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The solution properties of two different glycoforms of IgG1 (IgG1Cri and IgG1Wid) are compared using primarily sedimentation equilibrium analysis with two complementary analysis routines: SEDFIT-MSTAR and MULTISIG. IgGCri bears diantennary complex-type glycans on its Fc domain that are fully core fucosylated and partially sialylated, whilst on IgGWid, they are non-fucosylated, partially galactosylated and non-sialylated. IgGWid is also Fab glycosylated. Despite these differences, SEDFIT-MSTAR analysis shows similar weight average molar masses Mw of ~ (150 ± 5) kDa for IgGCri and ~ (154 ± 5) kDa for IgGWid and both glycoforms show evidence of the presence of a small fraction of dimer confirmed by MULTISIG analysis and also by sedimentation coefficient distributions from supportive sedimentation velocity measurements. The closeness of the sedimentation equilibrium behaviour and sedimentation coefficient distributions with a main peak sedimentation coefficient of ~ 6.4S for both glycoforms at different concentrations suggest that the different glycosylation profiles do not significantly impact on molar mass (molecular weight) nor conformation in solution.

Glycopeptide antibiotics are regularly used in ophthalmology to treat infections of Gram-positive bacteria. Aggregative interactions of antibiotics with mucins however can lead to long exposure and increases the risk of resistant species. This study focuses on the evaluation of potential interactions of the last line of defence glycopeptide antibiotic teicoplanin with an ocular mucin model using precision hydrodynamic and microscopic techniques: sedimentation velocity in the analytical ultracentrifuge (SV-AUC), dynamic light scattering (DLS) and atomic force microscopy (AFM). For the mixtures of teicoplanin at higher doses (1.25 mg/mL and 12.5 mg/mL), it was shown to interact and aggregate with BSM in the distributions of both sedimentation coefficients by SV-AUC and hydrodynamic radii by DLS. The presence of aggregates was confirmed by AFM for higher concentrations. We suggest that teicoplanin eye drop formulations should be delivered at concentrations of < 1.25 mg/mL to avoid potentially harmful aggregations and to reduce the risk of antimicrobial resistance

The evaluation of two terpene-derived polymers, termed TPA6 and TPA7, as possible consolidants for archaeological wood was carried out. The overall objective of this work was to expand the non-aqueous treatment toolkit which is available for the conservation of the highly degraded Oseberg collection. The wood artefacts which were found on the Oseberg ship were treated with alum in the early 20th century, leading to the formation of sulfuric acid and to the precarious state that they are in today. Some of these artefacts cannot be treated with conventional aqueous consolidants, like polyethylene glycol, due to their highly degraded and/or reconstructed nature. This study sought to examine the level of penetration of the polymers in archaeological wood and to evaluate their consolidative effect. Both TPA6 and TPA7 were soluble in isopropanol and had a Mw of 3.9 and 4.2 kDa respectively. A number of archaeological wood specimens were immersed in solutions of these polymers. Their penetration and effects were evaluated using weight and dimensional change, colour change, infrared spectroscopy, scanning electron microscopy and hardness tests. Both polymers successfully penetrated the wood specimens, with a higher concentration found on the surface versus the core. Additionally, both polymers appeared to increase the hardness of the specimen surfaces. Increasing the polymer concentration and soaking time in future investigations could potentially facilitate the penetration to the wood cores.

A recent investigation was aimed at obtaining structural information on a highly extended protein via SEC-MALS-SAXS. Significantly broadened elution peaks were observed, reminiscent of a phenomenon known as viscous fingering. This phenomenon is usually observed above 50 mg/mL for proteins like bovine serum albumin (BSA). Interestingly, the highly extended protein (Brpt5.5) showed viscous fingering at concentrations lower than 5 mg/mL. The current study explores this and other non-ideal behavior, emphasizing the presence of these effects at relatively lower concentrations for extended proteins. BSA, Brpt5.5, and a truncated form of Brpt5.5 referred to as Brpt1.5 are studied systematically using size-exclusion chromatography (SEC), sedimentation velocity analytical ultracentrifugation (AUC), and viscosity. The viscous fingering effect is quantified using two approaches and is found to correlate well with the intrinsic viscosity of the proteins – Brpt5.5 exhibits the most severe effect and is the most extended protein tested in the study. By AUC, the hydrodynamic non-ideality was measured for each protein via global analysis of a concentration series. Compared to BSA, both Brpt1.5 and Brpt5.5 showed significant non-ideality that could be easily visualized at concentrations at or below 5 mg/mL and 1 mg/mL, respectively. A variety of relationships were examined for their ability to differentiate the proteins by shape using information from AUC and/or viscosity. Furthermore, these relationships were also tested in the context of hydrodynamic modeling. The importance of considering non-ideality when investigating the structure of extended macromolecules is discussed.

