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Lipases are one of the widely used biocatalyst for various purposes and so they have wide range of applications. These water soluble enzymes have unique ability of catalyzing the hydrolysis of ester bonds in triacylglycerols. Lipase is the primary digestant used to break fats into fatty acids and glycerol. Lipase is produced by different methods and via variety of sources. Isolation of lipase is a challenging task as it should be cost effective and efficient one at industrial scale. Lipase has been isolated from marine algae which is easily available in shallow water and from culturing of the same. People are not using seaweeds as a food in India so it gives us a better chance and challenge to utilize it for different purposes. In this research work we isolated lipase from different types of seaweeds from nearby area, purified it and estimated the molecular weight of lipase. The obtained lipase was tested against different physical and chemical parameters such as effect of pH on activity and stability, effect of temperature on activity and effect of different substrates on the activity of lipase showing its optimum range and specificity for different substrates.
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Int. J. Pharm. Sci. Rev. Res., 27(1), July – August 2014; Article No. 34, Pages: 188-190 ISSN 0976 – 044X
International Journal of Pharmaceutical Sciences Review and Research
Available online at
© Copyright protected. Unauthorised republication, reproduction, distribution, diss emination and copying of this document in whole or in part is strictly p rohibited.
© Copyright protected. Unauthorised republication, reproduction, distribution,
Swapnil Suresh Patil, L. Jeyanthi Rebecca*
Department of Industrial Biotechnology, Bharath University, 173, Agharam Road, Selaiyur, Chennai, Tamilnadu, India.
*Corresponding author’s E-mail:
Accepted on: 29-04-2014; Finalized on: 30-06-2014.
A protease is a member of a very large group of enzymes that have various functions in the body. Proteases break the long chainlike
molecules of proteins into shorter fragments. Proteases are found in plants, animals, including microorganisms such as fungi,
bacteria and yeast. Protease is considered to be an important industrial enzyme accounting for about 60% worldwide sale of
enzymes. The influence of parameters such as temperature, pH were evaluated.
Keywords: Algae, Electrophoresis, Protein content, Protease, purification.
he ocean is considered to be the repository of many
unexploited natural resources. Protease hydrolyzes
the peptide bonds and is also termed as
proteinase.1 Protease constitutes one of the most
important groups of industrial group having application in
different sectors of industries. Protease is widely being
used in waste management, silver recovery from X-ray
films, leather industry, and detergent industry.2-4
Proteases have found applications in molecular biology
research as well.5 A protease from Bacillus
amyloliquefaciens may be used to promote flavor
production in cheddar cheese.6 Proteases are also being
used in meat processing, dairy, digestive aid7 Protease
has found an irreplaceable role in silk industry.8,9
Reports are available on protease production by fungal
species belonging to Aspergillus genera.10 There are
reports on the isolation of protease from plant leaves and
marine waste like fish scales, crab and prawn shells.11-14
Estimation of protein content by Bradford Method
The protein concentration was measured by the Bradford
method, using BSA as a standard. Dilute the Bradford
reagent and add the sample to the diluted reagent. The
colour was changed to dark blue. The absorbance was
taken at 590 nm.15
Protein precipitation using Ammonium sulphate
Ammonium sulphate was used to precipitate the protein.
Appropriate amount of ammonium sulphate is added to
the supernatant and centrifuged for 10 mins at 10000
rpm. The pellets were resuspended in suitable buffer.
Partial purification by Sephadex column
Sephadex was used to pack the column. The solution was
fed to the column for further purification.
Protease activity
0.5 ml of partially purified enzyme was mixed with 0.5 ml
casein. The reaction mixture was incubated for 1 hr at
37°C and stopped by addition of 1 ml of 10% TCA. The
mixture was centrifuged and supernatant was collected.
0.5 ml of supernatant was mixed with Na2CO3 and NaOH
and Folin Phenol. This mixture was shaken well and OD
was taken at 650nm. One protease unit is defined as the
amount of enzyme that releases 0.5 µg/ml/min tyrosine.
The protease activity was measured
Effect of pH and Temperature
The optimum pH of the enzyme was determined using
buffer at varying pH ranges of 3-12. The effect of
temperature on the protease activity was studied by
incubating the reaction mixture at different temperature
ranging from 20-90°C.
