OPTIMIZATION OF CITRIC ACID PRODUCTION BY SOLID STATE FERMENTATION USING ARTIFICIAL NEURAL NETWORK (ANN)

RICCA RAHMAN BINTI NASARUDDIN 0611866

SUPERVISOR: DR. MOHAMMED SAEDI JAMI CO-SUPERVISOR: ASSOC. PROC. DR. ZAHANGIR ALAM

BIOTECHNOLOGY ENGINEERING DEPARTMENT KULLIYYAH OF ENGINEERING INTERNATIONAL ISLAMIC UNIVERSITY MALAYSIA

INTRODUCTION

BACKGROUND
2. THE IMPORTANCE OF USING SOLID STATE FERMENTATION (SSF)

1. THE IMPORTANCES OF CITRIC ACID PRODUCTION Low toxic acidulant

Great demand Growing at the rate

35% In 2004: 1. Beverages 50% 2. Food 1520% 3. Soaps and detergents 1517% 4. Pharmaceutical / cosmetics 79% 5. Industrial 68%

Used with agro-industrial residues

(E.g.: EFB) reduce environmental problem regarding disposal of solid waste Lower energy requirement Produce less wastewater Use low volume equipment which is lower in cost but can effectively produce high concentrated product

Worldwide citric acid production is around 1.4 million tones per year. At current prices the market is worth about $1.5 billion.

Fermentation process is always complex. Many factors influence Citric Acid production Must apply optimization to maximize the yield and profit Common method used RSM Lack usage of ANN Comparisons between RSM and ANN No available research used ANN to optimize Citric Acid production by solid state fermentation

PROBLEM STATEMENT

In this study.

1. Optimize medium composition

5. Compare ANNs results with RSM

ARTIFICIAL NEURAL NETWORK

2. Optimize process conditions

4. Validate the ANN model by running experiment

3. Predict the output yield of citric acid production

LITERATURE REVIEW

CITRIC ACID PRODUCTION

Oil palm empty fruit bunch fiber is a lignocellulosic waste from palm oil mills. 1. low cost, 2. renewable and 3. widespread sources of sugars.

1. The cultures of Aspergillus niger are fed on a sucrose or glucose-containing medium to produce citric acid.

2. In term of production type, the use of

submerged fermentation is still dominating.

3. But now, the solid-state fermentation is creating new possibilities for producers. 4. The use of agro-industrial residues such as EFB as support in solid-state fermentation is economically important and minimizes environmental problems.
Rotary drum SSF

OPTIMIZATION
To optimize the process parameters maximize product increase profit Several factors influence fermentation process: 1. Medium compositions (sucrose, trace elements, simulator)

2. pH
3. Temperature 4. Agitation 5. Aeration 6. Moisture content - SSF Methods used for optimization: 1. RSM

2. ANN
3. Others (Genetic algorithm, CCD, Factorial Design)

WHAT IS RSM ?
1. Collection of mathematical and statistical techniques useful for the modeling and analysis of problems in which a response of interest is influenced by several variables and the objective is to optimize this response. 2. Normally use quadratic relationship. 3. A first-order model with two independent variables can be expressed as:

y = 0 + 1x1 + 2x2 + e
4. The approximating function with two variables is called a second-order model: y = 0 + 1x1 + 2x2 + 11x112 + 22x222 + 12x12 + e 5. Rapid and efficiently used with small amount of data, 6. The primary limitation of RSM occurs when the approximation offered by the quadratic function is inadequate.

