The 33rd Annual Meeting of the Thai Society for Biotechnology and International Conference (TSB 2021)
Effect of pH and incubation period on autolysis of spent Saccharomyces cerevisiae derived from sago bioethanol fermentation
Nurashikin Suhaili*, Nik Nur Aziati Mahmod, Awang Ahmad Sallehin Awang Husaini and Dayang Salwani Awang Adeni
Faculty of Resource Science and Technology, UNIMAS, 94300 Kota Samarahan, Sarawak, Malaysia
*Corresponding author's email: snurashikin@unimas.my
The 33rd Annual Meeting of the Thai Society for Biotechnology
and International Conference (TSB 2021)
Presentation Outline
03
02
01
PROBLEM STATEMENTS
AIM & OBJECTIVES
INTRODUCTION
07
05
06
04
METHODOLOGY
RESULTS & DISCUSSION
ACKNOWLEDGEMENTS
CONCLUSIONS & FUTURE WORK
01
INTRODUCTION
Baker’s yeast (Saccharomyces cerevisiae) is commonly utilised in the fermentation industry. More than 400 000 tons of spent Baker’s yeast are produced as a by-product by the brewing and bioethanol industries. The spent Baker’s yeast in general is normally discarded into the environment5. The spent Baker's yeast is a rich source of proteins, essential amino acids, RNA, vitamin B and minerals, which essentially serve as a raw material in the production of yeast extract23. The extracts from the spent yeast can serve as promising sources of valuable ingredients such as bio-active compounds and amino acids that can be used for the production of functional food and dietary supplements18.
Saccharomyces cerevisiae
The extracts can be recovered by disrupting the yeast cell walls with the use of endogenous enzymes via autolysis11. Autolysis is termed as an irreversible process in which the yeast cell is degraded by the endogenous enzyme activity when the stationary phase of the cell growth ended7.
Spent Baker's yeast
02
PROBLEM STATEMENTS
Previous reports on autolysis in the literature focused on the use of either fresh yeast or spent yeast from bioethanol fermentation using commercial feedstock. There is still limited information on the valorisation of spent Baker’s yeast generated from bioethanol production using agricultural waste. One of the potential agricultural wastes for producing bioethanol is sago fibre. The feasibility of sago fibre as a feedstock for bioethanol fermentation has been reported in our previous report by Awang-Adeni et al.4. Considering the potential of sago bioethanol in meeting the increasing industrial demand in the future, it is timely to uncover the potential direction of the waste generated from sago bioethanol production. In our prior report by Mamat et al.16, we reported the utility of liquid waste stream from sago bioethanol as a potential feedstock for recombinant laccase production. However, the information on the potential and fate of the solid waste generated from sago bioethanol production namely the spent S. cerevisiae still remains unexplored.
03
AIM & OBJECTIVES
AimTo investigate the feasibility of autolysis of spent Baker’s yeast generated from sago bioethanol fermentation Objectives To study the effect of pH and incubation time on protein and carbohydrate concentration released in the yeast lysate. In general, the present work provides useful insight into the development of on-site process stream, which may benefit the establishment of an integrated sago biorefinery in the future. Furthermore, the present work will help to promote sustainable development of sago bioethanol production with reduced waste whilst generating side revenue to the industry.
04
METHODOLOGY
What are we
going to learn?
How are we
going to learn?
Who I am?
Calendar
How are we going to evaluate?
What are we
going to need?
Resources
of interest
Video
05
RESULTS
OVERVIEW
- Effect of different initial pH values and incubation period on protein concentration of autolysate
- Effect of different initial pH values and incubation period on carbohydrate concentration of autolysate
- Morphological analysis of autolysed cells
05
RESULTS (cont)
Effect of pH and incubation time on protein concentration of the lysate
Autolysis at pH 3 and pH 5 produced the highest protein concentration in the lysate with the enhancement of 2.5-fold over that in the control sample. The activity of hydrolytic enzymes in yeast cells, especially proteases is greatly influenced by the pH of the autolysis20. Acidic pH is revealed to be beneficial for some key enzymes responsible for the autolysis such as protease A, which has an optimum pH between 2 and 615
Fig. 1: Changes of protein concentration of autolysate over time
05
RESULTS (cont)
Effect of pH and incubation time on carbohydrate concentration of the lysate
Autolysis at pH 3 and pH 5 produced the highest carbohydrate concentration in the lysate with the enhancement of 2-fold over that in the control sample. Considering the release of both protein and carbohydrate from the lysed yeast cells, it can be concluded that 72 hours of incubation is the best period for autolysis of spent Baker’s yeast in this work. Unnecessary extension of incubation period during the autolysis may not be favourable since it may increase the cost and also the risk of contamination due to the growth of pathogens over a long period. Hence, early insight into the feasibility of lysing the cell wall of spent Baker’s yeast generated from sago bioethanol production as demonstrated in this work will benefit the development of potential side process such as those that are related to yeast cell wall products that can be developed along with the sago bioethanol industry in the future.