The Oseberg Viking ship burial is one of the most extensive collections of Viking wooden artefacts ever excavated in Norway. In the early twentieth century, many of these artefacts were treated with alum in order to preserve them, inadvertently leading to their current degraded state. It is therefore crucial to develop new bioinspired polymers which could be used to conserve these artefacts and prevent further disintegration. Two hydroxylated polymers were synthesised (TPA6 and TPA7), using α-pinene- and oleic acid-derived monomers functionalised with an acrylate moiety. Characterisation using biomolecular hydrodynamics (analytical ultracentrifugation and high precision viscometry) has shown that these polymers have properties which would potentially make them good wood consolidants. Conformation analyses with the viscosity increment (ν) universal hydrodynamic parameter and ELLIPS1 software showed that both polymers had extended conformations, facilitating in situ networking when applied to wood. SEDFIT-MSTAR analyses of sedimentation equilibrium data indicates a weight average molar mass Mw of (3.9 ± 0.8) kDa and (4.2 ± 0.2) kDa for TPA6 and TPA7 respectively. Analyses with SEDFIT (sedimentation velocity) and MultiSig however revealed that TPA7 had a much greater homogeneity and a lower proportion of aggregation. These studies suggest that both these polymers—particularly TPA7—have characteristics suitable for wood consolidation, such as an optimal molar mass, conformation and a hydroxylated nature, making them interesting leads for further research.

Consumers are increasingly looking for new plant-based alternatives to substitute animal proteins in their diets but for some applications it can be difficult to achieve the desired product microstructure using only plant proteins. One approach to facilitate structuring is to mix these plant-based ingredients with a polysaccharide. Here, the phase behaviour and microstructure of quinoa protein isolate (QPI) in mixture with maltodextrin (MD) of two dextrose equivalents (DE 7 and 2) were investigated. The binodals of both QPI-MD phase diagrams showed an atypical shape, where the concentration of MD in the QPI-rich phase and of QPI in the MD-rich phase increased with overall biopolymer concentration. Molecular weight distribution and microstructure analyses revealed that both maltodextrins fractionated between the phases and were probably entrapped within the volume-spanning protein network in the QPI-rich phase, indicating a depletion flocculation mechanism of phase separation. The pre-heating of QPI and the removal of salt from the systems resulted in similarly atypical phase diagrams. The approach presented contributes to our understanding of the phase behaviour of mixtures between plant proteins and polysaccharides, while the results suggest that the formulation of plant-based products of predictable properties may be more challenging than anticipated.

There is currently a pressing need for the development of novel bioinspired consolidants for waterlogged, archaeological wood. Bioinspired materials possess many advantages, such as biocompatibility and sustainability, which makes them ideal to use in this capacity. Based on this, a polyhydroxylated monomer was synthesised from α-pinene, a sustainable terpene feedstock derived from pine trees, and used to prepare a low molar mass polymer TPA5 through free radical polymerisation. This polymer was extensively characterised by NMR spectroscopy (chemical composition) and molecular hydrodynamics, primarily using analytical ultracentrifugation reinforced by gel filtration chromatography and viscometry, in order to investigate whether it would be suitable for wood consolidation purposes. Sedimentation equilibrium indicated a weight average molar mass Mw of (4.3 ± 0.2) kDa, with minimal concentration dependence. Further analysis with MULTISIG revealed a broad distribution of molar masses and this heterogeneity was further confirmed by sedimentation velocity. Conformation analyses with the Perrin P and viscosity increment ν universal hydrodynamic parameters indicated that the polymer had an elongated shape, with both factors giving consistent results and a consensus axial ratio of ~ 4.5. These collective properties—hydrogen bonding potential enhanced by an elongated shape, together with a small injectable molar mass—suggest this polymer is worthy of further consideration as a potential consolidant.

The Oseberg ship is one of the most important archaeological testimonies of the Vikings. After excavation in 1904, the wooden gravegoods were conserved using alum salts. This resulted in extreme degradation of a number of the objects a hundred years later through acid depolymerisation of cellulose and lignin. The fragile condition of the artefacts requires a reconsolidation which has to be done avoiding water as solvent. We synthesized tert-butyldimethylsilyl (TBDMS) chitosan which is soluble in a 50:50 solution of ethyl acetate and toluene. Measurement of its molecular weight, to anticipate its penetration, provided a challenge as the density difference of the polymer and solvent was too small to provide adequate solute redistribution under a centrifugal field, so a two-stage process was implemented (i) determination of the weight-average molar mass of the aqueous soluble activated precursor, chitosan mesylate, Mw,mc using sedimentation equilibrium with the SEDFIT-MSTAR algorithm, and determination of the degree of polymerisation DP; (ii) measurement of the average degree of substitution DSTBDMS of the TBDMS group on each chitosan monosaccharide monomer unit using NMR, to augment the Mw,mc value to give the molar mass of the TBDMS-chitosan. For the preparation, we find Mw = 9.8 kg·mol−1, which is within the acceptable limit for penetration and consolidation of degraded wood. Future work will test this on archaeological wood from different sources.

