Clear Zone Research Articles

In vitro antimicrobial and antioxidant activities of Sambucus williamsii and Sambucus pendula

The present study aimed to compare the in vitro antimicrobial and antioxidant activities of Sambucus williamsii and Sambucus pendula. The antimicrobial activity of the two plants was evaluated using the disc diffusion and minimal inhibition concentration (MIC) method against three Gram-positive bacteria (Bacillus cereus ATCC 11778, Bacillus subtilis ATCC 9327 and Listeria monocytogenes ATCC 15313), four Gram-negative bacteria (Escherichia coli ATCC 15489, Psendomonas aeruginosa ATCC 27853, Pseudomonas fluorescens KCCM 41443 and Salmonella typhimurium KCCM 11862) and one yeast (Saccharomyces cerevisiae IFO 1950). The tested ether and ethyl acetate fractions of ethanol extract from Sambucus williamsii showed significant antimicrobial activity against Bacillus cereus and Pseudomonas fluorescens; conversely, the ether fraction of Sambucus pendula leaf showed no clear zone formation against any tested bacteria. In general, the fractions of the two Sambucus species exhibited a lower MIC against tested Gram-negative bacteria than the tested Gram-positive bacteria. The antioxidant activity was evaluated by 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging assay. The results showed that the half inhibitory concentration (IC50) for the ethanol extract of Sambucus williamsii was the lower value, which means the greater antioxidant activity. On the other hand, the IC50 value of the hot water extract of Sambucus pendula was the lower value. The richness of the total polyphenol contents of the two Sambucus implies their potential as raw material sources for the pharmaceutical and cosmeceutical industries.

Isolation and Identification of Soil-Derived Amylase-Producing Bacteria

Soil is a vital ecosystem for various life forms, including bacteria. Bacteria are the main group of soil microbes, mostly because of their ability to produce various extracellular enzymes. Amylases are the enzymes that break down starch, which are essential in biotechnology industries such as food, fermentation, textiles, and paper manufacturing. The demand for amylase is often high since many enterprises do not have access to local supplies of the enzyme. The objective of the current investigation is to isolate and identify bacterial isolates that can produce the enzyme amylase from soil samples. Soil samples were collected from the unexplored Biligiriranga hills (11.9988° N, 77.1398° E) in south-western Karnataka Bacterial isolates were isolated from soil samples and were primarily screened on starch agar medium to determine the amylase production. Three out of fifteen bacterial isolates that produced amylase were selected based on the highest clear zone around the bacterial colony. These three bacterial isolates were identified both microscopically and biochemically. Gram staining demonstrated that the isolates are rod-shaped and gram-positive organisms. Biochemical studies indicated that the isolates were positive for the catalase and starch hydrolysis tests, whereas negative for the potassium hydroxide and methyl red tests. Besides, 16S rRNA gene amplification and sequencing confirmed the isolate as Bacillus subtilis. The 16S rRNA sequence of B. subtilis strain B has been deposited in the NCBI GenBank. The optimal growth conditions observed in B. Subtilis. The optimal growth conditions observed in B. Subtilis were 37° C temperature at pH 7 and incubation period for 37h. Thus, the isolation and identification of soil-derived amylase-producing bacteria are crucial for understanding their potential applications in various industrial processes, such as bioremediation, waste management, and the production of enzymes for food, pharmaceuticals, and biofuel industries.

Antifungal Potential of Bacillus subtilis subsp. inaquosorum RLS76 for Management of Fusarium Wilt Disease of Bt- Cotton

