Applications In Detergent Industries Research Articles

Detergent-Compatible Amylases Produced by Fungal Species and Their Applications in Detergent Industry

Detergent enzymes are currently vital and important ingredients of modern powder and liquid detergents. They catalyze various biochemical processes intra-or extracellularly in the detergent industries. Microorganisms are the favored source of detergent compatible fungal amylases compared to plant or animal sources. Detergents cannot be produced in the developed countries without a mixture of detergent enzymes supplementation because they are essential components in the detergent industries. In countries under development, various produced detergents do not have enzymes. Amylases, proteases, pectinases, lipases, mannanases, and cellulases are generally the hydrolytic enzymes used in the detergent industries. They remove all soft, tough and stubborn starchy food, protein, pectin, fatty, mannane and cellulose-based stains attached to the various substrates like fabrics or glassware. A number of review articles on detergent-compatible bacterial and fungal enzymes exists, but none specifically covers detergent-compatible fungal amylases. In the present review, production and aspects of detergent fungal amylases are discussed with an emphasis on the stability of fungal amylases in the presence of detergent constituents. The article will help other worldwide research to study the properties of detergent pectinases and mannanases which are not extensively studied.

Purification and characterization of lipase enzyme from endophytic Bacillus pumilus WSS5 for application in detergent industry

Lipolytic enzymes from endophytic microbes have been the focus of intense and growing research in view of the applicability of such enzymes in various industrial applications and endophytic microorganisms may emerge as a potential source of lipases with distinct characteristics. The current work involved purification and characterization of the lipase from an endophytic-bacteria isolated from the stem of Withania somnifera L. Dunal. Based on the analysis of morphology and 16S rDNA sequence, the endophytic bacteria was identified as Bacillus pumilus and designated as B. pumilus WSS5. Purified lipase from the B. pumilus WSS5 was found to be of 27KDa as revealed by SDS-PAGE with optimum activity at 37 °C and pH 8. The stability of the enzyme in presence of various metal ions, surfactants, inhibitors, and organic solvents suggested that this enzyme could be exploited in leather and detergent industries. In addition, the stain removal potential of the crude and purified lipase was determined on cotton fabric pieces stained with vegetable oil. Lipase added to water along with detergent significantly enhanced the stain removal efficiency of a commercially available detergent. The current findings suggest the potential of B. pumilusWSS5 lipase owing to promising properties such as the alkali-thermostability, organic solvent-tolerance, and broad substrate specificity that makes it a strong candidate for application in detergent formulations, biotransformation, industries, and medicine. To our knowledge, this is the first report on the purification and characterization of lipase from bacterial endophyte isolated from the stem of Withania somnifera L. Dunal.

Purification and Characterization of Bromelain from Waste Parts of Ananas comosus for its Application in Detergent Industry

Purification and Characterization of Bromelain from Waste Parts of Ananas comosus for its Application in Detergent Industry

Purification, characterization and potential detergent industry application of a thermostable α-amylase from Bacillus licheniformis RA31

α-Amylase is an endoamylase catalysing the degradation of starch into maltose, maltotriose and glucose. The enzyme isolated from microbial sources possess unique properties of thermostability thus making it a useful tool in the detergent industry. Here, we explored a strain of thermophilic bacteria Bacillus licheniformis for its potential application in detergent industry. The B. licheniformis RA31 was isolated from soil samples of hot spring in Rampur district of Himachal Pradesh, India and grown on optimized media to produce thermostable α-amylase. The enzyme was ethanol precipitated, purified (12.93 fold, 55.52% yield and 621.93 U/mg specific activity) to homogeneity with a single band on SDS-PAGE (66 kDa) and native-PAGE (68 kDa). Purified enzyme displayed best activity in pH 8 buffer and 80% activity was retained in pH 7 and 10. It showed temperature optima at 70°C. Its activity was decreased at 70°C (70% after 4 h), 80°C (65% after 4 h) and 90°C (50% after 1 h). The enzyme was stimulated (126%; 5 mM) by barium chloride. It was relatively stable in the presence of commercial detergents (109-125%), SDS (84%), Tween 20 (88%), EDTA (72%) and β-ME (70% at 10 mM). Km and Vmax for the enzymatic hydrolysis of starch were 0.339 mg/mL and 1.450 mg/min, respectively. The enzyme revealed the highest specificity towards wheat starch granule (140% after 1 h) and SEM analysis displayed its biodegradation (2-10 h). Improved cleaning efficiency of potato curry stained fine cotton clothes were observed with enzyme assisted detergent advance treatment (0.02% w/v). The enzyme showed potential applications in detergent industry.

