title: Understanding the Relative Importance of the Effects of Soluble and Insoluble Inhibitors on the Enzyme Digestibility of Steam Pretreated Radiata Pine

authors: MacAskill, Jessica Jane

abstract: Lignocellulosic biomass use is one of the keys to growth and development of future biofuels and biochemicals. Amongst the lignocellulosic feedstocks, softwoods are particularly attractive as they have the potential to produce high sugar yields and are prevalent in many forests globally, currently they make up 95% of New Zealand’s forest estate.
One of the most promising approaches to the production of lignocellulosic biofuels is the sugar pathway. This pathway involves using enzymes to hydrolyse the carbohydrate polymers in the biomass to monomeric sugars, and then converting these sugars to ethanol. High sugar yields during enzymatic hydrolysis requires an effective pretreatment to disrupt and/or remove some of the lignin and hemicelluloses, so that the cellulose is more accessible to enzymes. Softwoods, such as Pinus radiata, are amongst the most recalcitrant substrates towards enzymatic hydrolysis, typically, they require more severe pretreatment conditions and higher enzyme doses than hardwood or agricultural residues.
Steam pretreatment is one simple and cost-effective pretreatment. Steam pretreatment involves heating the biomass in steam or water (150ºC to 250ºC), often in the presence of added acid catalysts. One disadvantage of steam pretreatment is that it can produce compounds that inhibit the subsequent enzymatic hydrolysis.
During enzymatic hydrolysis, these inhibitors, particularly lignin, lower the digestibility of the cellulose by non-productively binding the hydrolysis enzymes. This means that high enzyme doses are required increasing the conversion costs.
How these inhibitors are formed, how they work and what types of inhibitors are most deleterious during enzymatic hydrolysis is not well understood. The main objective of this thesis was to understand the relative roles of soluble and insoluble fibre components as inhibitors of enzyme hydrolysis of steam pretreated P. radiata as a function of pretreatment severity.
Samples: P. radiata sawdust was pretreated using steam explosion under six different pretreatment conditions and separated to produce both water-soluble (filtrate) and insoluble (substrate) samples. These samples were used for the rest of the research.

Insoluble inhibition: The digestibility of the insoluble substrates was examined using a commercial enzyme cocktail (Novozymes Cellic® CTec2). Results showed the digestibility of these substrates increased with increasing pretreatment severity due to increases in accessibility. However, when the substrates were tested after ball-milling to a common cellulose accessibility, as determined by Simons’ stain measurements, the digestibility decreased with increasing pretreatment severity. This showed that while increasing pretreatment severity led to greater enzyme inhibition, the inhibition was more than compensated for by increases in the accessibility.
As part of this work, modifications were made to the Simons’ stain method to ensure a robust and reliable method was available for measuring cellulose accessibility.
Soluble inhibition: The inhibitory effects of filtrates were investigated by comparing changes in the digestibility of both bleached kraft pulp and the insoluble steam pretreated substrates in the presence and absence of filtrates. The results showed that the inhibitory effects of the filtrates were minimal or non-existent under most conditions. It was therefore concluded that the insoluble components of steam pretreated P. radiata were more inhibitory than soluble components.
With the commercial enzyme cocktail CTec2, adding the filtrates back during hydrolysis led to small enhancements in digestibility. The results of subsequent experiments were consistent with the enhancements being due to components in the filtrates acting as reductants for the oxidative cellulase enzymes present in this cocktail.
Inhibition by lignin: Lignins were isolated from three insoluble substrates of differing pretreatment severity and their inhibitory effects on the digestibility of bleached kraft pulp were determined. Results showed: (i) that the inhibitory effects of added lignins increased with the amount of lignin added; (ii) that more severe pretreatments led to more inhibitory lignins; (iii) that adding the surfactant polyethylene glycol could overcome the inhibitory effects of these lignins. Additionally, the more severe pretreatments produced lignins with greater condensation and more phenolic groups, these in turn negatively correlated to digestibility, indicating the structural composition of lignin plays a role in the extent to which lignin is inhibitory.

