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Saccharification and liquefaction of cassava starch: an alternative source for the production of bioethanol using amylolytic enzymes by double fermentation process.

Pervez S, Aman A, Iqbal S, Siddiqui NN, Ul Qader SA - BMC Biotechnol. (2014)

Bottom Line: The distillate originated after recovery of bioethanol gave 53.0% yield.An improved and effective dual enzymatic starch degradation method is designed for the production of bioethanol using cassava starch.The technique developed is more profitable due to its fast liquefaction and saccharification approach that was employed for the formation of glucose and ultimately resulted in higher yields of alcohol production.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Karachi Institute of Biotechnology & Genetic Engineering (KIBGE), University of Karachi, Karachi -75270, Pakistan. ali_kibge@yahoo.com.

ABSTRACT

Background: Cassava starch is considered as a potential source for the commercial production of bioethanol because of its availability and low market price. It can be used as a basic source to support large-scale biological production of bioethanol using microbial amylases. With the progression and advancement in enzymology, starch liquefying and saccharifying enzymes are preferred for the conversion of complex starch polymer into various valuable metabolites. These hydrolytic enzymes can selectively cleave the internal linkages of starch molecule to produce free glucose which can be utilized to produce bioethanol by microbial fermentation.

Results: In the present study, several filamentous fungi were screened for production of amylases and among them Aspergillus fumigatus KIBGE-IB33 was selected based on maximum enzyme yield. Maximum α-amylase, amyloglucosidase and glucose formation was achieved after 03 days of fermentation using cassava starch. After salt precipitation, fold purification of α-amylase and amyloglucosidase increased up to 4.1 and 4.2 times with specific activity of 9.2 kUmg⁻¹ and 393 kUmg⁻¹, respectively. Concentrated amylolytic enzyme mixture was incorporated in cassava starch slurry to give maximum glucose formation (40.0 gL⁻¹), which was further fermented using Saccharomyces cerevisiae into bioethanol with 84.0% yield. The distillate originated after recovery of bioethanol gave 53.0% yield.

Conclusion: An improved and effective dual enzymatic starch degradation method is designed for the production of bioethanol using cassava starch. The technique developed is more profitable due to its fast liquefaction and saccharification approach that was employed for the formation of glucose and ultimately resulted in higher yields of alcohol production.

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Percent saccharification of cassava starch and glucose formation at different reaction time intervals.
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Figure 4: Percent saccharification of cassava starch and glucose formation at different reaction time intervals.

Mentions: Conversion of starchy materials into ethanol is an intricate process and several attempts have been made to produce bioethanol in commercially feasible quantities and to easily scale-up the methodology used. Cassava starch is a complex molecule containing amylose and amylopectin and for the production of bioethanol, first the starch molecules must be hydrolyzed into more simple sugars. Some pretreatment techniques including hot water and steam explosion treatment, alkaline and solvent pretreatment, acid hydrolysis and enzymatic degradation for the breakdown of complex starch molecule into simpler sugars have also been studied [34-37]. More recently, a new pre-treatment technique known as popping pre-treatment have gained attention for the hydrolysis of starchy feedstock [38]. However, enzymatic degradation using different hydrolases is mostly preferred because during acid hydrolysis the percent conversion of starch into reducing sugars is low as compared to the enzymatic degradation [39-41]. With the progression and advancement in enzymology, amylolytic enzymes are now preferable over conventional methods because enzymatic treatments lead towards high yield of glucose with reduced energy consumption. Therefore, in the current study gelatinized cassava starch was liquefied using α-amylase and was further saccharified by means of amyloglucosidase. However, before breaking starch into simple fermentable sugars, the time required for both the processes to occur effectively was also analyzed by incubating the gelatinized starch slurry with both partially purified amylolytic enzymes for different time intervals. Glucose was the main end-product which is required for production of bioethanol, therefore the concentration of glucose formation as well as percent saccharification was monitored throughout this study. Gelatinized cassava starch was mixed with partially purified α-amylase (9.2 kUmg-1) and amyloglucosidase (393.0 kUmg-1). It was observed that as the reaction time increases, the formation of glucose (40.0 gL-1) as well as percent saccharification (60.0%) also increased up to 90.0 minutes and beyond that both parameters become constant (Figure 4). This glucose containing mixture was further used for the production of ethanol. Efficiency of enzymatic liquefaction and saccharification also depends upon optimum enzyme activity as well as the purity of amylolytic enzymes as crude enzyme takes longer time period to completely hydrolyze starch molecule into glucose as compared to the purified enzyme (Table 3). This percent saccharification (60.0%) could also be further augmented by either improving the purity of enzyme or by incorporating other hydrolyase (xylanases, pectinases or cellulases) along with these amylolytic enzymes [42,43]. As reported earlier further increase in percent saccharification could also be achieved if the starch slurry was autoclaved before addition of amylolytic enzyme [44]. Similarly, Aggarwal et al.[45] and Soni et al.[46] have also discussed about the role of the purity level of amylolytic enzymes during starch hydrolysis. In the same way, Shanavas et al.[20] have also previously analyzed the effect of reaction time on saccharification of cassava starch and have obtained maximum percent saccharification after 30.0 minutes of incubation followed by slight increase when using commercially available starch hydrolyzing enzymes. On the contrary, Aggarwal et al.[45] reported maximum percent saccharification using crude amylolytic enzymes after 24 hours of incubation time. Very recently, Gohel et al. [47] used simultaneous saccharification and solid state fermentation for the production of ethanol using Indian sorghum feedstock and also incorporated acid fungal protease instead of urea for better ethanol yield.


