, potassium permanganate, by the hydrogen peroxide resulted from the conversion of glucose. The result showed a porous structure of alginate cryogel with pores diameter of 20-50 μm. The developed glucose biosensor was showed a linear response in the glucose detection from 1.0 to 5.0 mM with a regression of y = 0.01x+0.02 and R2 of 0.994. Furthermore, the glucose biosensor was showed a high operational stability up to 10 times of uninterrupted glucose detections.

A novel assay for the detection of bioactive volatiles evaluated by screening of lichen-associated bacteria

Volatile organic compounds (VOCs) produced by microorganisms are known both for their effect on pathogens and their role as mediators in various interactions and communications. Previous studies have demonstrated the importance of VOCs for ecosystem functioning as well as their biotechnological potential, but screening for bioactive volatiles remained difficult. We have developed an efficient testing assay that is based on two multi-well plates, separated by a sealing silicone membrane, two tightening clamps, and variable growth media, or indicators. The experiment design as presented here is a novel and robust technique to identify positive as well as negative VOC effects on the growth of a target organism and to test for specific substances e.g., hydrogen cyanide which can be detected with a suitable indicator. While the first pre-screening assay is primarily based on indicator color change and visible growth diameter reduction, we also introduce an advanced and quantitatively precise experiment design. This adaptation involves qPCR-based quantification of viable target cells after concluding the treatment with VOCs. Therefore, we chose preselected active isolates and compared the partial 16S rRNA gene copy number of headspace-exposed E. coli with non-treated controls. Separately obtained headspace SPME and GC/MS-based profiles of selected bacterial isolates revealed the presence of specific and unique signatures which suggests divergent modes of action. The assay was evaluated by screening 100 isolates of lung lichen-associated bacteria. Approximately one quarter of the isolates showed VOC-based antibacterial and/or antifungal activity; mainly Pseudomonas and Stenotrophomonas species were identified as producers of bioactive volatiles.

Biodegradation of One Ring Hydrocarbons (Benzene and Toluene) and Two Ring Hydrocarbons (Acenapthene and Napthalene) by Bacterial Isolates of Hydrocarbon Contaminated Sites Located in Chhattisgarh: A Preliminary Study

Aromatic hydrocarbons are common groundwater and soil contaminants associated with petroleum product releases. They are hydrophobic and can readily bio-accumulate in the environment. Some PAHs (Polyaromatic Hydrocarbons) are harmful and known to be carcinogenic, mutagenic or genotoxic. Soil samples from hydrocarbon contaminated sites of Chhattisgarh were taken aseptically and their physicochemical properties studied. Soil samples were cultured in BBH media from which a total of eight positive isolates were studied for utilization of four aromatic hydrocarbons viz. Benzene, Toluene, Acenapthene and Napthalene. After biochemical analysis the genera identified were Alacaligens, Arthobacter, Burkholderia, Psuedomonas, Ralstonia, Enterobacter, Micrococcus, Kluyvera. All the isolates except strains Burkholderia and Ralstonia were able to use all the aromatic hydrocarbons studied as sole source of carbon indicated by the color change of resazurine indicator. Strains Pseudomonas and Kluyvera were considered as best degraders since they degraded the broadest spectrum of tested aromatic hydrocarbons and they showed the highest microbial activity in the presence of hydrocarbons tested indicated by the rapidity of the color change of resazurine. Furthermore, strains Enterobacter and Micrococcus exhibited moderate microbial activity against all of the tested hydrocarbons. Isolate Ralstonia was able to utilize benezene, toluene and naphthalene but couldn’t utilize acenapthene. Isolate Burkholderia was able to utilize benzene, acenapthene and naphthalene, but their proliferation was not supported by toluene.