Purpose Insulin, in typical use, undergoes multiple changes in temperature; from refrigerator, to room temperature, to body temperature. Although long-term storage temperature has been well-studied, the short term changes to insulin are yet to be determined. Insulin detemir (IDet) is a clinically available, slow-acting, synthetic analogue characterised by the conjugation of a C14 fatty acid. The function of this modification is to cause the insulin to form multi-hexameric species, thus retarding the pharmacokinetic rate of action. In this investigation, the temperature dependence properties of this synthetic analogue is probed, as well as expiration. Methods Dynamic light scattering (DLS) and viscometry were employed to assess the effect of temperature upon IDet. Mass spectrometry was also used to probe the impact of shelf-life and the presence of certain excipients. Results IDet was compared with eight other insulins, including human recombinant, three fast-acting analogues and two other slow-acting analogues. Of all nine insulins, IDet was the only analogue to show temperature dependent behaviour, between 20 °C and 37 °C, when probed with non-invasive backscatter dynamic light scattering. Upon further investigation, IDet observed significant changes in size related to temperature, direction of temperature (heated/cooled) and expiration with cross-correlation observed amongst all 4 parameters. Conclusions These findings are critical to our understanding of the behaviour of this particular clinically relevant drug, as it will allow the development of future generations of peptide-based therapies with greater clinical efficacy.

Butvar B-98 and PDMS-OH both have a demonstrable ability as consolidants for archaeological wood. This makes them both potential treatment options for the Oseberg collection, which is one of the most important archaeological finds from the Viking era. Both Butvar B-98 and PDMS-OH are soluble in organic solvents, offering a useful alternative to aqueous-based consolidants. Extensive characterisation studies were carried out on both of these polymers, with the use of analytical ultracentrifugation and viscometry, for the benefit of conservators wanting to know more about the physical properties of these materials. Short column sedimentation equilibrium analysis using SEDFIT-MSTAR revealed a weight-average molar mass (weight-average molecular weight) Mw of (54.0 ± 1.5) kDa (kg · mol−1) for Butvar B-98, while four samples of PDMS-OH siloxanes (each with a different molar mass) had an Mw of (52.5 ± 3.0) kDa, (38.8 ± 1.5) kDa, (6.2 ± 0.7) kDa and (1.6 ± 0.1) kDa. Sedimentation velocity confirmed that all polymers were heterogeneous, with a wide range of molar masses. All molecular species showed considerable conformational asymmetry from measurements of intrinsic viscosity, which would facilitate networking interactions as consolidants. It is anticipated that the accumulated data on these two consolidants will enable conservators to make a more informed decision when it comes to choosing which treatment to administer to archaeological artefacts.

Multiple myeloma (MM) patients have increased risk of severe coronavirus disease 2019 (COVID-19) when infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Monoclonal gammopathy of undetermined significance (MGUS), the precursor of MM has been associated with immune dysfunction which may lead to severe COVID-19. No systematic data have been published on COVID-19 in individuals with MGUS. We conducted a large population-based cohort study evaluating the risk of SARS-CoV-2 infection and severe COVID-19 among individuals with MGUS. We included 75,422 Icelanders born before 1976, who had been screened for MGUS in the Iceland Screens Treats or Prevents Multiple Myeloma study (iStopMM). Data on SARS-CoV-2 testing and COVID-19 severity were acquired from the Icelandic COVID-19 Study Group. Using a test-negative study design, we included 32,047 iStopMM participants who had been tested for SARS-CoV-2, of whom 1754 had MGUS. Among these participants, 1100 participants, tested positive, 65 of whom had MGUS. Severe COVID-19 developed in 230 participants, including 16 with MGUS. MGUS was not associated with SARS-CoV-2 infection (Odds ratio (OR): 1.05; 95% confidence interval (CI): 0.81–1.36; p = 0.72) or severe COVID-19 (OR: 0.99; 95%CI: 0.52–1.91; p = 0.99). These findings indicate that MGUS does not affect the susceptibility to SARS-CoV-2 or the severity of COVID-19.