Fusarium wilt is a destructive disease affecting Bt cotton that is caused by Fusarium oxysporum f. sp. vasinfectum. This seed and soilborne disease pose a significant risk to Bt cotton production worldwide, including India. Therefore, to cope with this problem in an eco-friendly manner, we explored the potentiality of the rhizobacterial isolate possessing antifungal activity as a natural arsenal against the fungal pathogen fusarium wilt of Bt cotton. The rhizobacterial isolate RLS76 inhibited 85.39% of the fungal phytopathogen in the dual culture method. The RLS76 isolate's 16S rRNA partial gene sequence was determined to be Bacillus subtilis subsp. inaquosorum RLS76, with a closest phylogenetic affiliation of 99.53%. The crude extract of Bacillus subtilis subsp. inaquosorum RLS76 demonstrated 34.38% inhibition of the growth of the pathogen after a week of incubation by using an agar well diffusion method. Bacillus subtilis subsp. inaquosorum RLS76 produced volatile organic compounds that inhibited the fungal growth of phytopathogens by 54.84% in a closed environment. Also, GCMS analysis of the crude extract of Bacillus subtilis subsp. inaquosorum RLS76 identified four compounds, namely Pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-; Pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3-(2-methylpropyl); 2,5-cyclohexadiene-1,4-dione, 2,5-Dihydroxy-3-methoxy-6-methyl-; 9,12-Octadecadienoic acid (Z,Z)-,phenylmethyl ester. Out of four compounds, three exhibit antimicrobial activity except 2,5-Dihydroxy-3-methoxy-6-methyl. The RLS76 strain of Bacillus subtilis subsp. inaquosorum displayed a transition in hue, shifting from blue to orange, surrounding the colony on CAS medium by producing siderophore qualitatively. The quantitative siderophore produced in the crude extract was 86.06 SU after 60-72 h of incubation. The RLS76 strain of Bacillus subtilis subsp. inaquosorum also showed the ability to produce a proteolytic enzyme outside the cell. This enzyme caused a noticeable clear zone around the colony when the bacteria were grown on a skim-milk agar plate. The Bacillus subtilis subsp. inaquosorum RLS76 also exhibited phosphate solubilization activity with the clear halo around the colonies on NBRIP medium. All these results suggest that Bacillus subtilis subsp. inaquosorum RLS76 exhibits good biocontrol as well as a plant growth promoting candidate against the phytopathogen of Fusarium wilt of Bt cotton.

Isolation, screening and characterization of efficient cellulose-degrading fungal and bacterial strains and preparation of their consortium under in vitro studies.

In this investigation, cellulose-degrading fungi and bacteria were isolated from different partially decomposed cellulose-rich substrates, such as groundnut residues, rice straw, and rotten wood, following dilution plating techniques on carboxymethyl cellulose agar media and screening for potential cellulose degradation ability. The development of a clear halo zone surrounding the microbial colonies during the initial screening process using the Congo red test (20 isolates) suggested cellulose hydrolysis, and the highest cellulase production activity was implied by the isolates with the largest clear zone ratio (9 isolates). Using both macroscopic and microscopic examinations, as well as standard biochemical tests outlined in Bergey's Manual of Determinative Bacteriology, the genus-level identification of fungi and bacteria was accomplished. In order to molecularly identify the 4 isolatedfungal and bacterial strains at the species level after being ultimately selected for cellulase production potential under in vitro studies, fungal and bacterial DNA was extracted and amplified by PCR using the universal primers ITS1 and ITS4 for fungi (ITS rRNA, 5.8S rRNA) and 8F and 1492R for bacterial isolates (16S rRNA). After sequencing, the PCR results were compared to other comparable sequences in GenBank (NCBI). Based on the available NCBI data, phylogenetic analysis of their ribosomal gene partial sequences revealed that DAJ2 (PP086700) shares 100% homology with Aspergillus foetidus, DTJ4 (PP086699) shares 99.74% similarity with Trichoderma atrobrunnium, DBJ6 (PP082584) shares 100% identity with Priestia megaterium, and DMB9 (PP082585) shares 99.88% homology with Micrococcus yunnanensis. The cellulolytic potential of Phanerochaete chrysosporium is well established. Therefore, it was considered a standard culture for comparison and was collected from the MTCC, Chandigarh, India. Overall, all 4 selected isolates and the check organism were mutually compatible or synergistic with each other, and their consortium is useful for the accelerated decomposition of organic constituents during rapid composting.

