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Frontiers of Chemical Science and Engineering

ISSN 2095-0179

ISSN 2095-0187(Online)

CN 11-5981/TQ

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Front Chem Sci Eng    2011, Vol. 5 Issue (4) : 463-470    https://doi.org/10.1007/s11705-011-1112-1
RESEARCH ARTICLE
Application of statistical design for the production of inulinase by streptomyces sp. using pressmud
M. DILIPKUMAR1(), M. RAJASIMMAN1, N. RAJAMOHAN2
1. Department of Chemical Engineering, Annamalai University, Annamalainagar-608002, Tamilnadu, India; 2. Department of Chemical Engineering, Sohar University, Sohar, Sultanate of Oman
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Abstract

A Plackett-Burman design was employed for screening 18 nutrient components for the production of inulinase using streptomyces sp. and pressmud as the substrate via solid-state fermentation (SSF). From the experiments, three nutrients viz. yeast extract, FeSO4·7H2O, and NH4NO3 were found to be the most significant components. Hence these three components were selected and optimized using Response Surface Methodology (RSM). The optimum conditions are: yeast extract 0.00274 g/gds, FeSO4·7H2O 0.00011 g/gds and NH4NO3 0.00772 g/gds. The effect of the substrate concentration and initial moisture content were also studied. A substrate concentration of 12 g and an initial moisture content of 65% are optimum for the maximum production of inulinase (89 U/gds).
Keywords inulinase      pressmud      Response Surface Methodology (RSM)      streptomyces sp     
Corresponding Author(s): DILIPKUMAR M.,Email:mdilip_kumar@yahoo.co.in   
Issue Date: 05 December 2011
 Cite this article:   
M. DILIPKUMAR,M. RAJASIMMAN,N. RAJAMOHAN. Application of statistical design for the production of inulinase by streptomyces sp. using pressmud[J]. Front Chem Sci Eng, 2011, 5(4): 463-470.
 URL:  
https://academic.hep.com.cn/fcse/EN/10.1007/s11705-011-1112-1
https://academic.hep.com.cn/fcse/EN/Y2011/V5/I4/463
NutrientsComposition
Organic carbon20%-25%
Nitrogen0.90%-1.25%
Phosphorus2.50%-3.00%
Potassium1.00%-1.50%
Calcium11.00%
Magnesium1.65%
Sulfur0.23%
Copper52 ppm
Zinc69 ppm
Manganese898 ppm
Iron2000 ppm
Moisture70%
Tab.1  Composition of Pressmud
VariableLevels/g·((10 gds)-1)
Nutrient codeNutrientLow ( - 1)High ( + 1)
AYeast extract0.010.05
BBeef extract0.050.15
CMnSO4·7H2O0.10.5
DK2HPO40.020.07
ESoya bean cake0.40.8
FMgSO4·7H2O0.0020.012
GNH4Cl0.010.03
HKCl0.0050.015
J(NH4)2HPO40.050.3
KNH4NO30.050.1
LZnSO4·7H2O0.10.5
M(NH4)2SO40.060.1
NCorn steep liquor0.40.8
OPeptone0.050.15
PDextrose0.10.3
QFeSO4·7H2O0.00050.002
RKH2PO40.10.6
SUrea0.10.3
Tab.2  Nutrient screening using a Plackett-Burman design
Run No.ABCDEFGHJKLMNOPQRSInulinase activity /(U·gds-1)
11- 11- 11111- 1- 111- 111- 1- 1- 156.74
21- 11111- 1- 111- 111- 1- 1- 1- 1161.51
3- 1- 1- 1- 11- 11- 11111- 1- 111- 1151.12
4- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 1- 140.53
5- 1- 1- 11- 11- 11111- 1- 111- 11131.44