This study aims to understand possible effects of flavour compounds on the structure and conformation of endogenous proteins. Using methyl anthranilate (a grape flavour compound added to drinks, confectionery, and vape-liquids) and bovine serum albumin (BSA, a model serum protein) we designed experimental investigations using analytical ultracentrifugation, size exclusion chromatography small angle X-ray scattering, and fluorescence spectroscopy to reveal that methyl anthranilate spontaneously binds to BSA (ΔG°, ca. -21 KJ mol

Xanthan gum and scleroglucan, two rod-like polysaccharide hydrocolloids, are compared using a wide range of instrumental techniques and methods: steady shear flow, small amplitude oscillatory shear, first normal stress difference, capillary break-up and soft-contact tribology. The aqueous solutions of these two hydrocolloids with similar flow and viscoelastic profile show marked differences in capillary break-up time and apparent extensional viscosity. This result correlates with differences in first normal stress difference and, to a lesser extent, Stribeck curve behaviour. Formulating the hydrocolloids in concentrated sucrose solution (40 wt%) shifts relaxation profiles to longer times which greatly diminishes differences in rheological and lubrication behaviour. With exception of capillary break-up tests, no other methods showed statistically significant differences between the polysaccharides dissolved in the viscosified matrix. The toolbox of techniques is also applied to probe interactions of xanthan gum and scleroglucan with human whole saliva and bovine submaxillary mucin. We report no specific interactions between either hydrocolloid and salivary proteins and suggest that any cumulative effects must stem from specific sets of linear and non-linear rheological properties of saliva/hydrocolloid mixtures.

This work is part of a larger study, which aims to use soda lignin from straw as the starting point for a non-aqueous consolidant for highly degraded archaeological wood from the Oseberg collection. This wood was treated with alum salts in the early 1900s, is actively degrading and exists in varying states of preservation. Non-aqueous consolidants are an option to stabilize this wood mechanically in cases where it is too deteriorated to undergo aqueous-based retreatments, for example using polyethylene glycol. The aim of this study was to compare the extent of penetration of two soda lignin preparations in low- to medium-degraded archaeological pine. The soda lignins were dissolved in ethyl acetate and had two molecular weight groups: P1000 (molecular weight Mw of~3 kDa) and the ethyl acetate fraction FB01 (Mw of ~1 kDa). Penetration after immersion was evaluated by infrared spectroscopy and analytical pyrolysis. Treated specimens were also evaluated using weight and dimensional change and scanning electron microscopy. Both lignins penetrated into sample cores, but P1000 did not penetrate as well as FB01. This may be due to differences in their molecular weights, but also differences in polarity due to the presence of different functional groups.

Lignins are valuable renewable resources for the potential production of a large array of biofuels, aromatic chemicals and biopolymers. Yet native and industrial lignins are complex, highly branched and heterogenous macromolecules, properties that have to date often undermined their use as starting materials in lignin valorisation strategies. Reliable knowledge of weight average molar mass, conformation and polydispersity of lignin starting materials can be proven to be crucial to and improve the prospects for the success of such strategies. Here we evaluated the use of commonly-used size exclusion chromatography (SEC)—calibrated with polystyrene sulphonate standards—and under-used analytical ultracentrifugation—which does not require calibration—to characterise a series of lignin fractions sequentially extracted from soda and Kraft alkaline lignins using ethyl acetate, methyl ethyl ketone (MEK), methanol and acetone:water (fractions F01–F04, respectively). Absolute values of weight average molar mass (Mw) determined using sedimentation equilibrium in the analytical ultracentrifuge of (3.0 ± 0.1) kDa and (4.2 ± 0.2) kDa for soda and Kraft lignins respectively, agreed closely with previous SEC-determined Mws and reasonably with the size exclusion chromatography measurements employed here, confirming the appropriateness of the standards (with the possible exceptions of fraction F05 for soda P1000 and F03 for Indulin). Both methods revealed the presence of low (~ 1 kDa) Mw material in F01 and F02 fractions followed by progressively higher Mw in subsequent fractions. Compositional analysis confirmed > 90% (by weight) total lignins successively extracted from both lignins using MEK, methanol and acetone:water (F02 to F04). Considerable heterogeneity of both unfractionated and fractionated lignins was revealed through determinations of both sedimentation coefficient distributions and polydispersity indices. The study also demonstrates the advantages of using analytical ultracentrifugation, both alongside SEC as well as in its own right, for determining absolute Mw, heterogeneity and conformation information for characterising industrial lignins.