Sodium Dodecyl Sulphate-Polyacrylamide gel
electrophoresis (SDS-PAGE)
SDS-Polyacrylamide gel electrophoresis was performed
on slab gel with separating and stacking gels by the
method of (Laemmli, 1970).16
The present study was carried out to isolate and
characterize protease from algae. The protein content of
all the samples is shown in Table 1.
Table 3 and table 4 clearly show that parameters like
temperature and pH affect the activity of enzyme.
Maximum species showed highest activity at 40°C. The
optimum pH was checked for all the samples at varied pH
range. The optimum pH range was observed between 7-
7.5 depending on species, although the maximum activity
was seen at pH 7.2.
It can be observed from the above result that the isolated
enzyme from different species had molecular weight
Isolation and Characterization of Protease from Marine Algae
Int. J. Pharm. Sci. Rev. Res., 27(1), July – August 2014; Article No. 34, Pages: 188-190 ISSN 0976 – 044X
International Journal of Pharmaceutical Sciences Review and Research
Available online at
© Copyright protected. Unauthorised republication, reproduction, distribution, diss emination and copying of this document in whole or in part is strictly p rohibited.
© Copyright protected. Unauthorised republication, reproduction, distribution,
within the range of 30-45 kDa. The bands were compared
with the standard bacterial protease run alongside.
Table 1: Protein estimation of crude and purified samples
Species Name Protein content of
crude extract
Protein content
Ulva lactuca (Puducherry) 92 82
Ulva fasciata (Puducherry) 97 82
Enteromorpha compressa
(Puducherry) 86 77
Chaetomorpha antenna
(Puducherry) 105 90
Ulva lactuca
(Mahabalipuram) 92 77
flexuosa(Mahabalipuram) 110 90
Table 1 shows the protein content of 6 species of
seaweed. The highest protein content was seen in
Enteromorpha flexuosa (Mahabalipuram) and
Chaetomorpha antenna (Puducherry) while lowest was
found in Ulva lactuca (Mahabalipuram) and
Enteromorpha compressa (Puducherry).
Table 2: Activity of protease using Casein as substrate
Species Name Place
Protease activity
Ulva lactuca Puducherry 7.33
Ulva fasciata Puducherry 8.0
compressa Puducherry 6.74
antenna Puducherry 9.4
Ulva lactuca Mahabalipuram 6.55
flexuosa Mahabalipuram 9.6
The above Table shows Enteromorpha flexuosa showing
the highest activity while Ulva lactuca (Mahabalipuram)
showed the lowest.
Table 3: Effect of Temperature on Protease activity
Species Temperature Range (°C)
20 30 40 50 60 70
Ulva lactuca 6.33 6.71 7.35 6.4 5.20 4.42
Ulva fasciata 6.92 7.31 8.0 7.37 5.32 3.9
compressa 5.22 6.35 6.77 6.20 4.39 3.55
antenna 6.78 8.42 9.49 8.11 6.22 5.14
Ulva lactuca 4.52 5.91 6.61 5.33 4.59 4.38
flexuosa 6.55 8.61 9.38 7.52 6.17 5.21
Table 4: Effect of pH on Protease activity
Species pH range
6.5 7.0 7.5 8
Ulva lactuca 6.21 7.25 7.41 6.79
Ulva fasciata 6.42 8.0 8.22 7.66
Enteromorpha compressa 5.85 6.71 6.77 5.73
Chaetomorpha antenna 8.22 9.12 9.22 8.76
Ulva lactuca 5.73 6.43 6.47 5.95
Enteromorpha flexuosa 8.46 9.48 9.59 8.66
Figure 1: Estimation of molecular weight of algal protease
using SDS-PAGE
Note: Lane 1 - Ulva lactuca (Puducherry); Lane 2 - Ulva fasciata
(Puducherry); Lane 3 Ulva lactuca (Mahabalipuram); Lane 4
Enteromorpha compressa (Puducherry); Lane 5 Chaetomorpha
antenna (Puducherry); Lane 6 Enteromorpha flexuosa
(Mahabalipuram); Lane 7 – Standard bacterial Protease
The major aim of the study carried out was to investigate,
characterize and purify the protease enzyme from various
marine algae. The enzyme extracted from different
marine algae showed positive results for protease assay
confirming the presence of protease. The activity varied
with different species and temperature and pH.