WHAT IS ANN ?
NEURONS Computational modeling system based on the neural structure of the brain. NEURAL NETWORKS
Dendrites Axon Cell Body

Consist of three layers: 1. Input layer(s) 2. Hidden layer(s) 3. Output layer(s)

Resembles human brain in two respects: 1. Learning from examples 2. Stores knowledge
Major Components: 1. Weighing factors 2. Summation function 3. Transfer function 4. Scaling and limiting 5. Output function 6. Error function and back propagated value 7. Learning function

Input Layer

Hidden Layer

Output

Layer

Applications in Engineering Field: 1. Complex and non-linear problems 2. Prediction 3. Classification 4. Data Association 5. Data conceptualization 6. Data filtering

Processing Element Major Components: 1. Weighing factors 2. Summation function 3. Transfer function 4. Scaling and limiting 5. Output function 6. Error function and back propagated value 7. Learning function

1. TYPES OF NEURAL NETWORKS

2. NEURAL NETWORK TOPOLOGIES

1. 2. Feed Forward Neural networks Recurrent neural network

3. TYPES OF LEARNING

SUPERVISED LEARNING:
with teacher Provide correct output for every input pattern by determine and adjusting the weight to produce answer as close as possible to known correct answer .

Single Perceptron ARTIFICIAL NEURAL NETWORK

UNSUPERVISED LEARNING: without teacher

Multilayer Perceptron (MLP)

4. FEED FORWARD BACKPROPAGATION NN

Target

Input

Neural Network, including connections (weights) between neurons

Compare Output

Does not require a correct answer associated with each input pattern in the training data set. It explores the underlying structure in the data and organize categories based on this data

HYBRID LEARNING:
Adjust Weight

Combine supervised and unsupervised learning

RESEARCH STUDIES
Karnik et al. ,2007 A comparative study of the ANN and RSM modeling approaches for predicting burr size in drilling. Minimum absolute percentage error for ANN prediction is within range 1% - 0.14% lower than RSM which is 12% - 4.8%. Desai et al. ,2008 Comparison of ANN and RSM in fermentation media optimization: Case study of fermentative production of scleroglucan Average percentage error of ANN is 6.5 lower than RSM which is 20 Correlation coefficient of validation data for ANN is 0.98 higher than RSM which is 0.89

Kandimalla et al. 1999 Optimization of a vehicle mixture for the transdermal delivery of melatonin using ANN and RSM ANN can easily handle more than 4 input variables but for RSM, a large no. of input variables lead to a polynomial with many coefficient that involves tedious computation

Bagci and Isik, 2006 Investigation of surface roughness in turning undirectional GFRP composites by using RSM and ANN It was found that the maximum test errors were 6.30% and 6.36% by comparing roughness (Ra) values predicted from ANN model with those predicted RSM.

METHODOLOGY..

PART 1: SOFTWARE APPLICATION

Materials and equipments Building ANN for optimization

Sensitivity Analysis
MATERIALS AND EQUIPMENTS: 1. Software = MATLAB Version 2008a MATLAB: 1. Command-line functions in M-file 2. Toolboxes : nntool, nftool, nntraintool, nprtool

nntool

nftool

nntraintool

nprtool

nctool

1. Data:

Table 2: Experimental design data using CCD with the experimental and predicted value (using RSM) of citric acid production for medum compositions optimization.
Run number 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Sucrose a (% w/w) 8 6 4 6 6 4 4 6 8 8 4 6 6 6 6 3 6 8 9 6 Mineral Solution b (% w/w) 12 2 12 8 8 4 4 8 4 4 12 8 8 14 8 8 8 12 8 8 Inoculum c (% w/w) 20 16 20 16 16 20 12 16 12 20 12 16 16 16 16 16 10 12 16 22 Citric acid yield (g/kg-dry EFB) Experimental 259.23 267.47 256.52 334.68 334.23 208.69 236.65 332.44 259.22 260.65 250.02 333.88 333.08 292.42 334.14 247.46 257.79 285.06 278.9 217.41 Predicted (RSM) 258.1 268.15 242.52 332.81 332.81 213.22 231.32 332.81 266.75 247.19 257.02 332.81 332.81 300.18 332.81 249.8 256.12 274.06 288.06 230.58

a : 1% (w/w) = 33.3 g/kg-EFB

b: 1% (v/w) = Zn, 3; Cu, 3.3; Mn, 13.3 and Mg, 166.7 mg/kg-EFB c: 1% (v/w) = 6.7 x 1010 spores/kg-EFB

Table 3: Experimental design using CCD with the experimental and predicted value (using RSM) of citric acid production for process conditions optimization
Citric acid production Run Initial pH, A1