Hydrogen bonds or ‘H-bonds’ are polar, non-covalent bonds or interactions between a hydrogen atom (H) attached to a more electronegative atom, such as oxygen (O) or nitrogen (N), which partially pulls the electron cloud away from the H, leaving it electropositive—with another electronegative atom, such as O or N from a different molecule or from a different part of the same molecule. H-bond interactions play a huge role in the biochemistry of living processes, and in the structures and interactions of biological molecules, with each other and with different molecules including water. Nature's natural solvent, water, is itself a dynamic H-bonded polar structure, which strongly affects solubility and, as (dynamic) water of hydration, interactions between molecules. Compared with covalent and ionic bonds, H-bonds are individually much weaker (

Glycoconjugate vaccines use protein carriers to improve the immune response to polysaccharide antigens. The protein component allows the vaccine to interact with T cells, providing a stronger and longer-lasting immune response than a polysaccharide interacting with B cells alone. Whilst in theory the mere presence of a protein component in a vaccine should be sufficient to improve vaccine efficacy, the extent of improvement varies. In the present review, a comparison of the performances of vaccines developed with and without a protein carrier are presented. The usefulness of analytical tools for macromolecular integrity assays, in particular nuclear magnetic resonance, circular dichroism, analytical ultracentrifugation and SEC coupled to multi-angle light scattering (MALS) is indicated. Although we focus mainly on bacterial capsular polysaccharide-protein vaccines, some consideration is also given to research on experimental cancer vaccines using zwitterionic polysaccharides which, unusually for polysaccharides, are able to invoke T-cell responses and have been used in the development of potential all-polysaccharide-based cancer vaccines.A general trend of improved immunogenicity for glycoconjugate vaccines is described. Since the immunogenicity of a vaccine will also depend on carrier protein type and the way in which it has been linked to polysaccharide, the effects of different carrier proteins and production methods are also reviewed. We suggest that, in general, there is no single best carrier for use in glycoconjugate vaccines. This indicates that the choice of carrier protein is optimally made on a case-by-case basis, based on what generates the best immune response and can be produced safely in each individual case.Abbreviations: AUC: analytical ultracentrifugation; BSA: bovine serum albumin; CD: circular dichroism spectroscopy; CPS: capsular polysaccharide; CRM197: Cross Reactive Material 197; DT: diphtheria toxoid; Hib: Haemophilius influenzae type b; MALS: multi-angle light scattering; Men: Neisseria menigitidis; MHC-II: major histocompatibility complex class II; NMR: nuclear magnetic resonance spectroscopy; OMP: outer membrane protein; PRP: polyribosyl ribitol phosphate; PSA: Polysaccharide A1; Sa: Salmonella; St.: Streptococcus; SEC: size exclusion chromatography; Sta: Staphylococcus; TT: tetanus toxoid; ZPS: zwitterionic polysaccharide(s).

Circular dichroism (CD) (and synchrotron circular dichroism (SCD)) spectroscopy is a rapid, highly sensitive technique used to investigate structural conformational changes in biomolecules in response to interactions with ligands in solution and in film. It is a chiroptical method and at least one of the interacting molecules must possess optical activity (or chirality). In this review, we compare the capabilities of CD and SCD in the characterisation of celluloses and lignin polymers in archaeological wood. Cellulose produces a range of spectral characteristics dependent on environment and form; many of the reported transitions occur in the vacuum-ultraviolet region (< 180 nm) most conveniently delivered using a synchrotron source. The use of induced CD in which achiral dyes are bound to celluloses to give shifted spectra in the visible region is also discussed, together with its employment to identify the handedness of the chiral twists in nanocrystalline cellulose. Lignin is one target for the design of future consolidants that interact with archaeological wood to preserve it. It is reportedly achiral, but here we review several studies in which CD spectroscopy has successfully revealed lignin interactions with chiral enzymes, highlighting the potential usefulness of the technique in future research to identify new generation consolidants.

The global spread of enteric disease, the increasingly limited options for antimicrobial treatment and the need for effective eradication programs have resulted in an increased demand for glycoconjugate enteric vaccines, made with carbohydrate-based membrane components of the pathogen, and their precise characterisation. A set of physico-chemical and immunological tests are employed for complete vaccine characterisation and to ensure their consistency, potency, safety and stability, following the relevant World Health Organization and Pharmacopoeia guidelines. Variable requirements for analytical methods are linked to conjugate structure, carrier protein nature and size andO-acetyl content of polysaccharide. We investigated a key stability-indicating method which measures the percent free saccharide ofSalmonella entericasubspeciesentericaserovar Typhi capsular polysaccharide, by detergent precipitation, depolymerisation and HPAEC-PAD quantitation. Together with modern computational approaches, a more precise design of glycoconjugates is possible, allowing for improvements in solubility, structural conformation and stability, and immunogenicity of antigens, which may be applicable to a broad spectrum of vaccines. More validation experiments are required to establish the most effective and suitable methods for glycoconjugate analysis to bring uniformity to the existing protocols, although the need for product-specific approaches will apply, especially for the more complex vaccines. An overview of current and emerging analytical approaches for the characterisation of vaccines againstSalmonellaTyphi andShigellaspecies is described in this paper. This study should aid the development and licensing of new glycoconjugate vaccines aimed at the prevention of enteric diseases.