From this study, it is quite evident that marine algae are a
valuable source of Protease. Moreover, they can also be
screened for other industrially important enzymes.
1. Barrett AJ, Mcdonald JK, Nomenclature: Protease,
Proteinase and Peptidase. Biochem. J, 237, 1986, 935.
2. Ray A, Protease Enzyme- Potential Industrial Scope, Int. J.
Technol, 2, 2012, 1–4.
3. Fujiwara N, Tsumiya T, Katada T, Hosobuchi T, Yamamoto
K, Continuous Recovery Of Silver From Used X-Ray Films
Using A Proteolytic Enzyme, Process Biochem, 24, 1989,
4. Babu NKS, Lakshmi KD, Afr.J.Biotechnol., 4(7), 2005, 724-
30 kDa
Int. J. Pharm. Sci. Rev. Res., 27(1), July – August 2014; Article No. 34, Pages: 188-190 ISSN 0976 – 044X
International Journal of Pharmaceutical Sciences Review and Research
Available online at
© Copyright protected. Unauthorised republication, reproduction, distribution, diss emination and copying of this document in whole or in part is strictly p rohibited.
© Copyright protected. Unauthorised republication, reproduction, distribution,
5. János András Mótyán, Ferenc Tóth, József Tőzsér, Research
Applications Of Proteolytic Enzymes In Molecular Biology,
Biomolecules, 3, 2013, 923-942.
6. www1.Lsbu.Ac.Uk/Water/Enztech/Index.Html.
7. Chellapan S, C Jasmin, M Soorej, Basheer, KK Elayas, Sarita
G. Bhat, M. Chandrasekharn, Production, Purification And
Partial Characterization Of Novel Protease From Marine
Engyodontium Album Btmfs 10 Under Solid State
Fermentation, Process. Biochem., 41, 2006, 956-961.
8. Das Sumana, Mathummal Sudarshan, Ashoke Ranjan
Thakur, Shaon Raychaudhuri, Degumming Of Raw Silk
Fabric With Help Of Marine Extracellular Protease.
American Journal of Biochemistry And Biotechnology, 9(1),
2013, 12-18.
9. Yang Cao, Bochu Wang, Biodegradation Of Silk
Biomaterials, Int J Mol Sci., 10(4), 2009, 1514–1524.
10. Fan-Ching Y, IH Lin, Production Of Acid Protease Using Thin
Stillage From A Rice, Spirit Distillery By Aspergillus Niger,
Enzyme Microb. Technol., 23, 1998, 397-402.
11. L Jeyanthi Rebecca, S Sharmila, Merina Paul Das, F
Abraham Samuel, Production and Analysis of Protease
From Aspergillus Niger Using Fish Scales As Substrate,
Journal Of Chemical And Pharmaceutical Research, 4(10),
2012, 4597-4600.
12. L Jeyanthi Rebecca, S Sharmila, Merina Paul Das, TV
Rishikesh, S Ananda narasimhan, Production And Analysis
Of Protease And Amylase From Aspergillus Niger Using
Crab And Prawn Shell As Substrate, Journal Of Chemical
And Pharmaceutical Research, 4(10), 2012, 4542-4544.
13. S Sharmila, L Jeyanthi Rebecca, Merina Paul Das, Md
Saduzzaman, Isolation and Partial Purification Of Protease
From Plant Leaves, Journal Of Chemical And
Pharmaceutical Research, 4(8), 2012b, 3808-3812.
14. V Dhanalakshmi, L Jeyanthi Rebecca, G Revathi, S Sharmila,
Evaluation Of Basic Nutritional Parameters Of Seaweeds In
Coastal Tamilnadu, International Journal Of Biotechnology
And Biochemistry, 6(6), 2010, 921-928.
15. Bradford M, A Rapid And Sensitive Method For The
Quantization Of Microgram Quantities Of Protein Utilizing
The Principle Of Protein-Dye Binding, Anal. Biochem., 72,
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Source of Support: Nil, Conflict of Interest: None.
... A number of enzymes extracted from different marine algae show positive results for protease assay [44,45]. Their activity varied with different species, temperature and pH (Figs. 7 and 8). ...
