De-risking Pretreatment of Microalgae To Produce Fuels and Chemical Co-products.

Conversion of microalgae to renewable fuels and chemical co-products by pretreating and fractionation holds promise as an algal biorefinery concept, but a better understanding of the pretreatment performance as a function of algae strain and composition is necessary to de-risk algae conversion operations. Similarly, there are few examples of algae pretreatment at scales larger than the bench scale. This work aims to de-risk algal biorefinery operations by evaluating the pretreatment performance across nine different microalgae samples and five different pretreatment methods at small (5 mL) scale and further de-risk the operation by scaling pretreatment for one species to the 80 L scale. The pretreatment performance was evaluated by solubilization of feedstock carbon and nitrogen [as total organic carbon (TOC) and total nitrogen (TN)] into the aqueous hydrolysate and extractability of lipids [as fatty acid methyl esters (FAMEs)] from the pretreated solids. A range of responses was noted among the algae samples across pretreatments, with the current dilute Brønsted acid pretreatment using H2SO4 being the most consistent and robust. This pretreatment produced TOC yields to the hydrolysate ranging from 27.7 to 51.1%, TN yields ranging from 12.3 to 76.2%, and FAME yields ranging from 57.9 to 89.9%. In contrast, the other explored pretreatments (other dilute acid pretreatments, dilute alkali pretreatment with NaOH, enzymatic pretreatment, and flash hydrolysis) produced lower or more variable yields across the three metrics. In light of the greater consistency across samples for dilute acid pretreatment, this method was scaled to 80 L to demonstrate scalability with microalgae feedstocks.

Feedstock/pretreatment screening for bioconversion of sugar and lignin streams via deacetylated disc-refining.

Recent publications have shown the benefits of deacetylation disc-refining (DDR) as a pretreatment process to deconstruct biomass into sugars and lignin residues. Major advantages of DDR pretreatment over steam and dilute acid pretreatment are the removal of acetyl and lignin during deacetylation. DDR does not generate hydroxymethylfurfural (HMF) and furfural which are commonly produced from steam and dilute acid pretreatments. Acetate, lignin, HMF, and furfural are known inhibitors during enzymatic hydrolysis and fermentation. Another advantage of deacetylation is the production of lignin-rich black liquor, which can be upgraded to other bioproducts. Furthermore, due to the lack of sugar degradation during deacetylation, DDR has significantly less sugar loss than other pretreatment methods. Previous studies for DDR have primarily focused on corn stover, but lacked the investigative studies of other feedstocks. This study was designed to screen various DDR process conditions at pilot scale using three different feedstocks, including corn stover, poplar, and switchgrass. The impact of the pretreatment conditions was evaluated by testing hydrolysates for bioconversion to 2,3-butanediol. Pretreatment of biomass by DDR showed high-conversion-yields and 2,3-BDO fermentation production yields. Techno-economic analysis (TEA) of the pretreatment for biomass to sugar was also developed based on NREL's Aspen Model. This study shows that the cellulose and hemicellulose in poplar was more recalcitrant than herbaceous feedstocks which ultimately drove up the sugar cost. Switchgrass was also more recalcitrant than corn stover but less than poplar.

Aspects of occupational safety: a survey among European cancer nurses.

PURPOSE: Nurses are particularly at risk for occupational exposure to hazardous cancer drugs, risking both acute and chronic health effects. Knowledge on the implemented safety precautions into minimizing these risks is limited. METHODS: The European Cancer Nursing Index (ECNI) was developed by the European Oncology Nursing Society (EONS) to illustrate the development and status of this profession. In this study, anonymous online survey data on occupational safety reported by European cancer nurses as part of the ECNI 2022, was analysed. RESULTS: A total of 630 cancer nurses from 29 countries responded to the survey. A majority reported that written guidelines (n = 553, 88%) on safe handling and administration of hazardous drugs, personal protection equipment (PPE) and cytotoxic spillage kits (n = 514, 82%) were available at their workplaces. 130 (21%) nurses reported that wipe testing to assess any residual hazardous drugs on workplace surfaces were conducted systematically at their workplaces. 185 (29%) nurses reported that nurses sometimes or always continued with their regular tasks (including handling hazardous cancer drugs) during pregnancy and breast feeding. 185 (29%) also responded that nurses at their workplaces did not receive an introductory education program before handling hazardous drugs. In total, 346 (55%) of the nurses reported that their workplace had a freedom to speak-up guardian or whistle blower policy for members of staff. CONCLUSIONS: Even if most nurses report that there are safety routines in place at their workplaces, the results reveal several serious occupational risks for European nurses handling hazardous cancer drugs. Actions are needed to improve and optimize occupational safety for nursing staff.