Isolation and characterization of tanninolytic bacteria from sheep rumen contents: Assessment of tannin degrading, fibrolytic and feed digestibility potential

To counteract the negative effects of phytotannins, ruminants host a diverse community of bacteria that break down tannins. These bacteria not only possess tanninolytic activity but also have other functional relevance which has not been properly examined. Therefore, the present study was conducted to isolate and characterize tannin degrading bacteria (TDB) with possible fibrolytic activity from rumen of sheep fed tannin-rich diet. Based on the development of a clear zone on tannin-enriched agar plates, a total of 24 isolates were screened as tannin hydrolyzing bacteria. Majority of the isolates were gram positive cocci. The isolates could tolerate phenolic monomers such as ferulic acid, gallic acid, vanillic acid up to 30 mM and pyrogallol up to 10 mM however, the isolates failed to grow in presence of syringic acid, p-coumaric acid and p-hydroxybenzoic acid. Highest tannase activity was observed in isolate TDB23 while, lowest in isolates TDB2 and TDB5. In terms of fibrolytic activity, maximum endoglucanase, exoglucanase, and FPase activity was observed in isolate TDB9. In vitro digestibility of Prosopis cineraria leaves was considerably enhanced by inoculation of isolate TDB23 followed by TDB9. Therefore, the screened isolates demonstrated promising tannin and fibre degrading potential which can be further explored as direct-fed microbial in ruminants for effective utilization of tannin-rich fibrous feeds.

ANTIBACTERIAL AND DIASTASE ENZYME ACTIVITIES OF HONEY Apis mellifera FROM INDONESIA

The quality of monofloral honey from Apis mellifera as an antibacterial can be influenced by the activity of the diastase enzyme and the secondary metabolites from the nectar source. Therefore, this study aimed to compare the activity of the diastase enzyme and the effectiveness of monofloral honey derived from Apis mellifera bees as a natural antibacterial agent against Staphylococcus aureus and Escherichia coli. Sampling for monofloral honey was carried out from nine different nectar sources, namely Acacia carpa, Calliandra calothyrsus, Nephelium lappaceum, Melaleuca laucadendron, Ceiba pentandra, Mangifera indica, Coffea robusta 1, Coffea robusta 2, and Hevea brasiliensisensi. Furthermore, diastase content was determined using UV-Vis spectrophotometry and the bacterial inhibition zone using the disc diffusion method. The principal component analysis (PCA) was used to analyze the clustering of diastase enzyme and antibacterial activity. The results showed that the highest diastase activity in monofloral honey was Mangifera indica, Nephelium lappaceum, and Coffea robusta 2 at 20.00 DN. This was followed by Nephelium lappaceum, Ceiba pentandra, and Hevea brasiliensisensisensi at 10.00 DN, Acacia carpa at 6.67 DN, Coffea robusta 1 at 5.00 DN, and Calliandra calothyrsus 4.00 DN. The clear zones for Staphylococcus aureus on Coffea robusta 2, Acacia carpa, Nephelium lappaceum, Coffea robusta 1, Ceiba pentandra, Hevea brasiliensisensisensi, Nephelium lappaceum, Calliandra calothyrsus, and Mangifera indica were 19.47, 18.53, 17.73, 17.03, 16.12, 16.10, 16.03, 15.73, and 14.73 mm, respectively. Additionally, the clear zones for Escherichia coli on Ceiba pentandra, Coffea robusta 2, Acacia carpa, Coffea robusta 1, Melaleuca laucadendron, Mangifera indica, Hevea brasiliensisensisensi, Calliandra calothyrsus, and Nephelium lappaceum were 27.93, 26.13, 24.60, 24.53; 24.53, 24.07, 21.90, 21.60, and 21.53 mm, respectively. In conclusion, clustering analysis was conducted based on nectar sources to evaluate antibacterial and diastase activity. The clusters identified are cluster 1 consisting of Hevea brasiliensisensi, cluster 2 including Mangifera indica, and Nephelium lappaceum. Others are cluster 3 consisting of Acacia carpa and Calliandra calothyrsus, and clustergroup 4 including Nephelium lappaceum, Ceiba pentandra, Coffea robusta 1, and Coffea robusta 2. Therefore, it was necessary to carry out antibacterial testing of other bacteria, specifically Salmonella typhi, and determine the minimum inhibitory concentration (MIC) of honey with the best antibacterial activity in various concentration variations.