text: Understanding the Relative Importance of the Effects of Soluble and Insoluble Inhibitors on the Enzyme Digestibility of Steam Pretreated Radiata Pine Lignocellulosic biomass use is one of the keys to growth and development of future biofuels and biochemicals. Amongst the lignocellulosic feedstocks, softwoods are particularly attractive as they have the potential to produce high sugar yields and are prevalent in many forests globally, currently they make up 95% of New Zealand’s forest estate.
One of the most promising approaches to the production of lignocellulosic biofuels is the sugar pathway. This pathway involves using enzymes to hydrolyse the carbohydrate polymers in the biomass to monomeric sugars, and then converting these sugars to ethanol. High sugar yields during enzymatic hydrolysis requires an effective pretreatment to disrupt and/or remove some of the lignin and hemicelluloses, so that the cellulose is more accessible to enzymes. Softwoods, such as Pinus radiata, are amongst the most recalcitrant substrates towards enzymatic hydrolysis, typically, they require more severe pretreatment conditions and higher enzyme doses than hardwood or agricultural residues.
Steam pretreatment is one simple and cost-effective pretreatment. Steam pretreatment involves heating the biomass in steam or water (150ºC to 250ºC), often in the presence of added acid catalysts. One disadvantage of steam pretreatment is that it can produce compounds that inhibit the subsequent enzymatic hydrolysis.
During enzymatic hydrolysis, these inhibitors, particularly lignin, lower the digestibility of the cellulose by non-productively binding the hydrolysis enzymes. This means that high enzyme doses are required increasing the conversion costs.
How these inhibitors are formed, how they work and what types of inhibitors are most deleterious during enzymatic hydrolysis is not well understood. The main objective of this thesis was to understand the relative roles of soluble and insoluble fibre components as inhibitors of enzyme hydrolysis of steam pretreated P. radiata as a function of pretreatment severity.
Samples: P. radiata sawdust was pretreated using steam explosion under six different pretreatment conditions and separated to produce both water-soluble (filtrate) and insoluble (substrate) samples. These samples were used for the rest of the research.

Insoluble inhibition: The digestibility of the insoluble substrates was examined using a commercial enzyme cocktail (Novozymes Cellic® CTec2). Results showed the digestibility of these substrates increased with increasing pretreatment severity due to increases in accessibility. However, when the substrates were tested after ball-milling to a common cellulose accessibility, as determined by Simons’ stain measurements, the digestibility decreased with increasing pretreatment severity. This showed that while increasing pretreatment severity led to greater enzyme inhibition, the inhibition was more than compensated for by increases in the accessibility.
As part of this work, modifications were made to the Simons’ stain method to ensure a robust and reliable method was available for measuring cellulose accessibility.
Soluble inhibition: The inhibitory effects of filtrates were investigated by comparing changes in the digestibility of both bleached kraft pulp and the insoluble steam pretreated substrates in the presence and absence of filtrates. The results showed that the inhibitory effects of the filtrates were minimal or non-existent under most conditions. It was therefore concluded that the insoluble components of steam pretreated P. radiata were more inhibitory than soluble components.
With the commercial enzyme cocktail CTec2, adding the filtrates back during hydrolysis led to small enhancements in digestibility. The results of subsequent experiments were consistent with the enhancements being due to components in the filtrates acting as reductants for the oxidative cellulase enzymes present in this cocktail.
Inhibition by lignin: Lignins were isolated from three insoluble substrates of differing pretreatment severity and their inhibitory effects on the digestibility of bleached kraft pulp were determined. Results showed: (i) that the inhibitory effects of added lignins increased with the amount of lignin added; (ii) that more severe pretreatments led to more inhibitory lignins; (iii) that adding the surfactant polyethylene glycol could overcome the inhibitory effects of these lignins. Additionally, the more severe pretreatments produced lignins with greater condensation and more phenolic groups, these in turn negatively correlated to digestibility, indicating the structural composition of lignin plays a role in the extent to which lignin is inhibitory.

year: 2018

title: Taking the road less travelled: Exploring the link between curiosity and resilience with 1NZSAS

authors: Bojilova, Alexandrina

subjects: Resilience

abstract: The pivotal question- ‘what goes right?’ in resilience must be explored further. We need to reconstruct resilience- beyond recovery or equanimity and towards capaci-ty for learning and thriving. This research unveils curiosity as a vital enabler for resilience. The relationship is studied in the context of resilience as a holistic, multifaceted and developable capability.
The primary aim of this research is to understand ‘what goes right’ for individuals who perform and thrive in ongoing change, ambiguity and significant risks, and to study the contextual conditions enabling their resilience. The overarching re-search question asks: “How is resilience built, sustained and developed to support capacity for thriving?” or put simply, “What goes right?”. The three supporting questions are “In what ways does curiosity serve as a resilience enabler? “What are the heuristics individuals employ to sustain and grow resilient mindsets?”, and “What are the contextual conditions that support access to resilience?”.
This research employs the principles of social constructivism as a methodological framework. It uses semi-structured interviews and narratives to capture detailed insights from 35 New Zealand Special Air Service (1NZSAS) members, for whom resilience is a critical value-in-action. 
The present research extends understanding of and applied ways to resilience, by reconstructing it through the prisms of curiosity. It frames and explores the heu-ristics individuals nurture and rely upon to thrive in and grow through challenges and setbacks. This research also outlines the contextual conditions that enable capacity for resilience in some of the most demanding conditions. The present research presents an integrated model of the findings, using curiosity and resili-ence as the axis that shape, drive and sustain capacity for thriving in ambiguity and change. Further to academic contribution, this thesis offers invaluable contri-butions to practitioners and leaders, as well as individuals willing to broaden and build greater access to resilience.