Nursing care and management of adverse events for patients with BRAFV600E-mutant metastatic colorectal cancer receiving encorafenib in combination with cetuximab: a review.

Encorafenib is a B-Raf proto-oncogene serine/threonine-protein kinase (BRAF) inhibitor, approved in the EU and USA, in combination with the epidermal growth factor receptor (EGFR) inhibitor cetuximab, for the treatment of patients with BRAFV600E-mutant metastatic colorectal cancer (mCRC). In the pivotal BEACON CRC trial, patients achieved longer survival with encorafenib in combination with cetuximab vs. conventional chemotherapy. This targeted therapy regimen is also generally better tolerated than cytotoxic treatments. However, patients may present with adverse events unique to the regimen and characteristic of BRAF and EGFR inhibitors, which produce their own set of challenges. Nurses play an essential role in navigating the care of patients with BRAFV600E-mutant mCRC and managing adverse events that patients may experience. This includes early and efficient identification of treatment-related adverse events, subsequent management of adverse events and education of patients and their caregivers around key adverse events. This manuscript aims to provide support to nurses managing patients with BRAFV600E-mutant mCRC receiving encorafenib in combination with cetuximab, by summarising potential adverse events and providing guidance on how to manage them. Special attention will be paid to the presentation of key adverse events, dose modifications that may be required, practical recommendations and supportive care measures.

The impact of climate change on cancer nursing in Palestine.

Cancer is the third leading cause of death in Palestine, with many cancers diagnosed at a late stage. In contrast to the developed world, two thirds of cancer diagnoses occur between the ages of 15 and 64, moreover, 10% of all cancer diagnoses occur in children under the age of 10 (compared to 0.05% of all new cancer diagnoses in the UK). Cancer nursing as a speciality in Palestine is newly established in the last 5 years; partly helped by the introduction of the Higher Diploma in Cancer and Palliative Care Nursing, and more recently the delivery of the first intake of the Master of Science in Cancer and Palliative Care Nursing at Bethlehem University. There are many challenges faced by cancer patients and nurses in Palestine; there is only one facility in the West Bank that delivers radiotherapy, 2 PET-CT scanners for the whole of the West Bank, with no PET-CT or radiotherapy facilities in Gaza. There are 2 haematology units in the West Bank that perform autologous stem cell transplants for adults and any haematology patient (adult or child) requiring an allogeneic stem cell/bone marrow transplant has to be referred to neighbouring Israel or Jordan. Climate change might have both a direct and indirect impact on the growth of cancers and on cancer treatment and oncology nurses. Over the last 150 years the planet has warmed by over one degree Celsius resulting in disastrous consequences for the environment. Nurses make up the largest number of the healthcare workforce and are ideally placed to have a positive impact on the global warming crisis due to their leadership roles as well as their work in health promotion. They equally do a lot to help cancer patients to deal with its effects and often care for patients from marginalised groups. It is important for nurses to take the lead and move immediately to make health systems more resistant to climate change.

Mirvetuximab Soravtansine Combination Yields Encouraging Response Rates in Ovarian Cancer.

Mirvetuximab soravtansine in combination with bevacizumab (Avastin) to treat patients with platinum-agnostic ovarian cancer demonstrated a confirmed 64% overall response rate (ORR), regardless of platinum status, according to study results presented at the 2020 American Society of Clinical Oncology Virtual Scientific Program.

Salmon blood plasma: effective inhibitor of protease-laden Pacific whiting surimi and salmon mince.

