Sesame oil cake an inexpensive substrate for neutral protease production by penicillium chrysogenum ncim 737 in solid-state fermentation

B. Nagamani et al., IJSID, 2012, 2 (5), 436-447

ISSN:2249-5347
IJSID
International Journal of Science Innovations and Discoveries An International peer
Review Journal for Science

Research Article Available online through www.ijsidonline.info

SESAME OIL CAKE-AN INEXPENSIVE SUBSTRATE FOR NEUTRAL PROTEASE PRODUCTION BY
PENICILLIUM CHRYSOGENUM NCIM 737 IN SOLID-STATE FERMENTATION

Center for Biotechnology, Dept of Chemical Engineering, Andhra University, AP, India
B. Nagamani, M.V.V.Chandana Lakshmi*, V.Sridevi and P.Rajani

Neutral protease production under solid-state fermentation was carried out by
Received: 03-07-2012 ABSTRACT

using Penicillium chrysogenum NCIM 737. Among the six (green gram husk, black gram
Accepted: 18-10-2012

husk, rice bran, coconut oil cake, sesame oil cake and paddy straw + rice bran (7:3))
agro-industrial waste materials evaluated, sesame oil cake supported maximum protease
*Corresponding Author

production. The physiological parameters such as fermentation time, fermentation
temperature, pH, inoculum age, initial moisture content and the nutritional parameters
namely carbon, organic and inorganic nitrogen sources were optimized for the production
of protease. At 7 days of fermentation, 25°C, pH 7, 7-days old culture and at 45% initial
moisture content, protease activity of 172.5 U/gds was obtained. Further the activity was
raised to 197.5 U/gds by supplementing the substrate media with sucrose (1% w/w),
peptone (1% w/w) and ammonium chloride (1% w/w).
Address:
Name: Keywords: Solid-state fermentation, Sesame oil cake, Penicillium chrysogenum NCIM 737,
MVV. Chandana Lakshmi
INTRODUCTION
Place: Protease, Physiological parameters, Nutritional parameters.
Vishakapatnam, AP, India
E-mail:
mahantilakshmi@yahoo.com

INTRODUCTION

International Journal of Science Innovations and Discoveries, Volume 2, Issue 5, September-October 2012

436


Extra cellular enzymes are usually capable of digesting insoluble nutrient materials such as cellulose, protein, starch,
INTRODUCTION

and the digested products are transported into the cell where they are used as nutrients for growth. Some extra cellular
enzymes used in the food, dairy, pharmaceutical, textile industries, etc., are produced in large amounts by microbial synthesis1.
Enzymes will probably play a key role for environmental friendly cleanup processes because of biodegradability and efficient
processing in leather industry2. Proteolytic enzymes play an important role in the metabolism of almost all organisms (plants,
animals, fungi, bacteria and viruses). Investigation of proteases is a central issue in enzymology due to both their immense
physiological importance and wide application in research and economical activities 3. Commercial application of microbial
proteases is attractive due to the relative ease of large-scale production as compared to proteases from plants and animals4.
Most of the enzymes market is related to hydrolytic type of enzymes such as proteases, lipases and the cellulases. Although use
of enzymes has many advantages, the competitiveness of the enzymes compared to the chemicals is limited by their higher
production costs. Hence, the use of a less expensive material as substrate is an interesting option 5. Traditionally, synthetic
substrates were used for fermentations, which are now being largely replaced by agro- and agro-industrial by-products. These
not only provide a natural substrate for fungal growth and fermentation but they result in improved value of these agro-
industrial residues6.
Microbial proteases, which play a specific catalytic role in the hydrolysis of proteins, are indeed one of the most
important groups of industrial enzymes and account for about 60% of the total worldwide enzyme sales 7. Fungal proteases
offer a distinct advantage over the bacterial enzymes in terms of ease of downstream processing 8. Fungal neutral proteases are
the most important component of commercial fungal protease preparations, which have applications in baking, food
processing, animal feeds and pharmaceutical industries 9, 10. Neutral proteases usually have low thermostability but there are
reports of heat resistant neutral proteases, which can hydrolyze casein at fastest rate at 60 - 65°C11.
Species of Penicillium are also the producers of neutral proteases9. Though most of filamentous fungi are capable of
producing proteolytic enzymes, a demand for strain selection and fermentation media is an essential target in biotech
industry2. Protease producing strains can grow effectively in medium containing protein hydrolysates. Peptone is generally
used in culture media because of its rich amino acid and low-molecular weight peptide content10. Productions of cell and
enzymes in Solid-State Fermentation (SSF) were remarkably influenced by the water content, so water is the limiting factor for
fungal growth in SSF12, 13. SSF, generally known as the "bran process," was almost universally employed for the production of
fungal enzymes14. In recent developments, the organisms used in SSF produce high yields of pure enzymes, which are much
more efficiently produced than in submerged fermentations. Fungi play a key role in SSF, for their hyphal development allows
them to effectively colonise and penetrate the solid substrate12.
The present study was undertaken for the production of neutral protease under SSF by P. chrysogenum NCIM 737
using sesame oil cake as substrate, and to determine the effect of various physiological and nutritional parameters to enhance
the activity of protease.
MATERIALS AND METHODS

