title: Oxidative Decolouring of Bloodmeal Using Peracetic Acid

authors: Hicks, Talia

subjects: Bloodmeal

abstract: Bloodmeal is an ancillary product of the agricultural industry, containing 80 – 100 % protein, and has been used to create a bio-based plastic called Novatein Thermoplastic Protein®, with a proprietary blend of sodium sulfite, sodium dodecyl sulfate (SDS), water and triethylene glycol (TEG). Recent investigation has shown that the colour and odour of bloodmeal based thermoplastic can be significantly improved through the pre-treatment of bloodmeal with equilibrium peracetic acid. The resulting decoloured bloodmeal (DBM) no longer relies on the addition of sodium sulfite to be processed in conventional thermoplastic equipment. The purpose of this study was to gain a comprehensive understanding of the role by which peracetic acid decolours bloodmeal and the consequences of oxidation on composition, physico-chemical properties, protein structure and interactions and final polymer mobility.
Oxidative decolouring of haem is straightforward when the haem is freely accessible or in the form of oxyhaemoglobin and most common oxidants lead to cleavage of the porphyrin ring, resulting in a loss of colour saturation. However, after extensive thermal treatment during manufacture, the haem present in bloodmeal has migrated to inaccessible hydrophobic regions of the protein aggregates or has been oxidised to the form of methaemoglobin. Literature revealed that methaemoglobin catalytically removes hydrogen peroxide, lowering decolouring efficacy. Thus, peracetic acid is required when haem is present in the form of methaemoglobin, as is found in bloodmeal. Further, it was confirmed that a minimum of 3% w/w peracetic acid, in a 3:1 ratio to bloodmeal is required to obtain adequate decolouring (> 70% whiteness based on the RGB colour space).
The role of each component in equilibrium peracetic acid was established; water and acetic acid resulted in protein swelling but were found unlikely to influence accessibility of the oxidants to the haem sites and both hydrogen peroxide and peracetic acid were consumed in oxidising reactions. Acidic conditions were found to reduce the consumption of hydrogen peroxide, possibly through the inhibition of hydroxyl radical formation, resulting in less effective decolouring. Acetic acid was observed to have a protective effect on protein recovery as well as the resulting iron content, molecular mass distribution and secondary structure of protein.
Due to the large excess of peracetic acid solution required to facilitate diffusion and subsequent decolouring, a significant quantity remains unreacted in the recovered wastewater. Unreacted peracetic acid was shown to have potential for recycling, although in its immediate state it was insufficient to decolour fresh bloodmeal.
The protein-rich decoloured bloodmeal recovered had undergone a change in secondary structure composition and was found to contain evidence of a diffusion front, consistent with the heterogeneous phase decolouring mechanism. Oxidation with peracetic acid was found to result in less β-sheet aggregation compared to hydrogen peroxide treatment, although all treatments resulted in an increase in the quantity of disordered structures. Oxidation was also found to result in a significantly lowered glass transition temperature and along with an increased enthalpy of relaxation, evidenced a large improvement in polymer chain mobility compared with untreated bloodmeal.
DBM was found to be comprised of 90 – 99% w/w protein (1 – 10% w/w salt), with a significantly higher protein solubility and a similar volume weighted molecular mass compared with untreated bloodmeal. Further, DBM was found to have a greater composition of charged and polar amino acids, along with a large reduction in lysine and small reduction in aromatic and heterocyclic amino acids. Additionally, cystine crosslinks which stabilise bloodmeal were found to be partially oxidised to cysteine sulfonate compounds and cysteic acid (cleavage of the disulfide bond). Such a reduction in the quantity of amino acids which are capable of forming covalent networks (lysine, tyrosine and cysteine) support prior evidence that DBM no longer requires sodium sulfite to produce a thermoplastic.
The influence of SDS and TEG on secondary structure and chain mobility at ambient and elevated temperatures were explored. It was found that heating DBM and DBM with SDS alone is incapable of providing sufficient energy to induce mobility and chain rearrangement. However, the addition of TEG was found to facilitate chain mobility, and beyond 55 oC significant changes to the secondary structure composition was observed, first through the formation of α-helices and finally through the aggregation of chains into β-sheets. Wide angle X-ray scattering confirmed that the changes in ordered structures composition were reversible in DBM which contained both SDS and TEG.
The ability of TEG to plasticise DBM was more thoroughly explored, and it was found that prior to heating TEG was localised into regions either plasticiser-rich or -poor, and this resulted in the presence of two glass transition temperatures. After heating, the TEG was more homogeneously distributed, which resulted in the presence of one broad glass transition region.

