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Screening, Gene Sequencing and Characterising of Lipase for Methanolysis of Crude Palm Oil

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Abstract

Staphylococcus sp. WL1 lipase (LipFWS) was investigated for methanolysis of crude palm oil (CPO) at moderate temperatures. Experiments were conducted in the following order: searching for the suitable bacterium for producing lipase from activated sludge, sequencing lipase gene, identifying lipase activity, then synthesising CPO biodiesel using the enzyme. From bacterial screening, one isolated specimen which consistently showed the highest extracellular lipase activity was identified as Staphylococcus sp. WL1 possessing lipFWS (lipase gene of 2,244 bp). The LipFWS deduced was a protein of 747 amino acid residues containing an α/β hydrolase core domain with predicted triad catalytic residues to be Ser474, His704 and Asp665. Optimal conditions for the LipFWS activity were found to be at 55 °C and pH 7.0 (in phosphate buffer but not in Tris buffer). The lipase had a K M of 0.75 mM and a V max of 0.33 mM min−1 on p-nitrophenyl palmitate substrate. The lyophilised crude LipFWS performed as good as the commonly used catalyst potassium hydroxide for methanolysis of CPO. ESI-IT-MS spectra indicated that the CPO was converted into biodiesel, suggesting that free LipFWS is a worthy alternative for CPO biodiesel synthesis.

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Acknowledgments

Funding was provided by the Ministry of Education and Culture, the Republic of Indonesia through Beasiswa Unggulan Program as well as Research Incentive Program for Researcher and Engineer no. 02/SK/BPPI/2009. We thank Prof. Yana Maolana Syah for his ESI-IT-MS analysis of CPO and biodiesels as well as Mr. Tubagus Andhika Nugraha for critical reading of this manuscript.

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Authors and Affiliations

  1. Biochemistry Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Jl. Ganesha 10, Bandung, 40132, Indonesia

    Enny Ratnaningsih, Dewi Handayani, Fatiha Khairunnisa,  Ihsanawati, Sari Dewi Kurniasih, Bill Mangindaan & Zeily Nurachman

  2. Environmental Laboratory, Centre for Textile, Ministry of Industry, Jl. Jend. Ahmad Yani 390, Bandung, 40272, Indonesia

    Sinta Rismayani & Cica Kasipah

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  1. Enny Ratnaningsih
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Correspondence to Zeily Nurachman.

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Ratnaningsih, E., Handayani, D., Khairunnisa, F. et al. Screening, Gene Sequencing and Characterising of Lipase for Methanolysis of Crude Palm Oil. Appl Biochem Biotechnol 170, 32–43 (2013). https://doi.org/10.1007/s12010-013-0160-z

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  • DOI: https://doi.org/10.1007/s12010-013-0160-z

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