This retrospective investigation has established that the early theoretical attempts to directly incorporate the consequences of radial dilution into expressions for variation of the sedimentation coefficient as a function of the loading concentration in sedimentation velocity experiments require concentration distributions exhibiting far greater precision than that achieved by the optical systems of past and current analytical ultracentrifuges. In terms of current methods of sedimentation coefficient measurement, until such improvement is made, the simplest procedure for quantifying linear s-c dependence (or linear concentration dependence of 1/s) for dilute systems therefore entails consideration of the sedimentation coefficient obtained by standard c(s), g*(s) or G(s) analysis) as an average parameter ($$ \overline{s} $$ s ¯ ) that pertains to the corresponding mean plateau concentration (following radial dilution) ($$ \overline{c} $$ c ¯ ) over the range of sedimentation velocity distributions used for the determination of $$ \overline{s} $$ s ¯ . The relation of this with current descriptions of the concentration dependence of the sedimentation and translational diffusion coefficients is considered, together with a suggestion for the necessary improvement in the optical system.

Introduction Multiple myeloma (MM) patients have an increased risk of severe coronavirus disease 2019 (COVID-19) when infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Monoclonal gammopathy of undetermined significance (MGUS) precedes MM and related disorders and affects 4.2% of the general population over the age of 50 years. MM and MGUS are associated with immune dysfunction that is believed to contribute to the development of severe COVID-19. Currently, no systematic data on MGUS and COVID-19 have been published. We conducted a large population-based cohort study to evaluate whether MGUS was associated with SARS-CoV-2 infection and the development of severe COVID-19. Methods Data on all SARS-CoV-2 test results and COVID-19 severity was acquired from the COVID-19 Outpatient Clinic at Landspitali - The National University Hospital of Iceland. The first case of COVID-19 in Iceland was diagnosed on February 28 th, 2020. Since then, the Icelandic authorities have followed an aggressive strategy of SARS-CoV-2 testing and contact tracing. All SARS-CoV-2-positive individuals were immediately contacted and those with active infection were enrolled into telehealth monitoring consisting of repeated standardized interviews conducted by a nurse or physician. If clinical deterioration was detected, patients were assessed in person at the COVID-19 Outpatient Clinic and admitted if needed. Study participants were included from the Iceland Screens Treats or Prevents Multiple Myeloma study (iStopMM). The study is an ongoing population-based screening study for MGUS and randomized trial of follow-up strategies. Out of the 148,708 Icelanders who were born 1976 and earlier and were alive on September 9 th 2016, 80,759 (54%) provided informed consent for study participation and 75,422 (94%) of those provided a blood sample for MGUS screening by serum protein electrophoresis (SPEP) and free light chain (FLC) assay. MGUS was determined by current criteria using SPEP and FLC assay data. Individuals who had died, been diagnosed with MM and related disorders, or were undergoing treatment for smoldering MM prior to February 28 th were excluded. First, the association of MGUS and testing positive for SARS-CoV-2 was evaluated. We used a test negative design and included participants who had been tested at least once for SARS-CoV-2 between February 28 th and December 31 st, 2020. The association of MGUS and a positive test for SARS-CoV-2 was assessed using logistic regression, adjusted for sex and age. Next, the association of MGUS and severe COVID-19 was evaluated. Those who tested positive for SARS-CoV-2 were included unless they were hospitalized or living in a nursing home at diagnosis. Participants were followed until discharge from telehealth monitoring or until considered having severe COVID-19. Severe COVID-19 was defined as the composite outcome of the need for outpatient visit or hospital admission and death and as the composite outcome of hospital admission and death. Logistic regression was then performed adjusting for sex and age. Results Of the 75,422 individuals screened for MGUS, 32,047 (42%) were tested for SARS-CoV-2 during the study period of whom 1,754 had MGUS (5.5%). Those with MGUS were older (mean age 66.3 vs 59.1 p Of those who tested positive for SARS-CoV-2, a total of 230 had the composite outcome of requiring an outpatient visit or hospital admission, and death, and 117 had the composite outcome of hospital admission and death. After adjusting for age and sex, MGUS was not found to be associated with either endpoint (OR: 0.99; 95%CI: 0.52-1.91; p=0.99 and OR: 1.13; 95%CI: 0.52-2.46; p=0.76; Table; Figure B) Conclusions: In this large population-based study that included 75,422 individuals screened for MGUS, we did not find MGUS to be associated with SARS-CoV-2 susceptibility or COVID-19 severity. This is contrary to MM which is preceded by MGUS. These findings suggest that immunosuppression in MGUS differs significantly from that of MM and are important since they can inform management and recommendations for individuals with MGUS. Figure 1 Figure 1. Disclosures Kampanis: The Binding Site: Current Employment. Hultcrantz: Intellisphere LLC: Consultancy; Daiichi Sankyo: Research Funding; Curio Science LLC: Consultancy; Amgen: Research Funding; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding. Durie: Amgen, Celgene/Bristol-Myers Squibb, Janssen, and Takeda: Consultancy; Amgen: Other: fees from non-CME/CE services . Harding: The Binding Site: Current Employment, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Landgren: Amgen: Honoraria; Janssen: Honoraria; Celgene: Research Funding; Janssen: Other: IDMC; Janssen: Research Funding; Takeda: Other: IDMC; Amgen: Research Funding; GSK: Honoraria. Kristinsson: Amgen: Research Funding; Celgene: Research Funding.