Antibacterial activity of Shallots (Allium xwakegi Araki.) cultivars in Palu Valley against Salmonella Typhi ATCC 27870 through in vitro and in silico evaluation

Background and Objectives: Shallots, recognized for their minimal toxicity, cost-effectiveness, and widespread avail- ability, are increasingly considered a viable source of biological activity. This study evaluates the antibacterial efficacy of a specific shallot cultivar from Palu Valley, Indonesia, against Salmonella typhi, the pathogen responsible for typhoid fever. Materials and Methods: Utilizing thin-layer chromatography (TLC-bioautography) and gas chromatography-mass spec- troscopy (GC-MS), the study identifies active compounds in shallot ethanol extract and employs molecular docking to assess interactions between receptors and ligands. Results: Findings indicate significant antibacterial activity, with a notable inhibition zone diameter of 31.5 mm at spot Rf 0.28 in TLC bioautography and an optimum concentration of 2% yielding an average clear zone diameter of 28.27 mm in the agar diffusion test. GC-MS analysis reveals 41 compounds, predominantly dodecanoic acid and 1,2,3-propanetriyl ester. Ad- ditionally, molecular docking reveals the lowest binding affinity (-7.3 kcal/mol) for Ergost-8-En-3-Ol, 14-Methyl-, (3.Beta,5. Alpha.) against DNA gyrase. Conclusion: This study confirms Palu Valley shallot extract's potent antibacterial effect against Salmonella typhi, highlight- ing its therapeutic potential.

Isolation and characterization of cellulase producing bacteria from forest, cow dung, Dashen brewery and agro-industrial waste.

The continuous accumulation of waste, particularly from industries, often ends up in landfills. However, this waste can be transformed into a valuable resource through innovative methods. This process not only reduces environmental pollution but also generates additional useful products. This study aims to screen novel high-efficiency cellulose-degrading bacteria from cow dung, forest soil, brewery waste, and agro-industrial waste in the Debre Berhan area for the treatment of cellulose-rich agricultural waste. The serial dilution and pour plate method was used to screen for cellulolytic bacteria and further characterized using morphological and biochemical methods. From eleven isolates cow dung 1 (CD1), cow dung 6 (CD6) and cow dung (CD3) which produced the largest cellulolytic index (3.1, 2.9 and 2.87) were selected. Samples from forest soil, and spent grain didn't form a zone of clearance, and effluent treatment and industrial waste (IW9) shows the smallest cellulolytic index. Three potential isolates were then tested for cellulolytic activity, with cow dung 1 (CD1) displaying promising cellulase activity. These bacterial isolates were then identified as Bacillus species, which were isolated from cow dung 1 (CD1) with maximum cellulase production. Cow dung waste is a rich source of cellulase-producing bacteria, which can be valuable and innovative enzymes for converting lignocellulosic waste.

Mycodegradation of low-density polyethylene by Cladosporium sphaerospermum, isolated from platisphere

Plastic accumulation is a severe threat to the environment due to its resistivity to thermal, mechanical and biological processes. In recent years, microbial degradation of plastic waste disposal is of interest because of its eco-friendly nature. In this study, a total of 33 fungi were isolated from the plastisphere and out of which 28 fungal species showed halo zone of clearance in agarized LDPE media. The fungus showing highest zone of clearance was further used to evaluate its degradation potential. Based on morphological and molecular technique, the fungus was identified as Cladosporium sphaerospermum. The biodegradation of LDPE by C. sphaerospermum was evaluated by various methods. The exposure of LDPE with C. sphaerospermum resulted in weight loss (15.23%) in seven days, higher reduction rate (0.0224/day) and lower half-life (30.93 days). FTIR analysis showed changes in functional group and increased carbonyl index in LDPE treated with C. sphaerospermum. SEMimages evidenced the formation of pits, surface aberrations and grooves on the LDPE film treated with the fungus whereas the untreated control LDPE film showed no change. AFM analysis confirmed the surface changes and roughness in fungus treated LDPE film. This might be due to the extracellular lignolytic enzymes secreted by C. sphaerospermum grown on LDPE. The degradation of polyethylene by Short chain alkanes such as dodecane, hexasiloxane and silane were identified in the extract of fungus incubated with LDPE film through GC–MS analysis which might be due to the degradation of LDPE film by C. sphaerospermum. This was the first report on the LDPE degradation by C. sphaerospermum in very short duration which enables green scavenging of plastic wastes.