Saccharification and liquefaction of cassava starch: an alternative source for the production of bioethanol using amylolytic enzymes by double fermentation process.

Pervez S, Aman A, Iqbal S, Siddiqui NN, Ul Qader SA - BMC Biotechnol. (2014)

Percent saccharification of cassava starch and glucose formation at different reaction time intervals.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4048251&req=5

Figure 4: Percent saccharification of cassava starch and glucose formation at different reaction time intervals.
Mentions: Conversion of starchy materials into ethanol is an intricate process and several attempts have been made to produce bioethanol in commercially feasible quantities and to easily scale-up the methodology used. Cassava starch is a complex molecule containing amylose and amylopectin and for the production of bioethanol, first the starch molecules must be hydrolyzed into more simple sugars. Some pretreatment techniques including hot water and steam explosion treatment, alkaline and solvent pretreatment, acid hydrolysis and enzymatic degradation for the breakdown of complex starch molecule into simpler sugars have also been studied [34-37]. More recently, a new pre-treatment technique known as popping pre-treatment have gained attention for the hydrolysis of starchy feedstock [38]. However, enzymatic degradation using different hydrolases is mostly preferred because during acid hydrolysis the percent conversion of starch into reducing sugars is low as compared to the enzymatic degradation [39-41]. With the progression and advancement in enzymology, amylolytic enzymes are now preferable over conventional methods because enzymatic treatments lead towards high yield of glucose with reduced energy consumption. Therefore, in the current study gelatinized cassava starch was liquefied using α-amylase and was further saccharified by means of amyloglucosidase. However, before breaking starch into simple fermentable sugars, the time required for both the processes to occur effectively was also analyzed by incubating the gelatinized starch slurry with both partially purified amylolytic enzymes for different time intervals. Glucose was the main end-product which is required for production of bioethanol, therefore the concentration of glucose formation as well as percent saccharification was monitored throughout this study. Gelatinized cassava starch was mixed with partially purified α-amylase (9.2 kUmg-1) and amyloglucosidase (393.0 kUmg-1). It was observed that as the reaction time increases, the formation of glucose (40.0 gL-1) as well as percent saccharification (60.0%) also increased up to 90.0 minutes and beyond that both parameters become constant (Figure 4). This glucose containing mixture was further used for the production of ethanol. Efficiency of enzymatic liquefaction and saccharification also depends upon optimum enzyme activity as well as the purity of amylolytic enzymes as crude enzyme takes longer time period to completely hydrolyze starch molecule into glucose as compared to the purified enzyme (Table 3). This percent saccharification (60.0%) could also be further augmented by either improving the purity of enzyme or by incorporating other hydrolyase (xylanases, pectinases or cellulases) along with these amylolytic enzymes [42,43]. As reported earlier further increase in percent saccharification could also be achieved if the starch slurry was autoclaved before addition of amylolytic enzyme [44]. Similarly, Aggarwal et al.[45] and Soni et al.[46] have also discussed about the role of the purity level of amylolytic enzymes during starch hydrolysis. In the same way, Shanavas et al.[20] have also previously analyzed the effect of reaction time on saccharification of cassava starch and have obtained maximum percent saccharification after 30.0 minutes of incubation followed by slight increase when using commercially available starch hydrolyzing enzymes. On the contrary, Aggarwal et al.[45] reported maximum percent saccharification using crude amylolytic enzymes after 24 hours of incubation time. Very recently, Gohel et al. [47] used simultaneous saccharification and solid state fermentation for the production of ethanol using Indian sorghum feedstock and also incorporated acid fungal protease instead of urea for better ethanol yield.

Bottom Line: The distillate originated after recovery of bioethanol gave 53.0% yield.An improved and effective dual enzymatic starch degradation method is designed for the production of bioethanol using cassava starch.The technique developed is more profitable due to its fast liquefaction and saccharification approach that was employed for the formation of glucose and ultimately resulted in higher yields of alcohol production.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Karachi Institute of Biotechnology & Genetic Engineering (KIBGE), University of Karachi, Karachi -75270, Pakistan. ali_kibge@yahoo.com.

ABSTRACT

Background: Cassava starch is considered as a potential source for the commercial production of bioethanol because of its availability and low market price. It can be used as a basic source to support large-scale biological production of bioethanol using microbial amylases. With the progression and advancement in enzymology, starch liquefying and saccharifying enzymes are preferred for the conversion of complex starch polymer into various valuable metabolites. These hydrolytic enzymes can selectively cleave the internal linkages of starch molecule to produce free glucose which can be utilized to produce bioethanol by microbial fermentation.

Results: In the present study, several filamentous fungi were screened for production of amylases and among them Aspergillus fumigatus KIBGE-IB33 was selected based on maximum enzyme yield. Maximum α-amylase, amyloglucosidase and glucose formation was achieved after 03 days of fermentation using cassava starch. After salt precipitation, fold purification of α-amylase and amyloglucosidase increased up to 4.1 and 4.2 times with specific activity of 9.2 kUmg⁻¹ and 393 kUmg⁻¹, respectively. Concentrated amylolytic enzyme mixture was incorporated in cassava starch slurry to give maximum glucose formation (40.0 gL⁻¹), which was further fermented using Saccharomyces cerevisiae into bioethanol with 84.0% yield. The distillate originated after recovery of bioethanol gave 53.0% yield.

Conclusion: An improved and effective dual enzymatic starch degradation method is designed for the production of bioethanol using cassava starch. The technique developed is more profitable due to its fast liquefaction and saccharification approach that was employed for the formation of glucose and ultimately resulted in higher yields of alcohol production.

Show MeSH
Related in: MedlinePlus