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Frontiers in Biology

ISSN 1674-7984

ISSN 1674-7992(Online)

CN 11-5892/Q

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Front. Biol.    2018, Vol. 13 Issue (2) : 123-129    https://doi.org/10.1007/s11515-018-1485-3
RESEARCH ARTICLE
Media optimization for extracellular amylase production by Pseudomonas balearica vitps19 using response surface methodology
Moni Philip Jacob Kizhakedathil, Subathra Devi Chandrasekaran()
Department of Biotechnology, School of Biosciences and Technology, VIT University, Vellore – 632014, Tamil Nadu, India
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Abstract

BACKGROUND: In this study, we optimized the process for enhancing amylase production from Pseudomonas balearica VITPS19 isolated from agricultural lands in Kolathur, India.

METHODS: Process optimization for enhancing amylase production from the isolate was carried out by Response Surface Methodology (RSM) with optimized chemical and physical sources using Design expert v.7.0. A central composite design was used to evaluate the interaction between parameters. Interaction between four factors – maltose (C-source), malt extract (N-source), pH, and CaCl2 was studied.

RESULTS: The factors pH and CaCl2 concentration were found to affect amylase production. Validation of the experiment showed a nearly twofold increase in alpha amylase production.

CONCLUSION: Amylase production was thus optimized and increased yield was achieved.
Keywords Pseudomonas balearica VITPS19      alpha amylase      optimization      response surface methodology      central composite design      pH     
Corresponding Author(s): Subathra Devi Chandrasekaran   
Online First Date: 15 March 2018    Issue Date: 28 May 2018
 Cite this article:   
Moni Philip Jacob Kizhakedathil,Subathra Devi Chandrasekaran. Media optimization for extracellular amylase production by Pseudomonas balearica vitps19 using response surface methodology[J]. Front. Biol., 2018, 13(2): 123-129.
 URL:  
https://academic.hep.com.cn/fib/EN/10.1007/s11515-018-1485-3
https://academic.hep.com.cn/fib/EN/Y2018/V13/I2/123
Factors Independent variables Coded Levels
–a (-2) –1 0 + 1 + a ( + 2)
A Maltose % 0 0.5 1 1.5 2
B Malt extract % 0 0.25 0.50 0.75 1
C CaCl2% 0 0.05 0.10 0.15 0.2
D pH 6 6.25 6.5 6.75 7
Tab.1  Range of values for the response surface method- CCD
Runs number Coded value order A B C D Enzyme activity
(U)		 Predicted response
(U)
8 1 1.5 0.75 0.15 6.25 0.52 0.16
5 2 0.5 0.25 0.15 6.25 0.76 0.926
28 3 1 0.5 0.1 6.5 1.90 1.90
19 4 1 0 0.5 6.5 1.77 2.108
26 5 1 0.5 0.1 6.5 1.90 1.90
30 6 1 0.5 0.1 6.5 1.90 1.90
12 7 1.5 0.75 0.05 6.75 2.10 1.186
22 8 1 0.5 0.2 6.5 0.49 0.46
11 9 0.5 0.75 0.05 6.75 2.42 1.996
27 10 1 0.5 0.1 6.5 1.90 1.90
15 11 0.5 0.5 0.15 6.75 2.48 1.958
10 12 1.5 0.25 0.05 6.75 1.92 1.986
13 13 0.5 0.25 0.15 6.75 1.94 1.79
17 14 0 0.5 0.1 6.5 1.92 2.416
23 15 1 0.5 0.1 6 0.30 0.12
24 16 1 0.5 0.1 7 0.59 1.44
4 17 1.5 0.75 0.05 6.25 0.55 0.73
7 18 0.5 0.75 0.15 6.25 0.80 0.766
9 19 0.5 0.25 0.05 6.75 1.96 1.056
1 20 0.5 0.25 0.05 6.25 1.89 1.292
29 21 1 0.5 0.1 6.5 1.90 1.90
21 22 1 0.5 0 6.5 0.42 1.1
16 23 1.5 0.75 0.15 6.75 0.28 0.912
25 24 1 0.5 0.1 6.5 1.90 1.90
3 25 0.5 0.75 0.05 6.25 1.08 1.244
14 26 1.5 0.25 0.15 6.75 2.73 1.824
18 27 2 0.5 0.1 6.5 1.73 1.936
2 28 1.5 0.25 0.05 6.25 2.05 1.856
6 29 1.5 0.25 0.15 6.25 0.80 1.256
20 30 1 1 0.1 6.5 0.78 1.148
Tab.2  Response of the experimental design by RSM
Source Sum of squares df Mean square F value p-value Prob>F
Model 11.01 14 0.79 2.40 0.0479 Significant
A-A 0.32 1 0.32 0.99 0.3345
B-B 1.39 1 1.39 4.23 0.0563
C-C 0.58 1 0.58 1.77 0.2021
D-D 2.62 1 2.62 8.00 0.0121 Significant
AB 1.13 1 1.13 3.45 0.0819
AC 0.056 1 0.056 0.17 0.6837
AD 0.089 1 0.089 0.27 0.6103
BC 0.013 1 0.013 0.039 0.8467
BD 0.11 1 0.11 0.33 0.5751
CD 0.18 1 0.18 0.54 0.4711
A2 0.14 1 0.14 0.42 0.5279
B2 0.13 1 0.13 0.41 0.5316
C2 2.32 1 2.32 7.07 0.0171 Significant
D2 2.18 1 2.18 6.65 0.0202 Significant
Residual 5.24 16 0.33
Lack of Fit 5.24 10 0.52
Pure Error 0.000 6 0.000
Cor total 16.25 30
Tab.3  Analysis of variance (ANOVA) for the quadratic model of amylase production obtained from the experimental results
Std. Dev. 0.57 R-Squared 0.6774
Mean 1.47 Adj R-Squared 0.3951
C.V. % 39.03 Pred R-Squared -0.8581
PRESS 30.19 deq Precision 5.972
Tab.4  R-Squared, Adj R-Squared, Pred R-Squared, and Adeq Precision value of the model
Fig.1  (A) Three dimensional curves showing the effects of factors C (CaCl2) and D (pH). (B) Contour plots showing the effects of factors C (CaCl2) and D (pH).
Fig.2  Perturbation graph showing the effect of all independent variables on amylase production.
Fig.3  Plot between expected normal values versus residuals.
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