text: Oxidative Decolouring of Bloodmeal Using Peracetic Acid Bloodmeal is an ancillary product of the agricultural industry, containing 80 – 100 % protein, and has been used to create a bio-based plastic called Novatein Thermoplastic Protein®, with a proprietary blend of sodium sulfite, sodium dodecyl sulfate (SDS), water and triethylene glycol (TEG). Recent investigation has shown that the colour and odour of bloodmeal based thermoplastic can be significantly improved through the pre-treatment of bloodmeal with equilibrium peracetic acid. The resulting decoloured bloodmeal (DBM) no longer relies on the addition of sodium sulfite to be processed in conventional thermoplastic equipment. The purpose of this study was to gain a comprehensive understanding of the role by which peracetic acid decolours bloodmeal and the consequences of oxidation on composition, physico-chemical properties, protein structure and interactions and final polymer mobility.
Oxidative decolouring of haem is straightforward when the haem is freely accessible or in the form of oxyhaemoglobin and most common oxidants lead to cleavage of the porphyrin ring, resulting in a loss of colour saturation. However, after extensive thermal treatment during manufacture, the haem present in bloodmeal has migrated to inaccessible hydrophobic regions of the protein aggregates or has been oxidised to the form of methaemoglobin. Literature revealed that methaemoglobin catalytically removes hydrogen peroxide, lowering decolouring efficacy. Thus, peracetic acid is required when haem is present in the form of methaemoglobin, as is found in bloodmeal. Further, it was confirmed that a minimum of 3% w/w peracetic acid, in a 3:1 ratio to bloodmeal is required to obtain adequate decolouring (> 70% whiteness based on the RGB colour space).
The role of each component in equilibrium peracetic acid was established; water and acetic acid resulted in protein swelling but were found unlikely to influence accessibility of the oxidants to the haem sites and both hydrogen peroxide and peracetic acid were consumed in oxidising reactions. Acidic conditions were found to reduce the consumption of hydrogen peroxide, possibly through the inhibition of hydroxyl radical formation, resulting in less effective decolouring. Acetic acid was observed to have a protective effect on protein recovery as well as the resulting iron content, molecular mass distribution and secondary structure of protein.
Due to the large excess of peracetic acid solution required to facilitate diffusion and subsequent decolouring, a significant quantity remains unreacted in the recovered wastewater. Unreacted peracetic acid was shown to have potential for recycling, although in its immediate state it was insufficient to decolour fresh bloodmeal.
The protein-rich decoloured bloodmeal recovered had undergone a change in secondary structure composition and was found to contain evidence of a diffusion front, consistent with the heterogeneous phase decolouring mechanism. Oxidation with peracetic acid was found to result in less β-sheet aggregation compared to hydrogen peroxide treatment, although all treatments resulted in an increase in the quantity of disordered structures. Oxidation was also found to result in a significantly lowered glass transition temperature and along with an increased enthalpy of relaxation, evidenced a large improvement in polymer chain mobility compared with untreated bloodmeal.
DBM was found to be comprised of 90 – 99% w/w protein (1 – 10% w/w salt), with a significantly higher protein solubility and a similar volume weighted molecular mass compared with untreated bloodmeal. Further, DBM was found to have a greater composition of charged and polar amino acids, along with a large reduction in lysine and small reduction in aromatic and heterocyclic amino acids. Additionally, cystine crosslinks which stabilise bloodmeal were found to be partially oxidised to cysteine sulfonate compounds and cysteic acid (cleavage of the disulfide bond). Such a reduction in the quantity of amino acids which are capable of forming covalent networks (lysine, tyrosine and cysteine) support prior evidence that DBM no longer requires sodium sulfite to produce a thermoplastic.
The influence of SDS and TEG on secondary structure and chain mobility at ambient and elevated temperatures were explored. It was found that heating DBM and DBM with SDS alone is incapable of providing sufficient energy to induce mobility and chain rearrangement. However, the addition of TEG was found to facilitate chain mobility, and beyond 55 oC significant changes to the secondary structure composition was observed, first through the formation of α-helices and finally through the aggregation of chains into β-sheets. Wide angle X-ray scattering confirmed that the changes in ordered structures composition were reversible in DBM which contained both SDS and TEG.
The ability of TEG to plasticise DBM was more thoroughly explored, and it was found that prior to heating TEG was localised into regions either plasticiser-rich or -poor, and this resulted in the presence of two glass transition temperatures. After heating, the TEG was more homogeneously distributed, which resulted in the presence of one broad glass transition region.

year: 2015

title: Measuring the bulk impedance of brain tissue in vitro

authors: Wilson, Marcus T.

abstract: Theoretical and numerical models of brain activity suggest a link between seizures and electrical connectivity. We have therefore been motivated to measure electrical conductivity in brain tissue. Such measurements in vitro are difficult; it is necessary to use a conductive inorganic salt solution, artificial cerebrospinal fluid (ACSF), to keep the tissue alive. We have attempted to provide a robust method to make such measurements. Mouse brain tissue was sliced (400 μm) using established methods. Half the slices were placed in standard ACSF; half were placed in ACSF devoid of magnesium ions. The latter case promotes seizure activity. Electrical activity was measured with a tungsten electrode at various places on the slices. Sixty-nine samples of cortex (2 mm × 2 mm) were cut with a razor. Their areas were measured with a calibrated microscope. Each sample was placed between two flat Ag/AgCl electrodes in a Perspex sandwich. Excess ACSF was removed with filter paper. The impedance was measured at 25°C from 20 Hz to 2 MHz with an Agilent E4980A four-point impedance meter in a shielded room, using a low current. Between 1 kHz and 100 kHz the conductivity was approximately 0.2 S m⁻¹; outside this range dispersion occurred. Samples prepared in the magnesium-free ACSF had a conductivity about 10% lower. The Cole-Cole model of conductivity was fitted. There were few significant differences between the parameters for the different groups measured.