Benefits of Stevia (Stevia rebaudiana B.) and Sirih (Piper betle L.) Leaves Combination Againts Dental Care

Oral health is the most important part, to improve its hygiene by maintaining teeth. Cavities are a common problem. Betel leaf as an antibacterial. Stevia leaves as bacteriostatic and bactericidal also eliminate the cause of tooth decay and gingivitis. The purpose of the study was to determine the antibacterial activity of toothpaste combined with betel leaf extract with stevia leaves against the growth of Streptococcus mutans bacteria. The study used an experimental method consisting of making betel leaf and stevia leaf extracts using the maceration method. Preparation of toothpaste with a combination of betel leaf extract with stevia leaves at a concentration of 15% with the ratio of betel leaves and stevia leaves, namely formula 1 (5%: 10%), formula 2 (7.5%: 7.5%), formula 3 (10%: 5%). Testing antibacterial activity by measuring the clear zone using the disc paper method with blood agar media. Data analysis using ANOVA and Paired T test. The results of the study obtained formula (5%: 10%) clear zone of 16.42 mm, formula (7.5%: 7.5%) of 17.24mm, formula (10%: 5%) of 17.94mm, positive control of 18.66mm, negative control of 2.75mm. Conclusion 10:5 combination toothpaste has antibacterial activity.

Plackett–Burman screening of physico-chemical variables affecting Citrus peel-mediated synthesis of silver nanoparticles and their antimicrobial activity

With the growing resistance of pathogenic microbes to traditional drugs, biogenic silver nanoparticles (SNPs) have recently drawn attention as potent antimicrobial agents. In the present study, SNPs synthesized with the aid of orange (Citrus sinensis) peel were engineered by screening variables affecting their properties via Plackett–Burman design. Among the variables screened (temperature, pH, shaking speed, incubation time, peel extract concentration, AgNO3 concentration and extract/AgNO3 volume ratio), pH was the only variable with significant effect on SNPs synthesis. Therefore, SNPs properties could be enhanced to possess highly regular shape with zeta size of 11.44 nm and zeta potential of − 23.7 mV. SNPs purified, capped and stabilized by cloud point extraction technique were then checked for their antimicrobial activity against Bacillus cereus, Listeria innocua, Listeria monocytogenes, Staphylococcus aureus, Enterobacter cloacae, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella typhimurium and Candida albicans. The maximum antimicrobial activity of SNPs was recorded against E. coli, L. monocytogenes and C. albicans with clear zone diameter of 33.2, 31.8 and 31.7 mm, respectively. Based on minimum inhibition concentration and minimum bactericidal concentration of SNPs (300 mg/l) as well as their effect on respiratory chain dehydrogenases, cellular sugar leakage, protein leakage and lipid peroxidation of microbial cells, E. coli was the most affected. Scanning electron microscopy, protein banding and DNA fragmentation proved obvious ultrastructural and molecular alterations of E. coli treated with SNPs. Thus, biogenic SNPs with enhanced properties can be synthesized with the aid of Citrus peel; and such engineered nanoparticles can be used as potent antimicrobial drug against E. coli.

Lactic Acid Bacteria Isolates and the Microbiome of Cincalok, Tempoyak, and Mandai: A Traditional Fermented Food from Kalimantan Island, Indonesia.