The substrates used in this study namely Green gram husk, Black gram husk, Rice bran, Coconut oil cake,
Substrate

Sesame oil cake (SOC) and Paddy straw + Rice bran (7:3) were obtained from local grocery shop in Visakhapatnam.


437


The organisms used in the present study namely P. chrysogenum NCIM 737, Rhizopus oligosporus NCIM 1215 and
Microorganism and maintenance of culture

Acremonium chrysogenum NCIM 893 were obtained from National Collection of Industrial Microorganisms (NCIM), Pune,
India. The cultures were maintained on potato dextrose agar slants and were sub-cultured every month.

The inoculum was prepared by dispersing the spores from a week-old fungal slant culture in 0.1 % Tween-80 solution
Inoculum preparation

with a sterile inoculation loop.

Five grams of each substrate was taken in 250 mL Erlenmeyer flask separately were moistened with salt solution
Solid-State Fermentation

[composition (%w/v) (g/100mL): ammonium nitrate 0.5, potassium dihydrogen orthophosphate 0.2, sodium chloride 0.1 and
magnesium sulfate 0.1] and sterilized at 121.5°C for 15 min, cooled and then inoculated with 1 mL of fungal spore suspension
and incubated at 25°C for 7 days.

A solution of Tween-80 (0.1%) was added to 100 mL distilled water. 25 mL of water was added to 5 g of fermented
Extraction of crude enzyme

substrate and the substrate was homogenized on a rotary shaker at 180 rpm for 1h and then filtered. The solids were removed
by centrifuging the homogenate at 8000 x g at 4ºC for 15 min and the resultant clear supernatant was used for analytical
studies.

To 200 µL of crude enzyme extract, 500 µL of casein (1%) and 300 µL of 0.2 mol/L phosphate buffer (pH 7.0) were
Assay for neutral protease

added. The reaction mixture was incubated at 60°C for 10 min and arrested by the addition of 1mL of 10% trichloro acetic
acid16. The reaction mixture was centrifuged at 8000 x g at 40C for 15 min and to the supernatant, 5 mL of 0.4 mol/L Na 2CO3,
1mL of 3-fold diluted Folin and Ciocalteau’s phenol reagent were added. The resulting solution was incubated at room
temperature for 30 min and the absorbance of the blue color developed was read at 660 nm and its concentration was
determined using tyrosine standard curve. One unit of enzyme activity was defined as the amount of enzyme that liberated
one microgram of tyrosine from substrate (casein) per minute under assay conditions.
Standard graph for tyrosine

To a series of test tubes, 0.1mL, 0.2mL, 0.4mL, 0.6mL, 0.8mL, 1mL of standard solution of tyrosine (100 µg/mL) was
Procedure

taken and water is added to each test tube to make the solution up to 1mL. Each test tube contains 10, 20, 40, 60, 80, 100
µg/mL of tyrosine. To each test tube 5 mL of 0.5 M Na 2CO3 and 1mL of 3-fold diluted Folin and Ciocalteau’s Phenol reagent
were added and incubated for 30 min. The optical density of above solutions was measured at 660 nm (Fig. 1). Blank was
prepared with 1mL of water instead of tyrosine solution.

Seven different substrates like green gram husk, black gram husk, rice bran, coconut oil cake, sesame oil cake, paddy
Screening of substrates and fungal species

straw rice bran (7:3) were screened using three different fungal species namely Penicillium chrysogenum NCIM 737, Rhizopus
oligosporus NCIM 1215 and Acremonium chrysogenum NCIM 893 for neutral protease production using SSF.


438


OPTIMIZATION

The protocol adopted for the optimization of process parameters was to evaluate the effect of an individual parameter
Optimization of physiological parameters

at a time and to incorporate it at the standard level before optimizing the next parameter.

