Elsevier

New Biotechnology

Volume 27, Issue 1, 28 February 2010, Pages 25-32
New Biotechnology

Research paper
Enhanced production of xylanase by Aspergillus carneus M34 in solid-state fermentation with agricultural waste using statistical approach

https://doi.org/10.1016/j.nbt.2009.09.008Get rights and content

The production of a low molecular weight xylanase by Aspergillus carneus M34 was investigated in solid-state fermentation using agricultural waste as the substrate. Experimental designs were practiced for optimisation of the medium composition. When agricultural wastes of coba husk and corn steep liquor were used in the ratio of 4.5:0.5, a 22.5% increase of xylanase activity was observed compared with the medium containing only coba husk. The incubation time for xylanase production can be reduced from 12 days to six days by increasing the inoculum size to 2 × 107 spores mL−1. The optimal media compositions were as follows by using statistical approach: coba husk/corn steep liquor (4.5/0.5); NH4NO3, 32.4 g L−1; CaCl2, 1.34 g L−1; MnSO4·7H2O, 0.0124 g L−1. Xylanase activity of 1721 U g−1 substrate was obtained by A. carneus M34 in a six-day period at 30 °C, which was a 227% increase compared with that obtained before applying the Plackett–Burman and response surface methodology experimental design.

Introduction

Lignocellulose, the most abundant and renewable biomass available on earth, contains three major groups of polymers, that is, cellulose (30–45%), hemicellulose (25–45%), and lignin (15–30%) [1]. Xylan (mainly β-1, 4-linked d-xylose), which is a major structural component of plant cell walls, is the second most abundant biopolymer after cellulose and belongs to the group of complex structural polymers collectively referred to as hemicelluloses [2]. The hydrolysis of xylan requires the action of different types of enzymes, among which xylanases (1, 4-β-d-xylan xylanohydrolase, EC 3.2.1.8) [3] and β-xylosidases (1, 4-β-d-xylan xylohydrolase, EC 3.2.1.37) are the best characterized [4]. Xylanases have been used in many applications, including energy generation, waste treatment, production of chemicals, clarification of juices, and paper manufacture 1, 5. In addition, xylanases have potential for use in the production of xylooligosaccharides, ethanol, and other useful substances 6, 7.

Microbial xylanases have received a great deal of attention in recent years. A large number of microorganisms, including bacteria, fungi, actinomycetes, and yeasts have been reported to produce xylanolytic enzymes 4, 8, 9. Filamentous fungi are particularly interesting producers of xylanases from an industrial point of view owing to the fact that they produce extracellular xylanase [10]. Among xylanolytic microorganisms, Aspergillus has been shown to be an efficient producer of xylanases [11].

Solid-state fermentation (SSF) systems have generated much interest in recent years because they offer several economical and practical advantages over submerged cultivation. The advantages include the simplicity of the cultivation equipment, improved product recovery, reduced wastewater output, higher product concentration, lower capital investment, and lower plant operational costs 12, 13. This system has been successfully applied in the preparation of several high-value products, such as enzymes, organic acids, secondary metabolites, fuels, pesticides, and aromatic compounds 11, 14, 15. In addition, the use of solid-state cultivation has been reported to be more advantageous than submerged fermentation, because, in addition to lower production costs, better physiochemical properties of enzyme can be obtained in SSF [16].

Optimisation of fermentation conditions using conventional methods is time consuming and costly, especially for a large number of variables. Statistical experimental designs, such as PB (Plackett–Burman) and RSM (Response surface methodology), are alternative strategies that involve a minimum number of experiments for a large number of factors. These methods have been employed to improve the production of enzymes such as α-amylase [17], lipase [18], xylanase [3] and l-leucine amino peptidase [19].

A novel phytase-producing and xylanase-producing strain of A. carneus M34, which was isolated in our laboratory, produced low molecular weight xylanase, and the production was optimised in submerged fermentation [3]. The statistical experimental design strategies was that PB was used to determine which of the nutritional salt variables were the most relevant, and then RSM was used to determine their optimal values. The present investigation describes the successful optimisation for the production of this xylanase (group I of family 11 endoxylanases) [20] by A. carneus M34 in media containing agricultural waste using PB experimental design and RSM in solid-state fermentation.

Section snippets

Microorganism and culture condition

A. carneus M34 was isolated from Taiwan soil during a screening study for phytase-producing microorganisms and identified by the Food Industry Research and Development Institute (FIRDI) in Hsinchu, Taiwan. The microorganism was grown on potato dextrose agar (PDA) for 14 days at 30 °C and then stored at 4 °C. Stock cultures were transferred to fresh medium every five to six weeks and incubated under the same conditions. The conidial suspensions were prepared by adding 10 mL of sterilized 0.05%

Effect of different agricultural wastes on xylanase production and growth

The effects of various agricultural wastes on the xylanase activity of A. carneus M34 in SSF are given in Table 4. In these experiments, the nitrogen sources were from the 5 mL of nutrient salt solution that contained 5 g L−1 of NH4NO3. Growth on medium with coba husk and corn steep liquor at the ratio of 4.5:0.5 resulted in maximum production of xylanase activity (645 U g−1 dry substrate), followed by the 4:1 ratio of coba husk/corn steep liquor (601 U g−1 dry substrate) after six days of cultivation

Discussion

A major problem experienced by agro-based industries is the management of wastes. Improper methods of disposal of solid waste can cause various hazards, such as pollution of air, degradation of water quality, and negative impacts on public health. Although chemical treatment of solid wastes can be used, to date, emphasis has been on biological conversion of plant products. Fungi are capable of hydrolysing complex organic compounds as a major source of energy. Agricultural wastes such as banana

Conclusions

To the best of our knowledge, there are no reports of optimisation of xylanase production by A. carneus M34, the isolate from Taiwan soil, using statistical approach in solid-state fermentation with agricultural waste. In conclusion, the A. carneus M34 microorganism shows improved production of xylanase in SSF than in submerged fermentation [3]. By using cheap agricultural wastes, such as coba husk, a high activity of xylanase was obtained in six days. Since this enzyme has been commercially

Acknowledgement

We gratefully acknowledge the support for this research by the National Science Council, Taiwan, and Republic of China (NSC94-2313-B005-058).

References (38)

Cited by (0)

View full text