Indonesia has abundant traditional fermented food with various lactic acid bacteria (LAB), which can be developed into probiotics for pharmaceutical and functional food and feed products. This research is aimed at (1) obtaining and identifying LAB isolates and (2) studying the microbiome (bacterial diversity and abundance) of spontaneously-fermented traditional foods of Kalimantan Island, Cincalok, Tempoyak, and Mandai. To obtain LAB isolates, food samples were serially diluted and inoculated on MRS agar that contained 1% CaCO3 (MRSA). Isolates forming clear zones were purified and identified by DNA barcoding. The microbiome was studied using genomic-sequencing techniques and analysed for taxonomic composition. Seven pure isolates were obtained from Cincalok, two Tempoyak, and one Mandai. DNA barcoding revealed that the Cincalok seven isolates were Staphylococcus carnosus (strain HSP-S16), Tetragenococcus halophilus (FSB201), Corynebacterium phoceense, Vagococcus vulneris (SS1995), Enterococcus faecalis (S11-6), Pisciglobus halotolerans (C01), and Priestia filamentosa (P3.1); two from Tempoyak, Levilactobacillus brevis (E1D3BL1) and Lactiplantibacillus plantarum (UMCC-2996); and one from Mandai, Staphylococcus cohnii (XAAS.x13; non-LAB). The T. halophilus, E. faecalis, P. halotolerans, L. brevis, and L. plantarum belong to LAB. The P. halotolerans from Cincalok and non-LAB in these three fermented foods were the first documented report. The microbiome revealed the dominance of Firmicutes phyla in the fermented foods, with 93% in Cincalok, 89.94% in Tempoyak, and 60.32% in Mandai. On the genus level, Cincalok was dominated by Tetragenococcus 40.33%, Anaerococcus 23.29%, Vagococcus 9.27%, and Lactobacillus 6.84%. Meanwhile, Tempoyak was dominated only by Lactobacillus 89.94%. Mandai were dominated by Lactobacillus 31.97%, Proteus 17.14%, Aerococcus 16.85%, Mangrovibacter 15.15%, and Vagococcus 6.2%. However, Mandai's microbiome LAB was not culturable/isolated on MRSA. The plausibility is that those unculturable LAB require coculturing with other bacteria and additional media components to grow on MRSA. This study is the first report regarding the microbiome of Cincalok, Tempoyak, and Mandai, along with their culturable LAB isolates.

Enhancing the bioavailability and antioxidant activity of natamycin E235–ferulic acid loaded polyethylene glycol/carboxy methyl cellulose films as anti-microbial packaging for food application

This study investigated the development of biodegradable films made from a combination of polyethylene glycol (PEG), carboxymethyl cellulose (CMC) and mixtures from natamycin and ferulic acid. The films were characterized for their surface microstructure, antioxidant activity, thermal stability, mechanical properties, permeability and antifungal/bacterial activity. The addition of natamycin and ferulic acid to the film matrix enhanced antioxidant activity, thermal stability, antimicrobial activity, reduced the water vapor permeability (WVP) to 1.083 × 10−10 g × m−1s−1Pa−1, imparted opaque color and increased opacity up to 3.131 A mm−1. The attendance of natamycin and ferulic acid inside films created a clear roughness shape with agglomerates on the surface of films and caused a clear inhibition zone for Aspergillus niger, E. coli and C. botulinum. The utilization of PG/CMC/N-F packaging material on Ras cheese had a noticeable effect, resulting in a slight decrease in moisture content from 34.23 to 29.17 %. Additionally, it helped maintain the titrable acidity within the range of 0.99 % to 1.11 % and the force required for puncture from 0.035 to 0.052 N with non-significant differences. Importantly, these changes did not significantly affect the sensory qualities of Ras cheese during the storage period.

Purification, and characterization of detergent-compatible serine protease from Bacillussafensis strain PRN1: A sustainable alternative to hazardous chemicals in detergent industry

Owing to vast therapeutic, commercial, and industrial applications of microbial proteases microorganisms from different sources are being explored. In this regard, the gut microbiota of Monopteruscuchia were isolated and examined for the production of protease. All the isolates were primarily and secondarily screened on skim milk and gelatin agar plates. The protease-positive isolates were characterized morphologically, biochemically, and molecularly. Out of the 20 isolated strains,6 belonging to five different genera viz.Bacillus,Priestia,Aeromonas,Staphylococcus, and Serratia demonstrated proteolytic activity. Bacillussafensis strain PRN1 demonstrated the highest protease production and, thus, the largest hydrolytic clear zones in both skim milk agar (15 ± 1 mm) and gelatin (16 ± 1 mm) plates. The optimized parameters (time, pH, temperature, carbon, nitrogen) for highest protease activity and microbial growth of B.safensis strain PRN1 includes 72 h (OD600 = 0.56,1303 U/mL), pH 8 (OD600 = 0.83, 403.29 U/mL), 40 °C (OD600 = 1.75, 1849.11 U/mL), fructose (OD600 = 1.22, 1502 U/mL), and gelatin (OD600 = 1.88, 1015.33 U/mL). The enzyme was purified to homogeneity using salt-precipitation and gel filtration chromatography. The sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that the purified enzyme was a monomer of a molecular weight of ∼33 kDa. The protease demonstrated optimal activity at pH 8 and 60 °C. It was strongly inhibited by phenylmethylsulfonyl fluoride (PMSF), demonstrating that it belongs to the serine-proteases family. The compatibility of the enzyme with surfactants and commercial detergents demonstrates its potential use in the detergent industry. Furthermore, the purified enzyme showed antibacterial and blood-stain removal properties.