The production profile of neutral protease was studied by conducting the fermentation at different time intervals 1, 2, 3, 4, 5,
Optimization of fermentation time

6, 7, 8, 9, 10 days.

The inoculated substrates were incubated at different temperatures ranging from 20 to 45°C, to determine the optimum
Optimization of fermentation temperature

fermentation temperature for neutral protease production.

Optimum pH for neutral protease production was determined by conducting the fermentation at different pH 4, 5, 6, 7, 8, 9 and
Optimization of medium pH

10.

Different inoculum ages of 1-10 days were varied to determine the maximum production of neutral protease.
Optimization of inoculum age

Optimum initial moisture content for neutral protease production was determined by adjusting the initial moisture
Optimization of initial moisture content

content of the fermentation substrate to varying levels of 30, 35, 40, 45, 50 and 55%.
EFFECT OF NEUTIONAL PARAMETERS

Influence of various carbon supplements on enzyme production was studied by adding different sugars namely
Effect of carbon supplements

xylose, maltose, galactose, sucrose and lactose at 1 % (w/w) to the fermentation media.

Various organic nitrogen supplements - peptone, beef extract, yeast extract and malt extract at a concentration of 1 %
Effect of organic nitrogen supplements

(w/w) were added to the fermentation media to study its effect on protease production.

Different sources of inorganic nitrogen - KNO3, NH4Cl, NH4NO3 and (NH4)2SO4 at 1 % (w/w) were added to the
Effect of inorganic nitrogen supplements

fermentation medium to study its effect on enzyme production.

The pH optimum of the neutral protease enzyme was determined by using buffer solutions of different pH (Phosphate
Characterization of the extracted enzyme

buffer 7.0, Acetate buffer 5.0, Glycine-NaOH buffer 10.5) for enzyme assay. The buffers used were of the concentration 0.2
mol/L.
RESULTS AND DISCUSSION

The three fungal species were inoculated individually in the six agro-industrial wastes. The results in the present
Screening of microorganisms and substrates

study indicated that protease production varied with the type of agro-waste as shown in Table 1. The maximum activity of
130.0 U/gds was obtained, when P.chrysogenum NCIM 737 was inoculated in the substrate, sesame oilcake. This could be
attributed to solid materials dual role supply of nutrients to the microbial culture and anchorage for the growing cells.

439


Table 1: Screening of microorganisms and substrates for the protease production
Substrate
protease activity(U/gds) protease activity(U/gds) protease activity(U/gds)
P.chrysogenum R.oligosporous A. chrysogenum

Green gram husk 50.00 47.50 5.00
Black gram husk 17.50 7.50 10.00
Rice bran 50.00 20.00 82.50
Coconut Oil cake 17.50 10.00 11.25
Sesame oil cake 130.00 102.50 2.50
Paddy straw + Rice bran(7:3) 10.00 15.00 5.00
P.chrysogenum NCIM 737 proved to be the best strain for neutral protease production on sesame oilcake substrate
giving maximum enzyme activity. Hence, this Penicillium strain was selected to optimize the physiological and nutritional
parameters to enhance the enzyme production under SSF.

1.2

1.0
optical density at 660 nm

0.8

0.6

0.4

0.2

0.0
0 20 40 60 80 100

Figure 1. Standard graph for tyrosine
concentration of tyrosine (micro gram/ml)

The enzyme production was gradually increased with the passage of time and highest enzyme activity of 135.0 U/gds
Effect of fermentation time

was obtained after 7 days of incubation as shown in Fig.2. The subsequent decrease in the enzyme may be due to the
inactivation of the enzyme by other constituent proteases, the reduced availability of nutrients and production of toxic
metabolites11. Tremacoldi and Carmona, 2005 reported that the highest protease activity was obtained by Aspergillus clavatus
after 6 days of incubation for culture medium containing glucose and casein at 1% (w/v) as substrates 15.

The enzyme production was carried out by P.chrysogenum NCIM 737 at 20-45°C temperature range. Maximum
Effect of fermentation temperature

activity of protease, 147.5 U/gds was obtained at a temperature of 25°C as shown in Fig. 3. Further increase in temperature,
reduced the enzyme production. The reduction in enzyme activity may be due to the denaturation of the enzyme by losing its
catalytic properties at high temperature due to stretching breaking of weak hydrogen bonds with in the enzyme structure 11. In
earlier reports, Pushpa and Madhava Naidu, 2010 reported that the maximum production of protease from coffee by-products
using Aspergillus oryzae was obtained at temperature 30°C16.