The effect of salmon plasma (SP) from Chinook salmon on proteolytic inhibition was investigated. SP was found to inhibit both cysteine and serine proteases as well as protease extracted from Pacific whiting muscle. SP was found to contain a 55kDa cysteine protease inhibitor through SDS-PAGE inhibitor staining. Freeze dried salmon plasma (FSP) and salmon plasma concentrated by ultrafiltration (CSP) were tested for their ability to inhibit autolysis in Pacific whiting surimi and salmon mince at concentrations of 0.25%, 0.5%, 1%, and 2%. Pacific whiting surimi autolysis was inhibited by an average of 89% regardless of concentration while inhibition of salmon mince autolysis increased with concentration (p<0.05). CSP performed slightly better than FSP at inhibiting salmon mince autolysis (p<0.05). Serine protease inhibition decreased when SP heated above 40°C but was stable across a broad NaCl and pH range. Cysteine protease inhibitors exhibited good temperature, NaCl, and pH stability.

Genetic Determinants for Enzymatic Digestion of Lignocellulosic Biomass Are Independent of Those for Lignin Abundance in a Maize Recombinant Inbred Population.

Biotechnological approaches to reduce or modify lignin in biomass crops are predicated on the assumption that it is the principal determinant of the recalcitrance of biomass to enzymatic digestion for biofuels production. We defined quantitative trait loci (QTL) in the Intermated B73 × Mo17 recombinant inbred maize (Zea mays) population using pyrolysis molecular-beam mass spectrometry to establish stem lignin content and an enzymatic hydrolysis assay to measure glucose and xylose yield. Among five multiyear QTL for lignin abundance, two for 4-vinylphenol abundance, and four for glucose and/or xylose yield, not a single QTL for aromatic abundance and sugar yield was shared. A genome-wide association study for lignin abundance and sugar yield of the 282-member maize association panel provided candidate genes in the 11 QTL of the B73 and Mo17 parents but showed that many other alleles impacting these traits exist among this broader pool of maize genetic diversity. B73 and Mo17 genotypes exhibited large differences in gene expression in developing stem tissues independent of allelic variation. Combining these complementary genetic approaches provides a narrowed list of candidate genes. A cluster of SCARECROW-LIKE9 and SCARECROW-LIKE14 transcription factor genes provides exceptionally strong candidate genes emerging from the genome-wide association study. In addition to these and genes associated with cell wall metabolism, candidates include several other transcription factors associated with vascularization and fiber formation and components of cellular signaling pathways. These results provide new insights and strategies beyond the modification of lignin to enhance yields of biofuels from genetically modified biomass.

Crystal structures of acetate kinases from the eukaryotic pathogens Entamoeba histolytica and Cryptococcus neoformans.

Acetate kinases (ACKs) are members of the acetate and sugar kinase/hsp70/actin (ASKHA) superfamily and catalyze the reversible phosphorylation of acetate, with ADP/ATP the most common phosphoryl acceptor/donor. While prokaryotic ACKs have been the subject of extensive biochemical and structural characterization, there is a comparative paucity of information on eukaryotic ACKs, and prior to this report, no structure of an ACK of eukaryotic origin was available. We determined the structures of ACKs from the eukaryotic pathogens Entamoeba histolytica and Cryptococcus neoformans. Each active site is located at an interdomain interface, and the acetate and phosphate binding pockets display sequence and structural conservation with their prokaryotic counterparts. Interestingly, the E. histolytica ACK has previously been shown to be pyrophosphate (PP(i))-dependent, and is the first ACK demonstrated to have this property. Examination of its structure demonstrates how subtle amino acid substitutions within the active site have converted cosubstrate specificity from ATP to PP(i) while retaining a similar backbone conformation. Differences in the angle between domains surrounding the active site suggest that interdomain movement may accompany catalysis. Taken together, these structures are consistent with the eukaryotic ACKs following a similar reaction mechanism as is proposed for the prokaryotic homologs.

Novel pyrophosphate-forming acetate kinase from the protist Entamoeba histolytica.