Statistical optimization of a novel extracellular alkaline and thermostable amylase production from thermophilic Actinomadura keratinilytica sp. Cpt29 and its potential application in detergent industry

Amylolytic enzymes are biocatalysts, which are widely used in several industrial sectors. The present study was carried out to investigate amylase production by a thermophilic actinobacteria strain Actinomadura keratinilytica Cpt29. Initially, optimization of nutritional and environmental factors was performed using the One Variable at Time (OVAT) approach and Plackett–Burman design. Further, the optimum level of each factor was determined using Box–Behnken design (BBD). The different investigations employed showed that corn starch and tryptone were found to be the best carbon and nitrogen sources, respectively. Corn starch, tryptone, and pH seem to be the most influential variables for amylase production. Enhancement of 1.5-fold in the catalytic activity was reached after optimization. The obtained enzyme was characterized. Optimal enzyme activity was recorded with an alkaline pH of 8.0 at 70 °C and significantly enhanced by Ca 2+ and Mg 2+ . Good stability was retained over a broad range of pH and temperature. Also, the obtained amylase showed strong tolerance to the commercial detergents tested and excellent cleaning efficiency, which makes this enzyme an interesting new additive for detergent formulations. • Actinomadura keratinilytica Cpt29 thermophilic actinomycete used for amylase production. • Corn starch and tryptone are the best carbon and nitrogen sources according to the One-factor-at-a-time optimization. • Enhancements of 1.5 fold in amylase production with the Response Surface methodology optimization. • The amylase from Actinomadura keratinilytica Cpt29 exhibited promising properties for biotechnological applications.

Characterization of a novel lipase from Bacillus licheniformis NCU CS-5 for applications in detergent industry and biodegradation of 2,4-D butyl ester

Enzymatic degradation has become the most promising approach to degrading organic ester compounds. In this study, Bacillus licheniformis NCU CS-5 was isolated from the spoilage of Cinnamomum camphora seed kernel, and its extracellular lipase was purified, with a specific activity of 192.98 U/mg. The lipase was found to be a trimeric protein as it showed a single band of 27 kDa in SDS-PAGE and 81 kDa in Native-PAGE. It was active in a wide range of temperatures (5–55 °C) and pH values (6.0–9.0), and the optimal temperature and pH value were 40 °C and 8.0, respectively. The enzyme was active in the presence of various organic solvents, metal ions, inhibitors and surfactants. Both crude and purified lipase retained more than 80% activity after 5 h in the presence of commercial detergents, suggesting its great application potential in detergent industry. The highest activity was found to be towards medium- and long-chain fatty acids (C6-C18). Peptide mass spectrometric analysis of the purified lipase showed similarity to the lipase family of B. licheniformis. Furthermore, it degraded more than 90% 2,4-D butyl ester to its hydrolysate 2,4-D within 24 h, indicating that the novel lipase may be applied to degrade organic ester pesticides.

Purification and characterization of a novel extracellular, alkaline, thermoactive, and detergent-compatible lipase from Aeromonas caviae LipT51 for application in detergent industry

Lipase producer bacterium isolated from Erzurum was identified as Aeromonas caviae LipT51 (GenBank ID: MN818567.1) by 16S rDNA sequencing and conventional methods. Extracellular lipase was purified by ammonium sulphate precipitation, centrifugal filtration, and anion-exchange chromatography resulting in 6.1-fold purification with 28% final yield. Molecular weight was 31.6 kDa on SDS-PAGE. Lipase was stable over a broad range of pH (6–11) and temperature (25–70 °C), and showed optimum activity at pH 9 and 60 °C. Km and Vmax for pNPP hydrolysis were 0.88 mM and 34.2 U/mg protein, respectively. Ba2+, Ca2+, Co2+, Cu2+, Fe3+, and Mg2+ increased activity, while Mn2+, Mo2+, Ni2+, Zn2+, and other additives partially decreased. Activity and stability increased with laundry detergent and slightly decreased with handwash and dishwashing detergents. Alkaline and thermostable lipase from newly isolated A. caviae has been shown for the first time to be remarkably compatible with laundry detergent and improve washing performance by enhanced oil-stain removal.