611- 111- 1- 1- 1- 11- 11- 11111- 165.31
7- 1- 111- 111- 1- 1- 1- 11- 11- 111151.42
8- 11- 11111- 1- 111- 111- 1- 1- 1- 130.51
91- 1- 1- 1- 11- 11- 11111- 1- 111- 151.41
10- 11- 11- 11111- 1- 111- 111- 1- 162.30
111- 111- 1- 1- 1- 11- 11- 11111- 1- 171.81
12- 11111- 1- 111- 111- 1- 1- 1- 11- 131.30
13111- 1- 111- 111- 1- 1- 1- 11- 11- 162.61
1411- 1- 111- 111- 1- 1- 1- 11- 11- 1172.71
151- 1- 111- 111- 1- 1- 1- 11- 11- 11151.10
161111- 1- 111- 111- 1- 1- 1- 11- 1141.66
17- 1- 11- 11- 11111- 1- 111- 111- 121.14
1811- 1- 1- 1- 11- 11- 11111- 1- 11150.19
19- 111- 1- 1- 1- 11- 11- 11111- 1- 1120.78
20- 111- 111- 1- 1- 1- 11- 11- 1111173.43
Tab.3  Plackett-Burman experimental design matrix for screening of important variables for inulinase production
VariableCodeLevels /(g·(10 gds) -1)
- 1.68- 10+ 1+ 1.68
Yeast extractX10.010.020.030.040.05
FESO4·7H2OX20.00050.00080.00120.00160.002
NH4NO3X30.050.0620.0750.0870.1
Tab.4  Ranges of the independent variables used in RSM
Run No.X1X2X3Inulinase activity /(U·gds-1)
ExperimentalPredicted
101.6803333.84
21115149.09
31- 1- 15246.02
40007881.05
500- 1.684449.67
60008181.05
70007881.05
80008581.05
90008281.05
10- 1- 1- 16265.16
111- 115558.78
120008281.05
130- 1.6806259.40
1411- 13632.82
15- 1114148.23
16001.687163.57
17- 11- 14744.46
181.68004347.93
19- 1.68007063.30
20- 1- 116165.42
Tab.5  Central composite design (CCD) of factors in coded levels with enzyme activity as response
Fig.1  Pareto chart showing the effect of media components on inulinase activity
SourceCoefficient factorSum of squaresDFFP>F
Model81.05045100.12916.10<0.0001
X1-4.56975285.1918.100.0174
X2-7.59853788.51122.400.0008
X34.13041232.9916.620.0278
X1 × X21.8750028.1310.800.3924
X1 × X33.1250078.1312.220.1672
X2 × X30.875006.1310.170.6854
X1 × X1-8.991341165.07133.090.0002
X2 × X2-12.17332135.60160.66<0.0001
X3 × X3-8.637791075.24130.540.0003
Residual352.08108.780.0162
Lack of fit316.085
Pure error36.005
Cor total5452.2019
Tab.6  Analysis of Variance (ANOVA) for a response surface quadratic model for the production of inulinase
Fig.2  Predicted response versus actual response
Fig.3  3D plot showing the effect of yeast extract and FeSO·7HO on inulinase activity
Fig.4  3D plot showing the effect of yeast extract and NHNO on inulinase activity
Fig.5  3D plot showing the effect of FeSO·7HO and NHNO on inulinase activity
Fig.6  Effect of substrate concentration on inulinase production
Fig.7  Effect of initial moisture content on inulinase production
SubstrateMicroorganismActivity /(U·gds-1)Ref.
Wheat bran and rice huskCryptococcus aureus436.2[28]
Dry bagasseKluyveromyces marxianus19.4[29]
Wheat branStaphylococcus sp.107.64[30]
Kluyveromyces marxianus122.88
Sugarcane bagasseKluyveromyces marxianus250[31]
Wheat bran and Rice branPichia guilliermondii,291.0[32]
Pichia guilliermondii M-30455.9
PressmudStreptomyces sp.89This work
Tab.7  Ranges of the independent variables used in RSM
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