Acetate kinase (ACK) catalyzes the reversible synthesis of acetyl phosphate by transfer of the γ-phosphate of ATP to acetate. Here we report the first biochemical and kinetic characterization of a eukaryotic ACK, that from the protist Entamoeba histolytica. Our characterization revealed that this protist ACK is the only known member of the ASKHA structural superfamily, which includes acetate kinase, hexokinase, and other sugar kinases, to utilize inorganic pyrophosphate (PP(i))/inorganic phosphate (P(i)) as the sole phosphoryl donor/acceptor. Detection of ACK activity in E. histolytica cell extracts in the direction of acetate/PP(i) formation but not in the direction of acetyl phosphate/P(i) formation suggests that the physiological direction of the reaction is toward acetate/PP(i) production. Kinetic parameters determined for each direction of the reaction are consistent with this observation. The E. histolytica PP(i)-forming ACK follows a sequential mechanism, supporting a direct in-line phosphoryl transfer mechanism as previously reported for the well-characterized Methanosarcina thermophila ATP-dependent ACK. Characterizations of enzyme variants altered in the putative acetate/acetyl phosphate binding pocket suggested that acetyl phosphate binding is not mediated solely through a hydrophobic interaction but also through the phosphoryl group, as for the M. thermophila ACK. However, there are key differences in the roles of certain active site residues between the two enzymes. The absence of known ACK partner enzymes raises the possibility that ACK is part of a novel pathway in Entamoeba.

Direct detection of the acetate-forming activity of the enzyme acetate kinase.

Acetate kinase, a member of the acetate and sugar kinase-Hsp70-actin (ASKHA) enzyme superfamily, is responsible for the reversible phosphorylation of acetate to acetyl phosphate utilizing ATP as a substrate. Acetate kinases are ubiquitous in the Bacteria, found in one genus of Archaea, and are also present in microbes of the Eukarya. The most well characterized acetate kinase is that from the methane-producing archaeon Methanosarcina thermophila. An acetate kinase which can only utilize PP(i) but not ATP in the acetyl phosphate-forming direction has been isolated from Entamoeba histolytica, the causative agent of amoebic dysentery, and has thus far only been found in this genus. In the direction of acetyl phosphate formation, acetate kinase activity is typically measured using the hydroxamate assay, first described by Lipmann, a coupled assay in which conversion of ATP to ADP is coupled to oxidation of NADH to NAD(+) by the enzymes pyruvate kinase and lactate dehydrogenase, or an assay measuring release of inorganic phosphate after reaction of the acetyl phosphate product with hydroxylamine. Activity in the opposite, acetate-forming direction is measured by coupling ATP formation from ADP to the reduction of NADP(+) to NADPH by the enzymes hexokinase and glucose 6-phosphate dehydrogenase. Here we describe a method for the detection of acetate kinase activity in the direction of acetate formation that does not require coupling enzymes, but is instead based on direct determination of acetyl phosphate consumption. After the enzymatic reaction, remaining acetyl phosphate is converted to a ferric hydroxamate complex that can be measured spectrophotometrically, as for the hydroxamate assay. Thus, unlike the standard coupled assay for this direction that is dependent on the production of ATP from ADP, this direct assay can be used for acetate kinases that produce ATP or PP(i).

The anti-trypanosomal agent lonidamine inhibits Trypanosoma brucei hexokinase 1.

Glycolysis is essential to the parasitic protozoan Trypanosoma brucei. The first step in this metabolic pathway is mediated by hexokinase, an enzyme that transfers the gamma-phosphate of ATP to a hexose. The T. brucei genome (TREU927/4 GUTat10.1) encodes two hexokinases (TbHK1 and TbHK2) that are 98% identical at the amino acid level. Our previous efforts have revealed that TbHK2 is an important regulator of TbHK1 in procyclic form parasites. Here, we have found through RNAi that TbHK1 is essential to the bloodstream form parasite. Silencing the gene for 4 days reduces cellular hexokinase approximately 60% and leads to parasite death. Additionally, we have found that the recombinant enzyme is inhibited by lonidamine (IC(50)=850 microM), an anti-cancer drug that targets tumor hexokinases. This agent also inhibits HK activity from whole parasite lysate (IC(50)=965 microM). Last, lonidamine is toxic to cultured bloodstream form parasites (LD(50)=50 microM) and procyclic form parasites (LD(50)=180 microM). Interestingly, overexpression of TbHK1 protects PF parasites from lonidamine. These studies provide genetic evidence that TbHK1 is a valid therapeutic target while identifying a potential molecular target of the anti-trypanosomal agent lonidamine.