440


180

Protease activity (U/gds)
140

100

60

20

-20 0 2 4 6 8 10 12
Fermentation time (days)

Figure 2. Effect of fermentation time on the production of neutral protease by P.chrysogenum NCIM 737

180

140

100

60

20

-20 15 20 25 30 35 40 45 50
Fermentation temperature

Figure 3. Optimization of fermentation temperature for neutral protease production by P.chrysogenum NCIM 737

Protease production by microbial strains depends on the extra cellular pH because culture pH strongly influences
Effect of pH

many enzymatic processes and transport of various components across the cell membranes, which in turn support cell growth
and product production17,18. The enzyme synthesis was increased with increase of medium pH towards neutrality with a
maximum activity of 162.5 U/gds as shown in Fig. 4. Similar results were also reported by Paranthaman et al., 2009 that the
maximum production of neutral protease from rice mill waste using Aspergillus niger was obtained at pH 7.017.

The effect of inoculum age on protease production was studied by conducting the fermentation with different
Effect of inoculum age

inoculum ages, 1-10 days. It was observed that 7-days old culture gave maximum production of protease, 165.0 U/gds as
shown in Fig.5. Ikasari and Mitchell, 1994 reported that the 5-day old inoculum gave maximum protease yield with Rhizopus
oligosporus ACM 145F19.

441


200

160

120

80

40

0
2 4 6 8 10 12
pH

Figure 4. Effect of pH on the production of protease production by P.chrysogenum NCIM 737

200

160

120

80

40

0
0 2 4 6 8 10 12
Inoculum age (days)

Figure 5. Effect of inoculum age on the production of neutral protease by P.chrysogenum NCIM 737.

Initial moisture content is a crucial factor affecting the formation of products through SSF. The optimum initial moisture
Effect of initial moisture content

content for neutral protease production was determined by adjusting the initial moisture content of the fermentation
substrate to varying levels of 30, 35, 40, 45, 50 & 55 %. From Fig. 6 it was observed that moisture level of 45 % was found to
be optimum for neutral protease production (172.5U/gds). Sumantha et al., 2006 reported that, the moisture content of 44.4
% facilitated neutral protease production by Rhizopus microsporus NRRL 3671, on rice bran20.


442


200

160

120

80

40

0
25 30 35 40 45 50 55 60
Initial moisture content (%)

Figure 6. Effect of initial moisture content on the production of neutral protease by P.chrysogenum NCIM 737

Various carbon supplements namely xylose, maltose, galactose, sucrose and lactose of 1 % (w/w) were studied
Effect of carbon supplements

individually. From Fig. 7 it was observed that all the carbon supplements to the substrate show influence on the enzyme
production. Among the 5 different carbon sources, it was observed that sucrose showed the highest enzyme activity of
181.25U/gds. Sumantha et al., 2006 reported that sucrose was the best carbon source in the carbohydrate deficient substrates
and sucrose enhanced the protease production by Rhizopus microsporus NRRL 367120.

200
180

160
140
120
100
80
60
40
20
0
xylose maltose galactose sucrose lactose
Carbon sources (1 % (w/w))

Figure 7. Effect of carbon sources on the production of neutral protease by P.chrysogenum NCIM 737.

Different organic nitrogen supplements like peptone, beef extract, yeast extract and malt extract at a concentration of
Effect of organic nitrogen supplements

1 % (w/w) were examined for the maximum protease activity. Among the 4 different organic nitrogen supplements, peptone
enhanced the production of protease i.e., maximum activity of 190.0 U/gds was obtained as shown in Fig. 8. According to


443


Kalaiarasi and Sunitha, 2009 also reported that peptone was the best organic nitrogen supplement for the maximum
production of protease21.

195
190
Protease activity (U/gds) 185
180
175
170
165
160
155
150
145
peptone beef extract yeast extract malt extract
Organic nitrogen sources (1% (w/w))

Figure 8. Effect of organic nitrogen sources on the production of neutral protease by P.chrysogenum NCIM 737

Various inorganic nitrogen supplements namely KNO3, NH4Cl, NH4NO3, and (NH4)2SO4 at a concentration of 1 % (w/w)
Effect of inorganic nitrogen supplements

were studied to enhance the enzyme activity. From Fig. 9 it was observed that among the 4 different inorganic nitrogen
supplements maximum protease activity of 197.5 U/gds was obtained using NH 4Cl. In the work of Sindhu et al., 2009,
ammonium nitrate (0.5%) was the best inorganic nitrogen source for the maximum production of protease by Penicillium
godlewskii SBSS 25 22.