Fig. 2: Changes of total carbohydrate concentration of autolysate over time
05
RESULTS (cont)
Changes of surface morphology of lysed yeast cells derived from autolysis conducted at pH 3 and pH 5
The yeast cell wall, which has a thickness of 100-150 nm, is a key component that determines the overall shape of the yeast cells25. During the lysis of the cells, the vacuole will be ruptured leading to the release of the hydrolytic enzymes that are responsible for the degradation of the organelles and cell membrane components25. This in turn resulted in the wrinkling of the cell wall and subsequently the reduction of the diameter of the cells throughout the autolysis.
Fig. 3: Changes of surface morphology of (a) untreated yeast cells (control samples); (b) yeast cells derived from autolysis conducted at pH 3 and (c) yeast cells derived from autolysis conducted at pH 5.
06
CONCLUSIONS & FUTURE WORK
Highlight 1
Highlight 2
Highlight 3
The feasibility of autolysis of spent Baker’s yeast generated from sago bioethanol fermentation has been demonstrated.
The initial pH of 3 and incubation period of 72 hours were proven as the best conditions for the autolysis on the basis of protein and total carbohydrate released in the lysate with the enhancements of 2.5-fold and 2-fold over that in the control samples, respectively.
Future work should consider the influence of other parameters such as organic solvents and mechanical intervention on the efficiency of autolysis.
07
ACKNOWLEDGEMENTS
Ministry of Higher Education Malaysia, Fundamental Research Grant Scheme, RACER/1/2019/STG0S/UNIMAS//l Herdson Sago Mill
Thanks!
snurashikin@unimas.my
Esto es un párrafo listo para contener creatividad, experiencias e historias geniales.
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The 33rd Annual Meeting of the Thai Society for Biotechnology and International Conference (TSB 2021)
Effect of pH and incubation period on autolysis of spent Saccharomyces cerevisiae derived from sago bioethanol fermentation
Nurashikin Suhaili*, Nik Nur Aziati Mahmod, Awang Ahmad Sallehin Awang Husaini and Dayang Salwani Awang Adeni
Faculty of Resource Science and Technology, UNIMAS, 94300 Kota Samarahan, Sarawak, Malaysia *Corresponding author's email: snurashikin@unimas.my
The 33rd Annual Meeting of the Thai Society for Biotechnology and International Conference (TSB 2021)
Presentation Outline
03
02
01
PROBLEM STATEMENTS
AIM & OBJECTIVES
INTRODUCTION
07
05
06
04
METHODOLOGY
RESULTS & DISCUSSION
ACKNOWLEDGEMENTS
CONCLUSIONS & FUTURE WORK
01
INTRODUCTION
Baker’s yeast (Saccharomyces cerevisiae) is commonly utilised in the fermentation industry. More than 400 000 tons of spent Baker’s yeast are produced as a by-product by the brewing and bioethanol industries. The spent Baker’s yeast in general is normally discarded into the environment5. The spent Baker's yeast is a rich source of proteins, essential amino acids, RNA, vitamin B and minerals, which essentially serve as a raw material in the production of yeast extract23. The extracts from the spent yeast can serve as promising sources of valuable ingredients such as bio-active compounds and amino acids that can be used for the production of functional food and dietary supplements18.
Saccharomyces cerevisiae
The extracts can be recovered by disrupting the yeast cell walls with the use of endogenous enzymes via autolysis11. Autolysis is termed as an irreversible process in which the yeast cell is degraded by the endogenous enzyme activity when the stationary phase of the cell growth ended7.