ANALYSIS OF ECO ENZYME CHARACTERISTICS WITH VARIATION OF “TAPE” YEAST CONCENTRATIONS

Eco enzyme is a multipurpose liquid which produced from organic waste. The liquid made with the ratio of raw materials in the form of brown sugar: organic waste: water is 1:3:10 which is processed using a fermentation process for 90 days. One method to shorten the fermentation time is adding yeast as catalyst throughout the fermentation process. This research aims to analyze the characteristics of the eco enzyme with variations in “tape” yeast concentrations. Raw material used in this research included organic waste (orange and melon peels with a ratio of 1:1), brown sugar, water, and “tape” yeast (yeast concentrations of 1, 2, 3, 4 and 5%w/v) through fermentation process for ten days. The eco enzyme products were tested to determine the pH value, TDS value, acetic acid content, aroma, color and protease enzyme activity. The analysis results showed that the pH value of eco enzyme products was in the range of 3.9 to 4.1, TDS (Total Dissolved Solids) value of 1339 to 1405 ppm, acetic acid content of 0.81 to 1.08%w/w, had bright yellow and deep yellow color with a fermented aroma. The selected eco enzyme product in this research was on eco enzyme with yeast concentration of 3 %w/v which had pH value of 3.9 TDS value 1403 ppm, acetic acid content of 0.81%w/w, and the color was bright yellow with fermented aroma. However, there was no clear zone around the colony in the test indicating that there was no protease enzyme activity.

Analysis of Amylase Activity in Bacteria Isolated from Hot Spring of Pentadio Resort

The hot spring of Pentadio Resort is one of the geothermal areas likely to be the place of bacteria or microorganisms that can produce amylolytic enzymes. This research aims to analyze bacteria in the Pentadio Resort hot springs that possess the potential to produce amylase enzymes, analyze the characteristics of bacterial isolates that produce amylase enzymes in Pentadio Resort hot springs and determine the types of bacteria that produce amylase enzymes in Pentadio Resort hot springs. The research employs a qualitative and descriptive method. The research involved several methods, including sample collection, media preparation, isolation and selection of thermophilic bacterial isolates, purification of thermophilic bacterial isolates, selection of isolates producing amylase enzymes, identification of selected isolates (motility test, biochemical test, gram staining), and molecular identification. The research yielded seven isolates, and isolate code A6 demonstrated the potential to produce amylase enzymes with a clear zone size of 23,26 mm, bacillus, gram positive, and positive motility. Subsequently, molecular identifidation was carried out in which, based on the results of phylogenetic tree reconstractiopisalate A6 was related to the bacteria Bacillus cereus strain MD 152, which belongs to the Bacillus cereus bacteria

Screening of microscopic fungi isolated from charcoal and wooden samples for Protease production

Proteases, otherwise called proteinases or proteolytic chemicals, have a place with a gathering of hydrolases. Microorganisms are the most well-known wellsprings of business chemicals because of their physiological and biochemical properties. Fungus from charcoal and wooden sample were secluded by sequential weakening agar plate method. The fungal disengages were to begin with evaluation of their capacity to create protease based on zone of clearance on skimmed milk agar plate. The segregates that gave excessive positive consequences in screening were identified as Aspergillus fumigatus, Fusarium species, Mucor recemossus, Penicillin citrium with the aid of inspecting the morphology and Lacto - phenol cotton blue (LPCB) staining procedure. Among those disconnects, Aspergillus fumigatus which constrained from charcoal floor, exhibited the maximum noteworthy protease movement confirming the use of the clean sector across the colony that is brooded at room temperature at a pH of round 7 for a hundred and twenty hours at 28 ˚C. Three fungal disengages chosen by beginning tests were exposed to extracellular protease creation in protease creation medium. The outcomes showed that three fungi could be utilized in the creation of protease compounds and in the upscale for modern creation.