200
195

190
185
180
175
170
165
160
155
150
KNO3 NH4Cl NH4NO3 (NH4)2SO4
Inorganic nitorgen source (1 %(w/w))

Figure 9. Effect of inorganic nitrogen sources on the production of neutral protease by P.chrysogenum NCIM 737


444


Enzyme characterization

The enzyme showed the maximum specific activity at pH 7.0 (Fig. 10) indicating the instability of enzyme at other pH
pH

values. Sumantha et al., 2006 reported that the neutral protease from Rhizopus oligosporus NRRL 3671 also has optimum pH
7.0.

200

160

120

80

40

0
4 5 6 7 8 9 10 11
pH

Figure 10. pH curve of neutral protease of P.chrysogenum NCIM 737.

The present study focused on the use of P. chrysogenum NCIM 737 to convert agro-industrial by-products to a value-
CONCLUSION

added product, an enzyme-protease. Neutral protease production under SSF was carried out by using P. chrysogenum NCIM
737. At the optimum conditions of fermentation time 7 days, temperature 25 oC, pH 7.0, inoculum age 7 days, initial moisture
content 45% the protease activity found was 172.5 U/gds. In addition to physiological parameters, the chemical parameters
namely, carbon (sucrose), organic nitrogen source (peptone) and inorganic nitrogen source (ammonium chloride) at a
concentration of 1% w/w enhanced the protease activity to 197.5 U/gds. Proteases have found a wide range of applications in
various industries such as food, pharmaceuticals, detergents etc. From the results, it could be inferred that neutral protease
produced through SSF of the sesame oil cake by P.chrysogenum NCIM 737 could possibly find useful application in food
industries.

1. Vania Sousa Andrade, Leonie Asfora Sarubbo, Kasutaka Fukushima, Makoto Miyaji, Kazuko Nishimura, Galba Maria de
REFERENCES

Campos-Takaki, “Production of extracellular protease By mucor circinelloides using D-glucose as carbon source /
substrate”, Braz. J. Microbiol. 2002, 33, 106-110.
2. Chellapandi, P. “Production and Preliminary Characterization of Alkaline Protease from Aspergillus flavus and Aspergillus
terreus”, E-J. Chem., 2010, 7(2), 479-482.


445


3. Siddalingeshwara, K.G., Uday, J., Huchesh, C.H., Puttaraju , H.P., Karthic, J.,Sudipta, K.M., Pramod, T., Vishwanatha, T.
“Screening and characterization of protease from Bacillus sp”, Int. J. Appl. Biol. Phar.Tech. 2010, 1, 575-581.
4. Vamsi Krishna, K., Mayank Gupta, Nikhil Gupta, Hipal Gaudani, Soham Trivedi, Prasad Patil, Girish Gupta, Yogesh
Khairnar, Amol Borasate, Dharmendra Mishra, “Optimization of growth and production of protease by Penicillium
species using submerged fermentation”, Int. J. Microbiol. Res. 2009,1,14-18.
5. Drouin, M., Lai, C. K., Tyagi, R.D., Surampalli, R.Y. “Bacillus licheniformis proteases as high value added products from
fermentation of wastewater sludge: pre-treatment of sludge to increase the performance of the process”, Water Science
& Technology 2007,599-605.
6. Alagarsamy Sumantha, Chandran Sandhya, George Szakacs, Carlos R. Soccol Ashok Pandey, “Production and Partial
Purification of a Neutral Metalloprotease by Fungal Mixed Substrate Fermentation”,Food Technol. Biotechnol. 2005, 43(4),
313–319.
7. Wu, T.Y., Mohammad, A.W., Jahim, J. Md., Anuar, N. “Investigations on protease production by a wild-type Aspergillus
terreus strain using diluted retentate of pre-filtered palm oil mill effluent (POME) as substrate”,Enzy. Microbial Tech.,
2006, 39, 1223-1229.
8. Mohamed Hajji, Ahmed Rebai, Neji Gharsallah, Moncef Nasri, “Optimization of alkaline protease production by Aspergillus
clavatus ES1 in Mirabilis jalapa tuber powder using statistical experimental design”, Appl. Microbiol. Biotech. 2008, 79,
915-923.
9. Alagarsamy Sumantha, Christian Larroche, Ashok Pandey, “Microbiology and Industrial Biotechnology of Food-Grade
Proteases: A Perspective”, Food Technol. Biotechnol. 2006, 44(2), 211-220.
10. Barnali Ray Basu, Ajit K. Banik, Manas Das, “Production and characterization of extracellular protease of mutant
Aspergillus niger AB100 grown on fish scale”, World J. Microbiol. Biotechnol. 2008, 24, 449-455.
11. Ikram-ul-haq, Hamid mukhtar, Hina umber, “Production of Protease by Penicillium chrysogenum Through Optimization of
Environmental Conditions”, J. Agr. Soc. Sci. 2006, 2(1), 23-25.
12. Rob te Biesebeke, George Ruijter, Yovita S.P. Rahardjo, Marisca J.Hoogschagen, Margreet Heerikhuisen, Ana Levin,
Kenneth G.A. van Driel, Maarten A.I. Schutyser, Jan Dijksterhuis, Yang Zhu, Frans J. Weber, Willem M. de Vos, Kees A.M.J.J.
van den Hondel, Arjen Rinzema, Peter J. Punt, “Aspergillus oryzae in solid-state and submerged fermentations Progress
report on a multi-disciplinary project”,FEMS Yeast Res. 2002, 2, 245-248.
13. 13. Hideki Narahara, Yosuke Koyama, Toshiomi Yoshida, Poonsuk Atthasampunna, Hisaharu Taguchi, “Control of water
content in a solid state culture of Aspergillusoryzae”, J. Fermen. Tech. 1984, 62(5), 453-459.
14. 14. Hwa L. Wang, Janet B. vespa, Hesseltine, C. W., “Acid Protease Production by Fungi Used in Soybean Food
Fermentation”, Appl. Microbiol. 1974, 27(5), 906-911.
Celia R. Tremacoldi and Eleonora Cano Carmona, “Production of extracellular alkaline proteases by Aspergillus clavatus”,
World J. Microbiol. Biotech., 2005, 21,169-172.
15.