Spent Baker's yeast
02
PROBLEM STATEMENTS
Previous reports on autolysis in the literature focused on the use of either fresh yeast or spent yeast from bioethanol fermentation using commercial feedstock. There is still limited information on the valorisation of spent Baker’s yeast generated from bioethanol production using agricultural waste. One of the potential agricultural wastes for producing bioethanol is sago fibre. The feasibility of sago fibre as a feedstock for bioethanol fermentation has been reported in our previous report by Awang-Adeni et al.4. Considering the potential of sago bioethanol in meeting the increasing industrial demand in the future, it is timely to uncover the potential direction of the waste generated from sago bioethanol production. In our prior report by Mamat et al.16, we reported the utility of liquid waste stream from sago bioethanol as a potential feedstock for recombinant laccase production. However, the information on the potential and fate of the solid waste generated from sago bioethanol production namely the spent S. cerevisiae still remains unexplored.
03
AIM & OBJECTIVES
AimTo investigate the feasibility of autolysis of spent Baker’s yeast generated from sago bioethanol fermentation Objectives To study the effect of pH and incubation time on protein and carbohydrate concentration released in the yeast lysate. In general, the present work provides useful insight into the development of on-site process stream, which may benefit the establishment of an integrated sago biorefinery in the future. Furthermore, the present work will help to promote sustainable development of sago bioethanol production with reduced waste whilst generating side revenue to the industry.
04
METHODOLOGY
What are we going to learn?
How are we going to learn?
Who I am?
Calendar
How are we going to evaluate?
What are we going to need?
Resources of interest
Video
05
RESULTS
OVERVIEW
05
RESULTS (cont)
Effect of pH and incubation time on protein concentration of the lysate
Autolysis at pH 3 and pH 5 produced the highest protein concentration in the lysate with the enhancement of 2.5-fold over that in the control sample. The activity of hydrolytic enzymes in yeast cells, especially proteases is greatly influenced by the pH of the autolysis20. Acidic pH is revealed to be beneficial for some key enzymes responsible for the autolysis such as protease A, which has an optimum pH between 2 and 615
Fig. 1: Changes of protein concentration of autolysate over time
05
RESULTS (cont)
Effect of pH and incubation time on carbohydrate concentration of the lysate
Autolysis at pH 3 and pH 5 produced the highest carbohydrate concentration in the lysate with the enhancement of 2-fold over that in the control sample. Considering the release of both protein and carbohydrate from the lysed yeast cells, it can be concluded that 72 hours of incubation is the best period for autolysis of spent Baker’s yeast in this work. Unnecessary extension of incubation period during the autolysis may not be favourable since it may increase the cost and also the risk of contamination due to the growth of pathogens over a long period. Hence, early insight into the feasibility of lysing the cell wall of spent Baker’s yeast generated from sago bioethanol production as demonstrated in this work will benefit the development of potential side process such as those that are related to yeast cell wall products that can be developed along with the sago bioethanol industry in the future.
Fig. 2: Changes of total carbohydrate concentration of autolysate over time
05
RESULTS (cont)
Changes of surface morphology of lysed yeast cells derived from autolysis conducted at pH 3 and pH 5
The yeast cell wall, which has a thickness of 100-150 nm, is a key component that determines the overall shape of the yeast cells25. During the lysis of the cells, the vacuole will be ruptured leading to the release of the hydrolytic enzymes that are responsible for the degradation of the organelles and cell membrane components25. This in turn resulted in the wrinkling of the cell wall and subsequently the reduction of the diameter of the cells throughout the autolysis.
Fig. 3: Changes of surface morphology of (a) untreated yeast cells (control samples); (b) yeast cells derived from autolysis conducted at pH 3 and (c) yeast cells derived from autolysis conducted at pH 5.
06
CONCLUSIONS & FUTURE WORK
Highlight 1
Highlight 2
Highlight 3
The feasibility of autolysis of spent Baker’s yeast generated from sago bioethanol fermentation has been demonstrated.
The initial pH of 3 and incubation period of 72 hours were proven as the best conditions for the autolysis on the basis of protein and total carbohydrate released in the lysate with the enhancements of 2.5-fold and 2-fold over that in the control samples, respectively.
Future work should consider the influence of other parameters such as organic solvents and mechanical intervention on the efficiency of autolysis.
07
ACKNOWLEDGEMENTS
Ministry of Higher Education Malaysia, Fundamental Research Grant Scheme, RACER/1/2019/STG0S/UNIMAS//l Herdson Sago Mill
Thanks!
snurashikin@unimas.my
Esto es un párrafo listo para contener creatividad, experiencias e historias geniales.