Moisture content,
B1 78 (+1) 70 (0) 58 (-2) 70 (0) 70 (0) 78 (+1) 62 (-1) 70 (0) 70 (0) 70 (0) 70 (0) 82 (+2) 70 (0) 62 (-1) 70 (0) 78 (+1) 62 (-1) 70 (0)

Incubation temperature, C1 Predicted 28 (-1) 32 (0) 32 (0) 32 (0) 32 (0) 36 (+1) 36 (+1) 32 (0) 38 (+2) 32 (0) 26 (-2) 32 (0) 32 (0) 28 (-1) 32 (0) 36 (+1) 28 (-1) 32 (0) 229.68 368.16 195.36 368.16 368.16 256.88 233.46 315.96 263.75 368.16 220.80 233.97 368.16 161.96 285.19 194.35 180.53 368.16

(g/kg-EFB) Experimental 241.30 368.61 190.26 367.42 368.40 241.24 240.86 316.42 274.33 368.14 210.05 238.90 368.81 158.10 284.53 198.38 196.32 367.91

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

4 (-1) 6.5 (0) 6.5 (0) 6.5 (0) 6.5 (0) 4 (-1) 4 (-1) 3 (-2) 6.5 (0) 6.5 (0) 6.5 (0) 6.5 (0) 6.5 (0) 4 (-1) 10 (+2) 9 (+1) 9 (+1) 6.5 (0)

Table 4: Experimental design using CCD with the experimental and predicted value (using RSM) of citric acid production for aeration and agitation optimization.

Run 1 2 3 4 5 6 7 8 9 10 11

Agitation, Citric acid (g/kg-EFB) Aeration, B2 A2 (L/min) Experimental Predicted (times/day)

4 (-1) 12 (+1) 4 (-1) 12 (+1) 4 (-1) 12 (+1) 8 (0) 8 (0) 8 (0) 8 (0) 8 (0) 1 (-1) 1 (-1) 5 (+1) 5 (+1) 3 (0) 3 (0) 1 (-1) 5 (+1) 3 (0) 3 (0) 3 (0) 130.40 239.42 198.94 131.91 268.36 276.54 234.14 229.37 323.81 324.35 324.16 130.86 235.61 204.31 133.00 262.53 279.25 237.49 222.91 325.14 325.14 325.14

Building ANN
Data Collection, Processing and Analysis Determination of Number of Hidden Layers
Variations of the number of hidden neurons

Training

Combination of input variables

Carry out several training

Best model selection

Model Verification
The best model is selected according to : 1. Multiple linear regression, R 2. Mean Squared Error (MSE)

Results: R, MSE, number of hidden neurons, number of utilized weight

Methodology for training and validation

4 ANN models will be created using 4 sets of data

ANN MODEL 1: 1. Sucrose 2. Sago Starch 3. Cassava Flour 4. Urea 5. Methanol 6. KH2PO4 7. Fe 8. Zn 9. Mn 10. Cu 11. Mg ANN MODEL 2: 1. Sucrose 2. Mineral solutions (Zn, Cu, Mn, Mg) 3. Inoculum ANN MODEL 3: 1. Initial pH 2. Moisture Content 3. Temperature ANN MODEL 4: 1. Aeration 2. Agitation

Training

Experimental value of Citric Acid Production for each set of data (g/kg-EFB)

Make comparison and get the value of R

Prediction

Predicted Citric Acid Production (g/kg-EFB)

