Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172

In this study, the use of NaOH treatment and chemical-free aqueous solution for intracellular polyhydroxyalkanoates (PHAs) recovery from local isolate Gram-negative wild type bacteria Comamonas sp. EB172 at different NaOH concentrations, treatment times, temperatures and agitations is being evaluat...

Full description

Saved in:

Bibliographic Details
Main Author:	Mohammadi, Mitra
Format:	Thesis
Language:	English
Published:	2012
Subjects:	Poly-beta-hydroxyalkanoates Gram-negative bacteria
Online Access:	http://psasir.upm.edu.my/id/eprint/32155/1/FBSB%202012%2018R.pdf
Tags:	Add Tag No Tags, Be the first to tag this record!

id	my-upm-ir.32155
record_format	uketd_dc
spelling	my-upm-ir.321552015-01-19T03:46:36Z Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172 2012-02 Mohammadi, Mitra In this study, the use of NaOH treatment and chemical-free aqueous solution for intracellular polyhydroxyalkanoates (PHAs) recovery from local isolate Gram-negative wild type bacteria Comamonas sp. EB172 at different NaOH concentrations, treatment times, temperatures and agitations is being evaluated as alternative and appropriate methods instead of halogenated solvent extraction system. These methods consist of recovery steps followed by incubation, centrifugation and purification steps using ethanol and water washing. The results of this study were compared with PHA extraction from recombinant Cupriavidus necator as a control. The PHA recovered under the most favourable conditions was further characterized. The chemical-free aqueous solution method as a clean process and low concentration of NaOH treatment were successfully developed for PHA extraction from wild type Comamonas sp. EB172. More than 88% purity and 96% recovery yield of PHA were achieved by incubating the wild type bacteria under the mild alkaline treatment using 0.05 M NaOH at 4°C for 1 h. However, up to 96% purity and recovery yield can be obtained by incubation of recombinant C. necator with 0.05 M NaOH at 4°C for 3 h. The incubation of Comamonas sp. EB172 in the chemical-free aqueous solution at 30°C for 5 h could efficiently extract the PHA with more than 93% of recovery yield. Besides, shorter incubation time is required for PHA extraction from recombinant strain using the non-chemical method, which resulted in a PHA with 80.6% of purity and recovery yield of 96.1%. Protein determination and transmission electron microscopy images as well as gas chromatography analysis proved that improvement in cell wall permeability and cell membrane breakage are the possible mechanisms of NaOH treatment and chemical-free aqueous solution method on PHA recovery which was accompanied by considerable release of protein after the extraction step. It was found that the effectiveness of chemical and non-chemical treatments depends on the microbial strain. The initial intracellular PHA content also affected the effectiveness of the extraction methods for PHA recovery. Although, the NaOH treatment could recover purer PHA as compared to the chemical-free method, both processes were able to extract the polymer with high yield. The chemical-free aqueous solution method was found to be better than NaOH treatment for PHA extraction in respect of the final polymer molecular weight, which is in fact almost double that of the chloroform-extracted PHA as control. The overall results in this study indicated that the mild NaOH treatment and the chemical-free aqueous solution methods developed can serve as alternative recovery methods with high potential instead of the conventional halogenated solvent extraction process such as chloroform, since these new methods are environmentally more benign, effective and simple in operation. The recovery of intracellular PHA from Comamonas sp. EB172 cells via the recovery methods developed herein can contribute towards the sustainable process of PHA production using organic acids derived from the anaerobic treatment of palm oil mill effluent (POME). Poly-beta-hydroxyalkanoates Gram-negative bacteria 2012-02 Thesis http://psasir.upm.edu.my/id/eprint/32155/ http://psasir.upm.edu.my/id/eprint/32155/1/FBSB%202012%2018R.pdf application/pdf en public phd doctoral Universiti Putra Malaysia Poly-beta-hydroxyalkanoates Gram-negative bacteria Faculty of Biotechnology and Biomolecular Sciences
institution	Universiti Putra Malaysia
collection	PSAS Institutional Repository
language	English
topic	Poly-beta-hydroxyalkanoates Gram-negative bacteria
spellingShingle	Poly-beta-hydroxyalkanoates Gram-negative bacteria Mohammadi, Mitra Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172
description	In this study, the use of NaOH treatment and chemical-free aqueous solution for intracellular polyhydroxyalkanoates (PHAs) recovery from local isolate Gram-negative wild type bacteria Comamonas sp. EB172 at different NaOH concentrations, treatment times, temperatures and agitations is being evaluated as alternative and appropriate methods instead of halogenated solvent extraction system. These methods consist of recovery steps followed by incubation, centrifugation and purification steps using ethanol and water washing. The results of this study were compared with PHA extraction from recombinant Cupriavidus necator as a control. The PHA recovered under the most favourable conditions was further characterized. The chemical-free aqueous solution method as a clean process and low concentration of NaOH treatment were successfully developed for PHA extraction from wild type Comamonas sp. EB172. More than 88% purity and 96% recovery yield of PHA were achieved by incubating the wild type bacteria under the mild alkaline treatment using 0.05 M NaOH at 4°C for 1 h. However, up to 96% purity and recovery yield can be obtained by incubation of recombinant C. necator with 0.05 M NaOH at 4°C for 3 h. The incubation of Comamonas sp. EB172 in the chemical-free aqueous solution at 30°C for 5 h could efficiently extract the PHA with more than 93% of recovery yield. Besides, shorter incubation time is required for PHA extraction from recombinant strain using the non-chemical method, which resulted in a PHA with 80.6% of purity and recovery yield of 96.1%. Protein determination and transmission electron microscopy images as well as gas chromatography analysis proved that improvement in cell wall permeability and cell membrane breakage are the possible mechanisms of NaOH treatment and chemical-free aqueous solution method on PHA recovery which was accompanied by considerable release of protein after the extraction step. It was found that the effectiveness of chemical and non-chemical treatments depends on the microbial strain. The initial intracellular PHA content also affected the effectiveness of the extraction methods for PHA recovery. Although, the NaOH treatment could recover purer PHA as compared to the chemical-free method, both processes were able to extract the polymer with high yield. The chemical-free aqueous solution method was found to be better than NaOH treatment for PHA extraction in respect of the final polymer molecular weight, which is in fact almost double that of the chloroform-extracted PHA as control. The overall results in this study indicated that the mild NaOH treatment and the chemical-free aqueous solution methods developed can serve as alternative recovery methods with high potential instead of the conventional halogenated solvent extraction process such as chloroform, since these new methods are environmentally more benign, effective and simple in operation. The recovery of intracellular PHA from Comamonas sp. EB172 cells via the recovery methods developed herein can contribute towards the sustainable process of PHA production using organic acids derived from the anaerobic treatment of palm oil mill effluent (POME).
format	Thesis
qualification_name	Doctor of Philosophy (PhD.)
qualification_level	Doctorate
author	Mohammadi, Mitra
author_facet	Mohammadi, Mitra
author_sort	Mohammadi, Mitra
title	Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172
title_short	Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172
title_full	Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172
title_fullStr	Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172
title_full_unstemmed	Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172
title_sort	non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate comamonas sp. eb172
granting_institution	Universiti Putra Malaysia
granting_department	Faculty of Biotechnology and Biomolecular Sciences
publishDate	2012
url	http://psasir.upm.edu.my/id/eprint/32155/1/FBSB%202012%2018R.pdf
_version_	1747811646911283200

Non-halogenated recovery method of intracellular polyhydroxyalkanoates from local isolate Comamonas sp. EB172

Similar Items