Pushpa S.Murthy, and Madhava Naidu, M., “Protease production by Aspergillus oryzae in solid state fermentation
utilizing Coffee byproducts”, World Appl. Sci. J., 2010, 8(2), 199-205.
16.

17. Paranthaman, R., Alagusundaram, K. and Indhumathi, “Production of protease from rice mill wastes by Aspergillus niger
in Solid State Fermentation”, World J. of Agri. Sci., 2009, 5(3), 308-312.


446


18. Prakasham, R.S., Subba Rao, Ch. and Sarma, P.N., Green gram husk—an inexpensive substrate for alkaline protease
production by Bacillus sp. in solid-state fermentation. Biores. Tech., 2006, 97, 1449–1454.
19. Ikasari, L., Mitchell, D.A. “Protease production by Rhizopus oligosporus in solid state fermentation”, World J. Microbiol.
Biotechnol. 1994, 10, 320-324.
20. Alagarsamy Sumantha, Paul Deepa, Chandran Sandhya, George Szakacs, Carlos Ricardo Soccol, Ashok Pandey, “Rice Bran
as a Substrate for Proteolytic Enzyme Production”, Braz. Arch. Biol. Technol. 2006, 49(5), 843-851.
21. Kalaiarasi, K., Sunitha, P. U. “Optimization of alkaline protease production from Pseudomonas fluorescens isolated from
meat waste contaminated soil”, African JBiotechnol. 2009, 8(24), 7035-7041.
22. Sindhu, R., Suprabha, G. N. and Shashidhar, S., “Optimization of process parameters for the production of alkaline
protease from Penicillium godlewskii SBSS 25 and its application in detergent industry”, African Journal of
MicrobiologyResearch, 2009, 3(2), pp. 515-522.


447

Sesame oil cake an inexpensive substrate for neutral protease production by penicillium chrysogenum ncim 737 in solid-state fermentation

Recommended

Recommended

More Related Content

What's hot

What's hot (20)

Similar to Sesame oil cake an inexpensive substrate for neutral protease production by penicillium chrysogenum ncim 737 in solid-state fermentation

Similar to Sesame oil cake an inexpensive substrate for neutral protease production by penicillium chrysogenum ncim 737 in solid-state fermentation (20)

More from ijsidonlineinfo

More from ijsidonlineinfo (13)

Recently uploaded

Recently uploaded (20)

Sesame oil cake an inexpensive substrate for neutral protease production by penicillium chrysogenum ncim 737 in solid-state fermentation