Production and characterisation of lipase for application in detergent industry from a novel Pseudomonas helmanticensis HS6

The aim of this study was to explore novel source of lipase from biodiversity hot spot region of Sikkim with activity at broad temperature range for application in detergent industry. Among the isolates, Pseudomonas helmanticensis HS6 showed activity at wide range of temperatures was selected for lipase production. Statistical optimisation for enhanced production of lipase resulted in enhancement of lipase activity from 2.3 to 179.3 U/mg. Lipase was purified resulting in 18.78 fold purification, 5.58% yield and high specific activity of 3368 U/mg. The partially purified lipase was found to be active in wide range of temperature (5–80 °C) and pH (6–9), showing optimum activity at 50 °C at pH 7. Peptide sequences on mass spectrometric analysis of purified lipase showed similarity to lipase family protein of three species of Pseudomonas. Both crude and purified lipase retained residual activity of 40–80% after 3 h of incubation with commercial detergents suggesting its application in detergent industry.

Exploration of detergent-stable alkaline α-amylase AA7 from Bacillus sp strain SP-CH7 isolated from Chilika Lake

An alkali-tolerant bacterial isolate CH7 was isolated from Chilika Lake sediments that produced α-amylase at high pH. It showed >15 mm zone diameter at pH 11.0 on agar plate containing 1% starch hence considered for study. It survived up to 10% NaCl concentration on nutrient agar plates. Taxonomically it was identified as Bacillus sp. strain SP-CH7 having GenBank Accession No. KU761266. Optimized alkaline α-amylase, AA7 showed 202.857 specific activity (IU/mg), 65.95 purification fold and 25.94 yield (%) respectively. Molecular weight was revealed by SDS-PAGE as 55 kDa. The Km value was 0.688 mg/mL and so was 2.342 IU/mL Vmax obtained for AA7 towards starch. This enzyme showed 92.79% and 91.14% stability at pH 11.0 and 13.0 respectively and up to 75 °C stability was also noted. Enzyme activity was stimulated by Co2+, Na+ and Ca2+. It was stable in presence of EDTA, SDS, all the powdered detergents as well as with liquid detergent “Ezee” which vouches for its potential application in Detergent industries.

Characterization of partially purified alkaline protease secreted by halophilic bacterium Citricoccus sp. isolated from agricultural soil of northern India

Abstract The present study reports the isolation of protease producing bacteria Bact2 from agricultural soil of Regional Centre of Soil Salinity Research Institute, Lucknow, India. It was identified as Citricoccus sp. (KC522120.1) by 16S rRNA gene sequencing. The optimum pH and temperature for maximum enzyme production was observed to be at 10 and 40 °C respectively. The enzyme protease was partially purified and recovery of enzyme was 77.16%. A 1.66 fold increase in enzyme activity was also observed in comparison to original. After purification, the partially purified enzyme (PPE) was characterized. The optimum pH and temperature of partially purified enzyme (PPE) activity was observed to be at 10 and 40 °C respectively. The activity of PPE was enhanced in presence of Ca2+ and Mg2+ while it was inhibited by Cu++ and Na+. The PPE was completely inhibited by Phenyl methyl sulphonyl fluoride (PMSF) indicating that it belongs to serine protease group. Finally, enzyme was found to be compatible with detergents and effective in removing stains of tea and ink indicating it's application in detergent industry. Besides, to our knowledge till date production of alkaline protease from Citricoccus sp. has been reported first time in the present manuscript.

Biochemical characterization and homology modelling of cold-active alkophilic protease from Northwestern Himalayas and its application in detergent industry

Abstract Proteolytic enzymes are one of the largest classes of commercial enzymes used in industrial applications especially alkophilic protease is required by detergent industry. Cold-active alkophilic protease producing psychrotrophic bacterium was isolated from soil sediment of Wular lake Kashmir and identified as Bacillus subtilis WLCP1. The molecular weight of purified protease was 38 kDa, determined by SDS-PAGE. The optimal pH of the purified protease was 10.0 and was stable at pH 7.0–11.0. The optimum temperature of the enzyme was 15 °C, lowest to be reported for protease producing Bacillus sp. and can work up to 40 °C at pH 10. The protease activity was inhibited by phenylmethylsulfonyl fluoride (PMSF) while improved by Ca2+ and Cu2+ indicating the enzyme is serine alkaline protease. Purified genomic DNA from Bacillus subtilis WLCP1 was cloned and sequenced that encodes a protein of 378 amino acids. Two 3D structure models were built using the structure of Subtilisin E and SUBTILISN BPNꞌ as templates and concluded by Ramachandran Plot. The purified protease from Bacillus subtilis WLCP1 with detergent enhanced the removal of blood stains under cold conditions, making it extremely useful as detergent additive for cold washing due to its ability to work in alkophilic pH and low temperature ranges, and first ever to be reported from the North Western Himalayas, exploring the range of habitat for industrial enzymes.