... The optimal pH and temperature of the alkaline protease from the marine yeast A. pullulans have been found to be 9.0 and 45 • C, respectively. Studies have shown that the enzyme is activated by Cu 2+ and Mn 2+ and inhibited by Hg 2+ , Fe 2+ , Fe 3+ , Zn 2+ , and Co 2+ [45]. The use of proteases from Bacillus sp. and lactic bacteria [46][47][48] are most common. ...
... The use of proteases from Bacillus sp. and lactic bacteria [46][47][48] are most common. On the other hand, alkaline protease from the marine yeast A. pullulans has potential uses in production of bioactive peptides from shrimp (Trachypenaeus curvirostris) muscle and spirulina (Arthospira platensis) powder [45]. Fibrinolytic and fibrinogen clotting enzymes are two specific proteases with fibrinolytic activity found in a number of genus Codium including Codium intricatum (CIPs) [49]. ...
Marine environment consists of the largest sources diversified genetic pool of material with an enormous potential for a wide variety of enzymes including proteases. A protease hydrolyzes the peptide bond and most of proteases possess many industrial applications. Marine proteases differ considerably from those found in internal or external organs of invertebrates and vertebrates. In common with all enzymes, external factors such as temperature, pH and type of media are important for the activity, catalytic efficiency, stability and proper functioning of proteases. In this review valuable characteristics of proteases in marine organisms and their applications are gathered from a wide literature survey. Considering their biochemical significance and their increasing importance in biotechnology, a thorough understanding of marine proteases functioning could be of prime importance.
... (6.74-9.60 U·mg −1 ), and Chaetomorpha antenna (9.40 U·mg −1 ) [34]. Table 1. ...
... (6.74-9.60 U·mg −1 ), and Chaetomorpha antenna (9.40 U·mg −1 ) [34]. ...
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Thrombosis is characterized by the pathological formation of fibrin clots within a blood vessel, leading to the obstruction of blood flow. Fibrinolytic enzymes from microorganisms have been shown to be more efficient and safer in dissolving clots. Then, this study aimed to evaluate the cell growth and fibrinolytic enzyme production of Tetradesmus obliquus under different cultivation conditions. T. obliquus grew under autotrophic and mixotrophic conditions using different concentrations of corn steep liquor (0.25 ≤ CSL ≤ 4.00%). The cells were concentrated and lysed via two different methods (sonication or homogenization) to trigger the release of the enzyme. It was precipitated via acetone or ammonium sulfate additions and purified using ion exchange chromatography. The highest biomass productivity (Px = 130 ± 12.8 mg∙L−1day−1), specific growth rate (µmax = 0.17 ± 0.00 day−1), and fibrinolytic activity (391 ± 40.0 U∙mg−1) was achieved on a mixotrophic cultivation at a 0.25% CSL concentration. The results showed that the homogenizing method had better performance in the release of enzyme, and the precipitation with acetone obtained the highest fibrinolytic activity (567 ± 49.3 U∙mg−1). The purified enzyme showed a specific activity of 1221 ± 31 U∙mg−1 and a molecular mass of 97 kDa. So, the fibrinolytic enzyme from T. obliquus had higher activity when compared to the other fibrinolytic enzymes, being a potential source for the development of therapeutic agents in thrombosis treatment. Additional studies are needed to investigate the biochemical properties and biological profile of this enzyme.
... It is also reported that seaweeds have great potential nutritional values with components such as polysaccharide, protein, vitamins and minerals, and can be used as a source of nutrients for humans and animals (Sakhthivel and Devi 2015). Moreover, some seaweeds and other types of algae have been found to have a considerable caseinolytic activity (Pérez-Lloréns et al. 2003;Sharmila et al. 2012;Bele et al. 2014;Patil and Rebecca 2014) and thus have the potential to be explored as rennet substitutes for the coagulation of milk in cheese making. ...
... They found that the water extract of U. lactuca was able to clot milk within 5 h. Recent studies about algal proteases have focused on the analysis of their proteolytic and caseinolytic activities, but ignored their milk-clotting properties (Pérez-Lloréns et al. 2003;Sharmila et al. 2012;Bele et al. 2014;Patil and Rebecca 2014). ...