Background: In addition to serum protein electrophoresis (SPEP) and serum immunofixation (IFE), measurement of serum free light chains (FLC) has become the hallmark for detection, prognostication, and monitoring of monoclonal gammopathies. However, serum FLC levels are heavily dependent on kidney function due to discrepancy in the rate of kidney clearance of the FLC. As chronic kidney disease (CKD) is common in the general population, it is possible that a large number of individuals have false FLC results. A kidney reference interval (eGFR < 60) has previously been published based on small numers (N=688) for serum FLC ratio (0.37-3.10) instead of the standard reference interval (0.26-1.65), but no kidney reference exists for kappa (3.3-19.4mg/L) or lambda (5.7-26.3mg/L). The aim of this study was to define and improve the reference interval for individuals with various degree of decreased kidney function and assess its effect on prevalence of light chain monoclonal gammopathy of undetermined significance (LC-MGUS) among CKD patients. Methods: A total of 80,759 participants of the Iceland Screens, Treats or Prevents Multiple Myeloma (iStopMM) study were included. Participants were screened by SPEP and IFE as well as by serum FLC assay (Freelite®). Serum creatinine (SCr) value closest to the time of screening was used to calculate (CKD-EPI) eGFR. Participants with M-protein, eGFR >60 mL/min/1.73 m2, missing SCr measurements or more than 1 year from the SCr measurement to the iStopMM screening were excluded. Correlation was assessed graphically and using the Spearman correlation coefficient. A nonparametric bootstrapping method was used to calculate the 95% CI. Partitioning was determined based on the proportion of subgroups (sex, age, eGFR) outside the whole group reference interval (4.1%). However, intervals were not partitioned if subgroups included few participants and/or if deemed more applicable and reasonable for clinical practice. LC-MGUS was defined as FLC ratio outside the reference interval and raised FLC level without evidence of monoclonal heavy chain on SPEP or IFE or end-organ damage attributed to the plasma cell proliferative disorder. Results: Of 75,422 individuals who were screened, 6,503 (12%) participants had eGFR < 60 mL/min/1.73 m2, without evidence of monoclonality on SPEP or IFE and were analysed further. The median (IQR) kappa level was 21.7 (16.6-29.4) mg/L, lambda level 19.0 mg/L (14.8-25.0) and FLC ratio 1.16 (0.97-1.39). Serum FLC levels increased with decreasing eGFR: serum free kappa (ρ= -0.44, p < 0.001), lambda (ρ= -0.39, p < 0.001), and FLC ratio (ρ= -0.15, p < 0.001). Using current reference intervals, 60% and 21% of persons had values outside the normal range for kappa and lambda, respectively. The FLC ratio was outside the standard reference interval in 9% and outside the kidney reference interval in 0.6% of participants. Based on these findings, we established new reference intervals for FLC and FLC ratio (Table). Participants on dialysis (N=26) had significantly higher median (IQR) serum free kappa 29.3 mg/L (26.1-57.6, p = 0.01) and lambda 25.1 mg/L (19.5-52.6, p = 0.01) than participants with eGFR 30-59, but no significant difference compared to participants with eGFR < 30 (p = 0.57). The FLC ratio in participants on dialysis was 1.22 (0.96-1.37), which was similar to participants with eGFR 45-59 (p = 0.63) and 30-44 (p = 0.34). Participants on dialysis had significantly lower FLC ratio than participants with eGFR < 30, or 1.31 (1.10-1.57, p = 0.02). Utilising our novel reference intervals, the crude prevalence of LC-MGUS in all participants with eGFR < 60 was 75 (1.2%), with no difference between participants with eGFR of 45-59, 30-44 and < 30, respectively. When the crude rate of LC-MGUS was compared to a rate based on previous reference intervals, the crude rate of both kappa and lambda LC-MGUS increased in participants with eGFR 45-59, eGFR 30-44 and eGFR < 30 (p Conclusion: Current reference intervals for serum FLC and FLC ratio are inaccurate for patients with decreased kidney function. Here we propose new reference intervals for serum FLC and FLC ratio for use in patients with CKD which also seem to be accurate in patients on dialysis. Implementing these novel reference intervals is likely to increase the sensitivity and specificity of the analyses and lead to a more accurate diagnosis of monoclonal gammopathy in individuals with kidney dysfunction. Figure 1 Figure 1. Disclosures Kampanis: The Binding Site: Current Employment. Hultcrantz: Intellisphere LLC: Consultancy; Curio Science LLC: Consultancy; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Research Funding; Daiichi Sankyo: Research Funding. Durie: Amgen, Celgene/Bristol-Myers Squibb, Janssen, and Takeda: Consultancy; Amgen: Other: fees from non-CME/CE services . Harding: The Binding Site: Current Employment, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Landgren: Janssen: Research Funding; Amgen: Honoraria; Janssen: Honoraria; Celgene: Research Funding; Janssen: Other: IDMC; Takeda: Other: IDMC; Amgen: Research Funding; GSK: Honoraria. Kristinsson: Amgen: Research Funding; Celgene: Research Funding.