Isolation and production of proteases from bacteria isolate from Bekasam and Rusip (Traditional Food from South Sumatera, Indonesia)

The research aims to assay enzyme activity of bacteria from traditional fermentation product (bekasam and rusip). The bacteria were inoculated in skim milk agar (SMA), and will produce clear zone. The best isolates from 9 isolates of bekasam and 9 isolates from rusip will be choosen, and the best result from bekasam and rusip were A3K2 and B3K2, respectively. Index proteolitic of A3K2 and B3K2 isolates were 10.0 and 7.0, respectively. Optimum time production of protease from A3K2 and B3K2 were 16 h and 32 h, respectively.

Calliandra haematocephala’s phytochemical analysis and antifungal effectiveness against fish pathogen in the lake of Bhopal

Aquaculture has been a major part of our Economy from ancient times and antimycotic resistant fish illnesses have emerged as a major threat in aquaculture from a very long time. As routinely used anti-fungal medications have become less effective and traditional medicinal herbs have been identified as a possible medication source that can counteract antifungal resistance. The current study will look at the antifungal efficacy of leaf and flower extracts of the nutritional and traditional medicinal plant called Calliandra haematocephala against 3 common fish fungal pathogens (Achlya, Saprolegnia and Aphanomyces) that were isolated from Lower Lake of Bhopal (M.P.) and causes illness in fishes present in Lower Lake, Bhopal (M.P.). The leaves and flowers of Calliandra haematocephala were extracted sequentially with ethanol as a solvent, and the presence of possible phytoconstituents was qualitatively evaluated. Flavonoids, phenol, alkaloids, carbohydrates, tannins and saponins were present in both flowers and leaves and steroids were found exclusively in the ethanolic extract of the leaf only not in flower extract. The antifungal activity of ethanolic extract was determined using the agar well diffusion technique. The ethanolic fraction of the flower demonstrated a wide range of activity against fish pathogens, with a maximal inhibitory or clearance activity against Saprolegnia in a amount of 20 μL (4 mm) and in leaves it showed maximum clearance zone in Achlya and Saprolegnia in 20μL (4 mm). In conclusion, Calliandra haematocephala’s extracts consist of potential secondary metabolites that might be employed as antifungal treatments for 3 prevalent fish Pathogens, and the leaf ethanolic extracts have more inhibitory efficacy than the flower ethanolic extract.

Antimicrobial Activities against Skin Flora from Tart-cherry Extracts (Prnus cerasus)

Purpose: In this study, antibacterial and anti-oxidant activities of Tart-cherry hydrothermal extracts (TCW) and ethanol extracts (TCE) against dermatological bacteria were measured, and the cell viability and anti-inflammatory effects were investigated in RAW264.7 macrophages to determine their characteristics for use as functional cosmetic materials. Methods: Anti-oxidant activity was determined using the DPPH method. The antibacterial activity against dermatophilosis was confirmed by the size of the clear zone formed by DISK diffusion and MIC measurement. The activity against an antibiotic-resistant MRSA strain was also measured. Anti-inflammatory response was identified by measuring cytotoxicity and NO activity in RAW264.7 macrophages, in which inflammatory reactions were induced with LPS. Results: Anti-oxidant activity of the TCE was 85.6%, and of TCW was 88.4% at 500 μg/mL. The size of the clear zone was confirmed using Cutibacterium acnes (18 mm), <i>Staphylococcus epidermis</i> (<i>S. epidermis</i> ) (4 mm), and <i>Staphylococcus aureus</i> (<i>S. aureus</i> ) (16 mm). No antifungal activity against <i>Candida albicans</i> was observed. RAW264.7 macrophages showed a survival rate of >90% at <250 μg/mL of extract. Additionally, a concentration-dependent inhibition of NO production was observed. Conclusion: Tart-cherry extracts possessed high anti-oxidant activity and anti-inflammatory efficacy. In addition to the antibacterial activity against the skin-residing bacteria, <i>S. aureus</i> and <i>S. epidermis</i>, high activity against the acne-causing strain <i>C. acnes</i> suggests a sufficient potential of Tart-cherry extracts as a raw material for the preparation of cosmetics and bioproducts.