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Cheese-making involves milk coagulation as a crucial step where calf rennet has been used traditionally as the milk-clotting enzyme in the cheese industry. This study investigated milk-clotting enzymes from seven species of seaweed and evaluated the parameters for their isolation and partial purification including post-harvest processing, extraction and purification methods. The saturation degree of ammonium sulphate and the precipitation stages were evaluated to obtain optimal purification conditions, and three desalting methods, namely dialysis, desalting column and the combination of dialysis and desalting column, were investigated to determine the most suitable method for ammonium sulphate removal. Protein extracts of all seven species of seaweeds exhibited some caseinolytic activity, but the extract of unprocessed (whole) dried samples had higher protein yields and stronger caseinolytic activities. The extract from one species, Gracilaria edulis, demonstrated the ability to clot milk. The evaluation of the purification method for G. edulis extract revealed an optimum single step of 50% saturation and ammonium sulphate precipitation with dialysis as the desalting method.
... Moreover, protease activity of T. obliquus cultivated on 0.25% CSL is higher than those produced by different marine algae, such as Ulva lactuca (6.55 -7.33 U•mg -1 ), Ulva fasciata (8.00 U•mg -1 ), Enteromorpha sp. (6.74 -9.60 U•mg -1 ), and Chaetomorpha antenna (9.40 U•mg -1 ) [32]. Dry cell concentration (mg·L -1 ) Time (days) 6 No significant difference in fibrinolytic activities was observed between autotrophic (430.46 ± 40.19 U•mg -1 ) and 0.25% CSL (391.34 ± 40.03 U•mg -1 ) cultivations, which were higher than 0.50% CSL (135.63 ± 6.98 U•mg -1 ). ...
Full-text available
Thrombosis is characterized by the pathological formation of fibrin clots within a blood vessel, leading to the obstruction of blood flow. Fibrinolytic enzymes from microorganisms have exhibited promising effects to dissolve clots in a more efficient and safe way. Then, the aim of this study was to evaluate the biomass and fibrinolytic enzyme production of Tetradesmus obliquus under autotrophic and mixotrophic conditions using different concentration of corn steep liquor (CSL). Different extraction and precipitating methods were tested, and the enzyme was purified by ion exchange chromatography. More advantageous culture condition was mixotrophic using 0.25% CSL, showing the highest values of biomass productivity (Px = 169.3 ± 44.36 mg∙L-1day-1) and specific growth rate (µmax = 0.17 ± 0.00 day-1), and significant fibrinolytic production (391.34 ± 40.03 U∙mg-1). Moreover, fibrinolytic activity was higher when extracted by homogenization and precipitated using acetone, which exhibited clear zone of fibrin degradation in the fibrinolytic plate assay. Additionally, the purified enzyme showed specific activity of 1176.90 ± 140.37 U∙mg-1 and molecular weight around 97 kDa. Finally, the enzyme has higher enzymatic activity than various fibrinolytic enzymes, and the obtained enzyme has potential to be developed as a therapeutic agent in thrombosis treatment. Additional studies are need to investigate the biochemical properties and biological profile of this enzyme.
... The authors did not perform the biochemical characterization of extracted enzyme regarding pH and temperature. Patil and Rebecca (2015) reported protease production using casein as substrate, and found specific activity values of 7.33 and 6.55 U/mg for Ulva lactuca, 8 U/mg for Ulva fasciata, 6.74 U/mg for Enteromorpha compressa, 9.6 U/mg for E.flexuosa and 9.4 U/mg for Chaentomorpha antenna. The effect of pH on protein activity was evaluated, and the algae with best results, C. antenna and E. flexuosa, showed stability in the pH range between 6.5 and 8.0. ...
... Highest proteolytic activity (112.46 ± 5.04 U mg −1 ) was achieved using 1.5% corn steep liquor, which is higher than those obtained from six different species of algae, with activities ranging from 6.55-9.6 U mg −1 [34]. Concentration of 0.4% corn steep liquor was reported to optimize the ...