EXAMPLE OF COMMAND-LINE FUNCTION

% Building Neural Network Model % Define the input parameters and values % Sucrose with unit (% w/w) S =[8 6 4 6 6 4 4 6 8 8 4 6 6 6 6 3 6 8 9 6]; % Mineral Solution with unit (% w/w) MS=[12 2 12 8 8 4 4 8 4 4 12 8 8 14 8 8 8 12 8 8]; % Inoculum with unit (% w/w) I =[20 16 20 16 16 20 12 16 12 20 12 16 16 16 16 16 10 12 16 22]; % Assigning the input parameters [inputs]=[S;MS;I]; % Define the target values which is the citric acid production (CA) targets_CA=[259.23 267.47 256.52 334.68 334.23 208.69 236.65 332.44 259.22 260.65 250.02 333.88 333.08 292.42 334.14 247.46 257.79 285.06 278.9 217.41]; % Creating the network net=newff(inputs,targets_CA,50,{},'trainbfg'); % Train the network [net,tr]=train(net,inputs,targets_CA); % Predicted Output predicted_CA = sim(net,inputs); predicted_CA'; plotregression(targets_CA,predicted_CA)

SENSITIVITY ANALYSIS
Linear Correlation in MATLAB (corrcoef) Correlation coefficient quantifies the strength of a linear relationship between

two variables.
The correlation coefficients range from -1 to 1, where 1. Values close to 1 suggest that there is a positive linear relationship between the data columns.

2. Values close to -1 suggest that one column of data has a negative linear
relationship to another column of data (anti-correlation). 3. Values close to or equal to 0 suggest there is no linear relationship between the data columns

This command function will also give the p-value of each relationship. Each pvalue is the probability of getting a correlation as large as the observed value by random chance, when the true correlation is zero. Small p-value give better correlation (normally less than 0.05).

EXAMPLE OF COMMAND-LINE FUNCTION

% Sensitivity analysis using Linear Correlation % The response is the of Citric Acid Production in (g/kg-EFB) Output=[30.56 77.49 5.995 7.537 5.267 64.84 23.16 66.12 7.577 18.83 128.9 13.11]; % The 11 independent variables as input variable are defined as follows:

S=[3 3 0 0 3 3 0 3 0 0 3 0]; % Sucrose - unit is [%(w/w)]

% Calculate the correlation and p-value. % If p(i,j) is less than 0.05, then the correlation r(i,j) is significant. 'S' [r,p]=corrcoef(S,Output)

ans = S r= 1.0000 0.6665 p= 1.0000 0.0179

0.6665 1.0000 0.0179 1.0000

PART 1: EXPERIMENT
Preparation of materials, media and equipment Experimental procedure for solid state bioconversion Using optimal conditions obtained from ANN models Harvesting and extraction of citric acid

Determination of citric acid

However, the experiment will only be run if the optimum value of the parameters of each set are different from the optimum value obtained from RSM

RESULTS..

PRELIMINARY RESULTS
SENSITIVITY ANALYSIS

Graph of correlation coefficient value (r) for each media constituent

Graph of correlation coefficient value (r) for each parameters of media optimization

Number of Hidden Neurons: 20

Number of Hidden Neurons: 30

Number of Hidden Neurons: 50

ANN MODEL 2

METHOD No. of hidden neurons R value

ANN MODEL

RSM

20

30

50

0.8464

0.94987

0.93218

0.985

EXPECTED RESULTS
1. ANN Models more stable 2. The value of R should be higher than the value of R obtained by RSM 3. The best model can be selected from the best model of each set of data that give highest value of R 4. The optimum value of each parameter can be obtained from the best model 5. The optimum output can be predicted using the optimum value of parameters 6. The results of ANN cab be compared with the result from RSM

CONCLUSION
1. ANN can be used to optimize fermentation process. 2. Data from existing studies were used for training ANN model. 3. ANN can be used with multiple parameters as input. 4. ANN can be retrain with different number of hidden neurons

and other parameter to obtain better result.

5. MATLAB provide command-line functions and neural network toolboxes that help to build ANN model. 6. Model from the preliminary work is still unstable and need improvement

GANTT CHART
2009 No Tasks 1 1 Revise and improve the existing preliminary ANN model. Obtain the best model for optimization Get the optimum parameters and predicted output from the first model Build ANN models for other data and perform optimization Determine the optimum parameters and predicted output from the ANN models Plan and run experiment for validation if necessary Analyse results and write final report Preparation for FYP 2 presentation November 2 3 4 1 January 2 3 4 1 February 2 3 4 1 2010 March 2 3 4 1 April 2 3 4

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