text: Taking the road less travelled: Exploring the link between curiosity and resilience with 1NZSAS The pivotal question- ‘what goes right?’ in resilience must be explored further. We need to reconstruct resilience- beyond recovery or equanimity and towards capaci-ty for learning and thriving. This research unveils curiosity as a vital enabler for resilience. The relationship is studied in the context of resilience as a holistic, multifaceted and developable capability.
The primary aim of this research is to understand ‘what goes right’ for individuals who perform and thrive in ongoing change, ambiguity and significant risks, and to study the contextual conditions enabling their resilience. The overarching re-search question asks: “How is resilience built, sustained and developed to support capacity for thriving?” or put simply, “What goes right?”. The three supporting questions are “In what ways does curiosity serve as a resilience enabler? “What are the heuristics individuals employ to sustain and grow resilient mindsets?”, and “What are the contextual conditions that support access to resilience?”.
This research employs the principles of social constructivism as a methodological framework. It uses semi-structured interviews and narratives to capture detailed insights from 35 New Zealand Special Air Service (1NZSAS) members, for whom resilience is a critical value-in-action. 
The present research extends understanding of and applied ways to resilience, by reconstructing it through the prisms of curiosity. It frames and explores the heu-ristics individuals nurture and rely upon to thrive in and grow through challenges and setbacks. This research also outlines the contextual conditions that enable capacity for resilience in some of the most demanding conditions. The present research presents an integrated model of the findings, using curiosity and resili-ence as the axis that shape, drive and sustain capacity for thriving in ambiguity and change. Further to academic contribution, this thesis offers invaluable contri-butions to practitioners and leaders, as well as individuals willing to broaden and build greater access to resilience.

year: 2021

title: The COVID-19 domestic vaccine pass: Implications for Māori

authors: Kukutai, Tahu

abstract: The New Zealand government has introduced a COVID-19 domestic vaccine pass to be used in conjunction with the COVID-19 Protection Framework. The Framework is likely to be activated soon after Cabinet meets on 29 November 2021. The pass will be necessary to access places and events that require proof of vaccination under the Framework.
This brief does not argue for or against the introduction of a domestic vaccine pass, but rather discusses key issues that it raises for Māori, and suggests actions to address them. We see four key issues:
• the lack of Māori involvement, as a Tiriti partner, on either the design or implementation of the COVID-19 domestic vaccine pass;
• disproportionate restriction on Māori mobility due to lower Māori vaccination rates;
• privacy and data security concerns;
• uneven implementation that could increase discrimination against Māori and other groups considered to pose a risk to others’ safety.
To respond to these issues we recommend that the implementation of the pass be designed in partnership with Māori and comply with Māori data sovereignty requirements. As Tiriti partners, Māori should expect that the pass will keep their communities safe, while providing opportunities to enact manaakitanga in the matrix of care, and the mana to manage their own affairs.

text: The COVID-19 domestic vaccine pass: Implications for Māori The New Zealand government has introduced a COVID-19 domestic vaccine pass to be used in conjunction with the COVID-19 Protection Framework. The Framework is likely to be activated soon after Cabinet meets on 29 November 2021. The pass will be necessary to access places and events that require proof of vaccination under the Framework.
This brief does not argue for or against the introduction of a domestic vaccine pass, but rather discusses key issues that it raises for Māori, and suggests actions to address them. We see four key issues:
• the lack of Māori involvement, as a Tiriti partner, on either the design or implementation of the COVID-19 domestic vaccine pass;
• disproportionate restriction on Māori mobility due to lower Māori vaccination rates;
• privacy and data security concerns;
• uneven implementation that could increase discrimination against Māori and other groups considered to pose a risk to others’ safety.
To respond to these issues we recommend that the implementation of the pass be designed in partnership with Māori and comply with Māori data sovereignty requirements. As Tiriti partners, Māori should expect that the pass will keep their communities safe, while providing opportunities to enact manaakitanga in the matrix of care, and the mana to manage their own affairs.

year: 2021-11-30