A fibrinolytic enzyme from the microalga Chlorella vulgaris was cultivated in mixotrophic conditions (corn steep liquor 1%), with maximum cell concentration and productivity of 1637.45 ± 15 mg L⁻¹ and 181.93 mg L⁻¹ day⁻¹, respectively. The enzyme was purified by a single chromatographic step, using ion exchange chromatography, with peaks correlated to 2 bands with respective molecular weights of 100.3 kDa and 45 kDa, determined by SDS-PAGE. A fibrin zymography was performed and found that only the 45 kDa band has fibrinolytic activity. After purification, fibrinolytic activity was estimated at 1834.6 U mg⁻¹ and 226.86 mm², by fibrin plate assays. The enzyme activity was enhanced in the presence of Fe²⁺ and inhibited by phenylmethane sulfonyl fluoride (PMSF) and ethylenediamine tetracetic acid (EDTA), which suggest it to be a metal-dependent serine protease. The extract also showed a red blood cell lysis <4% and in vitro thrombolytic activity of 25.6% in 90 min of reaction. These results indicate that the fibrinolytic enzyme from C. vulgaris may have potential applications in the prevention and treatment of thrombosis.
... Different bacteria, fungi, and actinomycetes are the major source of microbial protease enzyme. Protease are also isolated from marine algae (Swapnil and Jeyanthi Rebecca, 2014; Sharmila et al., 2012). Protease have been partially purified from marine waste like fish scales and crab and prawn shells (Jeyanthi Rebecca et al., 2012a; Jeyanthi Rebecca et al., 2012b) They have extensive industrial applications in the production and processing of detergent, food, pharmaceuticals, leather, textiles etc. (). ...
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Actinomycetes are an important group of microorganisms that has wide industrial application in the production of antibiotics and enzymes. In this study 109 isolates of Actinomycetes were isolated from marine soil samples collected from the coastal area of Chennai, India. The isolates were identified by morphological studies. Form the total 109 isolates, 40 showed proteolytic activity which was indicated by growth and clear zone formation on casein-skim milk agar media plate. Among the 40 protease producing strains the enzyme activity was highest in 8 strains namely, MB22, MB41, MB16, MB47, BN3, BN16, NK17, NK15. Potential strains were further used for protease production and for the purpose of screening for fibrinolytic enzymes.
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Protease occurs naturally in all organisms and is an essential constituent for all the existing live forms. Microorganisms such as bacteria and fungi and yeast are the main source of protease enzyme. They act as an important industrial enzyme occupying for about 60% of total enzyme market. In this study, protease was isolated from various leaves such as Coriandrum sativum, Nicotiana tobaccum, Murraya koenigii, and Moringa oleifera was partially purified. Then their specific activity and optimum pH were checked. Among the four plant species, the protease activity was found to be more in Nicotiana tobaccum (5.6 units/mg of protein) followed by Moringa oleifera (2.46 units/mg of protein), Murraya koenigii (2.02 units/mg of protein) and least in Coriandrum sativum (1.56 units/mg of protein). The optimum pH was found to be 7.2 for all the samples.
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Protease secreting microbe was isolated and characterized on the basis of their morphological, biochemical, physiological and 16S rDNA based molecular properties. The extracellular protease was quantified and characterized. Protease was used for different time (4, 8, 12 and 24 h) at different temperature (RT and 37°C) for optimization of the degumming process for raw silk fabric with enzyme dosage (0.2-1 unit/cm 2 of fabric). Post-enzymatic treatment, the fabric quality and texture was compared with conventionally treated as well as untreated fabric in terms of degumming loss, tensile strength and yarn count and colour fastness to light/water. The isolate SM1 (Bacillus thuringensis) was able to grow in Carbon Minimal Salt Medium (CMSM) with jaggery or tamarind as the carbon source (0.3% w/v). Energy Dispersive X-Ray Fluorescense (EDXRF) data showed intracellular accumulation of heavy metal by the isolate. Extracellular protease was able to degum silk fabric within 4 h at RT with enzyme concentration of 0.8unit/cm 2 and the maximum degumming loss was 21.72%. Post enzymatic degumming, a shiny texture was observed under Environmental Scanning Electron Microscope (ESEM) and the yarn volume also increased. Utilization of CMSM made the process cost effective during large scale application. Intracellular metal accumulation and growth in a wide range of temperature and pH made the isolate a potential candidate for bioremediation. Extracellular protease with significant degumming property could be used as an eco friendly approach as compared to the conventional chemical treatment.
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About 1,234,332 tons of marine food is consumed on a weekly basis all around the world leading to the production of marine food processing waste like fish scales, crab and prawn shells, fins bones etc. In the present study crab and prawn shells were collected from a local fish market powdered and used as a substrate for Aspergillus niger and the activity of protease and amylase were analysed after three days. Two mutated strains of A. niger, were also used apart from the wild type A. niger strain.