text: Measuring the bulk impedance of brain tissue in vitro Theoretical and numerical models of brain activity suggest a link between seizures and electrical connectivity. We have therefore been motivated to measure electrical conductivity in brain tissue. Such measurements in vitro are difficult; it is necessary to use a conductive inorganic salt solution, artificial cerebrospinal fluid (ACSF), to keep the tissue alive. We have attempted to provide a robust method to make such measurements. Mouse brain tissue was sliced (400 μm) using established methods. Half the slices were placed in standard ACSF; half were placed in ACSF devoid of magnesium ions. The latter case promotes seizure activity. Electrical activity was measured with a tungsten electrode at various places on the slices. Sixty-nine samples of cortex (2 mm × 2 mm) were cut with a razor. Their areas were measured with a calibrated microscope. Each sample was placed between two flat Ag/AgCl electrodes in a Perspex sandwich. Excess ACSF was removed with filter paper. The impedance was measured at 25°C from 20 Hz to 2 MHz with an Agilent E4980A four-point impedance meter in a shielded room, using a low current. Between 1 kHz and 100 kHz the conductivity was approximately 0.2 S m⁻¹; outside this range dispersion occurred. Samples prepared in the magnesium-free ACSF had a conductivity about 10% lower. The Cole-Cole model of conductivity was fitted. There were few significant differences between the parameters for the different groups measured.

year: 2015

title: Te Karauna Hou: the senior Ngati Rahiri rangatira

authors: Hart, Philip

abstract: Te Karauna Hou, the principal Ngati Rahiri rangatira living at Te Aroha in 1880, had a distinguished whakapapa linking him to several hapu. Before settling permanently at Te Aroha in the 1870s he lived in several places, especially at Kaitawa, on the southern outskirts of Thames. One of the principal rangatira in Hauraki, he was loyal to the Crown during the Waikato War, and later assisted Pakeha settlement. To emphasize his mana and that of his hapu, he held big festivities at Kaitawa and at Omahu pa at Te Aroha. 
In 1871, when Ngati Haua won a (temporary) victory in the land court over the ownership of the Aroha Block, Karauna took control of it on behalf of the Marutuahu confederation, and subsequently kept Ngati Haua at bay. For a time he opposed road-making on this block, but later agreed to it, for financial reasons. Like all rangatira, he sought to maximize his ownership of as many blocks of land as possible, sometimes having his lies exposed through his contradicting his earlier evidence. He also denied receiving money for land when his denials were easily disproved. Despite leasing and selling land, selling timber and gum, and opening his land to miners, he often struggled financially.  
Karauna claimed to have found gold in Hauraki in 1852, and was willing to open his land at Thames to miners, but did not invest in any claims before the Te Aroha rush, when he attempted to extract a bonus of £1,000 from the government for opening the field. After his death in 1885, Pakeha remembered with gratitude his friendly attitude to them, but they also remembered his drunkenness, which meant he lost the respect of both Maori and Pakeha in his latter years.

text: Te Karauna Hou: the senior Ngati Rahiri rangatira Te Karauna Hou, the principal Ngati Rahiri rangatira living at Te Aroha in 1880, had a distinguished whakapapa linking him to several hapu. Before settling permanently at Te Aroha in the 1870s he lived in several places, especially at Kaitawa, on the southern outskirts of Thames. One of the principal rangatira in Hauraki, he was loyal to the Crown during the Waikato War, and later assisted Pakeha settlement. To emphasize his mana and that of his hapu, he held big festivities at Kaitawa and at Omahu pa at Te Aroha. 
In 1871, when Ngati Haua won a (temporary) victory in the land court over the ownership of the Aroha Block, Karauna took control of it on behalf of the Marutuahu confederation, and subsequently kept Ngati Haua at bay. For a time he opposed road-making on this block, but later agreed to it, for financial reasons. Like all rangatira, he sought to maximize his ownership of as many blocks of land as possible, sometimes having his lies exposed through his contradicting his earlier evidence. He also denied receiving money for land when his denials were easily disproved. Despite leasing and selling land, selling timber and gum, and opening his land to miners, he often struggled financially.  
Karauna claimed to have found gold in Hauraki in 1852, and was willing to open his land at Thames to miners, but did not invest in any claims before the Te Aroha rush, when he attempted to extract a bonus of £1,000 from the government for opening the field. After his death in 1885, Pakeha remembered with gratitude his friendly attitude to them, but they also remembered his drunkenness, which meant he lost the respect of both Maori and Pakeha in his latter years.

year: 2016