Laundry Detergent Compatibility of Papain Like Protease Purified From Piper Betel Leaves

Enzymes have wide applications in detergent industry from early 1900’s. Mostly, clothes are soiled by protein based grime. Most of the detergents have either amylase / protease. Various sources were scrutinized for potent protease activity and Betel leaves were selected, the enzyme was then isolated, purified to homogeneity by ammonium sulphate precipitation, DEAE-Cellulose and gel permeation chromatographic techniques. The enzyme was monomeric in nature with a molecular mass of 38kDa as determined by native PAGE and SDS-PAGE. The enzyme shows maximum activity at 60oC and pH 4.0. The Km and Vmax of the enzyme were 4x10-3M and 54µmol/min/mg respectively. The enzyme was categorically inhibited by PCMB and iodo-acetamide suggesting it to have papain like nature. The stability of the enzyme is assessed over the stretch of alkaline pH and temperature. This evaluation validates the stability of the enzyme and its use in detergent formulations. It was evident that after adding the enzyme preparation the stains (tea, chocolate, blood) were removed much better than that of the controls, which affirms that papain like enzyme from betel leaves, enhances detergent activity.

Identification and characterization of a novel cold-tolerant extracellular protease from Planococcus sp. CGMCC 8088.

A psychrophilic extracellular protease was isolated from the marine bacterium Planococcus sp. M7 found in the deep-sea mud of the Southern Indian Ocean. The mature protease is about 43kDa and contains 389 amino acids. Sequence alignment revealed that the protease whose catalytic triad was comprised of Ser224, Lys249, and Gln253 contains a catalytic module belonging to the serralysin-type protease family 41, and displays 46.55% identity with the experimentally verified serine protease from Bacillus subtilis str. 168. The enzyme displayed an alkaline mesophilic preference with an optimum pH of 10.0 and an optimum temperature of 35°C. The enzyme retained its activity from 5 to 35°C and was resistant to repeated freezing and thawing, but was completely inactivated at 55°C. Calcium ions had a protective effect against thermal denaturation. More than 60% of the maximum activity was retained at pH values in the range of 5.0-11.0. Almost no activity loss was detected after 1h of incubation at pH 8.0-10.0 and 20°C, or with 1.0% SDS. Most important, this protease also showed good stability and compatibility with the standard enzyme-free detergent, which indicates its special interest for applications in detergent industry.

Role of Q177A and K173A/Q177A substitutions in thermostability and activity of the ELBn12 lipase.

Thermostable lipases have many applicationsin detergent industries and in organic synthesis.Thereare many ways to improve thermal stability of enzymes,for example, higher hydrophobicity, greater structural packing, higher content of the charged residues, and lower thermolabile ones. In this study, thermolabile Gln (sensitive to higher temperatures)was substituted with Alain native ELBn12 and mutated K173A lipases to examine its effect on thermal stability and activity of the lipases. Single (Q177A) and double mutants (K173A/Q177A) were expressed inEscherichia colipLysS. The Q177A variant increased both activity and thermostability of the lipase, whereas K173A/Q177A had a negative effect on the lipase activity and only had better thermal stability than the native at 50°C. pH stability of the double mutant between 9.0 and 11 was also lower than the other variants. Stability analysis in the presence of chemicals showed that Q177A mutant had better activity with 50% (v/v) organic solvents. On the other hand, K173A lipase showed increased activity with 1% (w/v) nonionic surfactant, and finally K173A/Q177A had better stability with 10 mM metal ions compared to the native lipase.

Purification and characterization of lipase by Bacillus methylotrophicus PS3 under submerged fermentation and its application in detergent industry.

Lipase production bacterial isolate was isolated from soil of service station and identified as Bacillus methylotrophicus PS3 by 16SrRNA with accession number |LN999829.1|. Lipase enzyme was purified by sequential methods of ammonium sulfate precipitation and Sephadex G-100 gel column chromatography. The molecular weight of purified enzyme was 31.40 kDa on SDS-PAGE. This purification procedure resulted in 2.90-fold purification of lipase with a 24.10% final yield. The purified lipase presented maximal hydrolytic activity at a temperature of 55 °C, and pH of 7.0. Lipase activity was stimulated by Triton X-100 and SDS with Mg2+ and Ca2+ metals employ a positive effect and outlast its stable in organic solvent i.e. methanol and ethanol.