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Proteolytic enzymes (also termed peptidases, proteases and proteinases) are capable of hydrolyzing peptide bonds in proteins. They can be found in all living organisms, from viruses to animals and humans. Proteolytic enzymes have great medical and pharmaceutical importance due to their key role in biological processes and in the life-cycle of many pathogens. Proteases are extensively applied enzymes in several sectors of industry and biotechnology, furthermore, numerous research applications require their use, including production of Klenow fragments, peptide synthesis, digestion of unwanted proteins during nucleic acid purification, cell culturing and tissue dissociation, preparation of recombinant antibody fragments for research, diagnostics and therapy, exploration of the structure-function relationships by structural studies, removal of affinity tags from fusion proteins in recombinant protein techniques, peptide sequencing and proteolytic digestion of proteins in proteomics. The aim of this paper is to review the molecular biological aspects of proteolytic enzymes and summarize their applications in the life sciences.
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Atualmente, há uma tendência de utilizar baixas temperaturas em limpeza de lavanderia por razões ambientais e econômicas, situações estas que tornam indispensáveis o uso de enzimas em produtos detergentes. Devido às lipases serem excelentes catalisadoras em soluções e em interface água-óleo, elas são potencialmente apropriadas para aplicação na remoção de manchas de gorduras em lavanderia industrial e detergentes domésticos. O efeito de diferentes detergentes comerciais e surfactantes sobre a atividade enzimática da lipase de Fusarium oxysporum foi observado através de ensaios com p-nitrofenilpalmitato (pNPP). A enzima foi compatível com vários surfactantes iônicos e não-iônicos como também com detergentes comerciais. Atividade lipolítica foi fortemente inibida por Sulfato Dodecil de Sódio (SDS), mas não por Triton X-100 e Triton X-114. As melhores condições de ensaio observadas para esta lipase foram pH 8,0 e 50ºC. A enzima foi estável em pH alcalino e manteve 93% da atividade residual durante 1 h de incubação a 60ºC. A maior atividade lipolítica foi medida com triglicerídeos de ácidos graxos de cadeia média e longa (C8-C18). A especificidade hidrolítica da enzima em vários óleos e gorduras testados foi variável. Todas estas propriedades e sua resistência a vários surfactantes e tolerância a detergentes comerciais fazem desta lipase um aditivo potencial para formulação de detergentes.
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Industrial enzymes are produced by various organisms using a wide variety of substrates. The present study was carried out to investigate the production of protease by Aspergillus niger using fish scale as the growth media. Fish scale, the chief waste material of fish processing industries, was enzymatically hydrolysed by protease produced by A. niger. Two mutated strains of A. niger, namely, AB30 and AB60 were also used apart from the wild type A. niger strain. Trichloroacetic acid buffer was used to maintain the optimum moisture content of the substrate. No separate nutrient sources apart from the fish scales were used. This proves the cost effectiveness and efficiency of this process. The enzyme activity was analyzed from the 5 th day and it was further analyzed for three consecutive days and the results were plotted and analyzed.
Enzymes are vitally important for the existence of life. Civilizations have used enzymes for thousands of years without understanding what they were or how they work. Over the past several generations, science has unlocked the mystery of enzymes and has applied knowledge to make better use of it in an ever growing number of applications. Enzymes play crucial role in producing the food we eat, the clothes we wear and even in producing fuel for our automobiles. They are important in combating environmental pollution. Leather processing is one of the important industries closely related to everyday life. Proteases execute a large variety of functions and have important biotechnological applications. They represent one of the largest groups of industrial enzymes and find application in detergents industry, leather industry, food industry, pharmaceutical industry and bioremediation processes (Anwar and Saleemuddin, 1998; Gupta et al. 2002). Probably the largest application of proteases is in laundry detergents, where they help removing protein based stains from clothes (Banerjee et al. 1999). In textile industry, proteases may also be used to remove the stiff and dull gum layer from the raw silk fibre to achieve improved luster and softness. They can modify the surface of wool and silk fibres to provide new and unique finish. Proteases have been used in the hide-dehairing process, where dehairing is carried out at pH values between 8 and 10.