Isolation and Screening of Protease Producing Bacteria from Local Environment for Detergent Additive

Proteases are among the most important hydrolytic enzymes that found in every organism to undertake important physiological functions. They are multipurpose enzymes used in various industries such as detergent, silver recovery, food, pharmaceutical, leather, and textile industries. This work aimed to produce protease from indigenous microbes for use as detergent additive. Isolation of protease producer was undertaken using skim milk agar medium. Crude enzyme was characterized in terms of wash and stain removal tests. A total of 188 protease positive bacteria were isolated from seven samples collected from Arba Minch town. Out of 36 alkaline protease producing bacteria, one isolate designated as Bacillus sp. Cab44 was selected. The optimum activity was reached at pH 9 and 50°C. The enzyme was stable in the pH range of 7 to 10. It retained 75%, 86% and 72% activity after one hr pre-incubation at 50°C, in 15% H2O2 and 0.3% commercial detergent respectively. The enzyme activity was increased by Mg2+, Cu2+, and Mn2+, was not affected by Ca2+ but decreased by Zn2+, Hg2+ and Fe2+. It removed stains of blood and egg yolk on cotton cloth at pH 9, 40°C, 5.07 U/ml in 30-40 min. These properties suggest that protease from Bacillus sp. Cab44 could find potential application in detergent industries as good candidate of additive in detergent formulation which have economic implication.

Production and Partial Characterization of α-Amylase Enzyme from Bacillus sp. BCC 01-50 and Potential Applications.

Amylase is an industrially important enzyme and applied in many industrial processes such as saccharification of starchy materials, food, pharmaceutical, detergent, and textile industries. This research work deals with the optimization of fermentation conditions for α-amylase production from thermophilic bacterial strain Bacillus sp. BCC 01-50 and characterization of crude amylase. The time profile of bacterial growth and amylase production was investigated in synthetic medium and maximum enzyme titer was observed after 60 h. In addition, effects of different carbon sources were tested as a substrate for amylase production and molasses was found to be the best. Various organic and inorganic compounds, potassium nitrate, ammonium chloride, sodium nitrate, urea, yeast extract, tryptone, beef extract, and peptone, were used and beef extract was found to be the best among the nitrogen sources used. Temperature, pH, agitation speed, and size of inoculum were also optimized. Highest enzyme activity was obtained when the strain was cultured in molasses medium for 60 h in shaking incubator (150 rpm) at 50°C and pH 8. Crude amylase showed maximal activity at pH 9 and 65°C. Enzyme remained stable in alkaline pH range 9-10 and 60–70°C. Crude amylase showed great potential for its application in detergent industry and saccharification of starchy materials.

Evaluation of a novel thermo-alkaline Staphylococcus aureus lipase for application in detergent formulations.

An extracellular lipase of a newly isolated S. aureus strain ALA1 (SAL4) was purified from the optimized culture medium. The SAL4 specific activity determined at 60 °C and pH 12 by using olive oil emulsion or TC4, reached 7215 U/mg and 2484 U/mg, respectively. The 38 NH2-terminal amino acid sequence of the purified enzyme starting with two extra amino acid residues (LK) was similar to known staphylococcal lipase sequences. This novel lipase maintained almost 100% and 75% of its full activity in a pH range of 4.0–12 after a 24 h incubation or after 0.5 h treatment at 70 °C, respectively. Interestingly, SAL4 displayed appreciable stability toward oxidizing agents, anionic and non-ionic surfactants in addition to its compatibility with several commercial detergents. Overall, these interesting characteristics make this new lipase promising for its application in detergent industry.

Production and characterization of surfactant-stable fungal keratinase from Gibberella intermedia CA3-1 with application potential in detergent industry

AbstractSurfactant-stable keratinases with good properties are promising candidates for extensive applications in detergent industries. A novel fungal keratinase-producing strain,

Optimization of alkaline protease production from a locally isolated Bacillus sp. ZR-5: Potential application as a detergent additive

The aim of this study was to optimize protease production using a novel Bacillus sp. ZR-5 strainisolated from the soil, and evaluate its application in detergent industry. The proteolytic activityof the strain was demonstrated using gelatin hydrolysis screening test. Protease productionoptimization was carried out using a two-step approach: a conventional method in order toidentify the best carbon and nitrogen sources followed by the application of response surfacemethodology (RSM) to optimize the factors, which include temperature, pH and incubationtime. Glucose or fructose (5 g/L), wheat bran (5 g/L), temperatures of 25 and 55°C, pH 10.0 andan approximate incubation time of 44 h, were determined as the optimal conditions according tooptimization processes. Validation tests were carried out under these conditions and the resultswere in good agreement with RSM predicted data. The in-gel activity (zymogram) test showedtwo hydrolytic zones with 66.2 and 36.5 kDa molecular weight on the casein containingpolyacrylamide gel. The high compatibility in the presence of detergent powder and washingperformance test suggested that the crude enzyme could be an appropriate choice as a detergentadditive in detergent industries.