Introduction Monoclonal gammopathy of undetermined significance (MGUS) is a plasma cell disorder preceding multiple myeloma and related disorders, present in 4.2% of the population over the age of 50. Although usually asymptomatic, MGUS has been associated with various health-related problems, including thrombosis, infections, fractures, neuropathy, and death. Because MGUS is asymptomatic, its diagnosis is typically incidental, during clinical workup for unrelated medical issues, and therefore most individuals remain undiagnosed. Consequently, MGUS cohorts in past studies may have suffered from more comorbidities than the actual population with MGUS. This might have introduced selection bias in previous studies on MGUS, which extent has not been studied in a systematic way. Therefore, previously reported associations between MGUS and various medical issues might not be as profound as formerly observed. The aim of this study was to compare characteristics of incidentally diagnosed MGUS versus MGUS diagnosed by systematic screening, with particular focus on demographics, comorbidities, and MGUS-related factors. Methods The study is based on the Iceland Screens, Treats, or Prevents Multiple Myeloma (iStopMM) study. iStopMM is a population-based screening study for MGUS and a randomized controlled trial of follow-up strategies that has included 54% (n = 80,759) of the Icelandic population above 40 years of age. In total, 75,422 participants were screened for MGUS by serum protein electrophoresis (SPEP) and free light chain (FLC) assay. Information on which individuals had incidentally diagnosed MGUS (clinical MGUS) prior to participation in iStopMM were gathered from the Icelandic cancer registry and laboratory results from Landspítali University Hospital and Læknasetrið, the only laboratories in Iceland that perform SPEP. M-protein concentration, MGUS isotype and FLC ratio were obtained from the original screening samples. Comorbidity data was acquired from two high-quality national registries: Hospital Discharge Register and Register of Primary Health Care Contacts, with >95% completeness and accuracy. MGUS diagnosed by screening was further classified into MGUS with or without M-proteins (light-chain MGUS). Those with clinical MGUS were used as the reference group in all analyses. Since all individuals with clinical MGUS had M-proteins, individuals with light-chain MGUS were excluded from this study. T-test and chi-square test were used for demographic comparison; linear regression adjusting for sex and age for continuous variables, and logistic regression adjusting for sex and age for comparison of categorical variables, regarding MGUS-related factors and comorbidities. Results The study cohort consisted of 3,300 individuals who had MGUS with M-proteins; 224 individuals with clinical MGUS and 3,076 with screened MGUS. The clinical MGUS group was significantly older (p Discussion In this large population-based study including 75,000 screened individuals, we found clinical MGUS cases to be older, more likely to live in Iceland's capital area, and have a higher M-protein concentration than those found to have MGUS while screened on the iStopMM study. Individuals with clinical MGUS also had a higher number of underlying comorbidities and were 1.5-3.3 times more likely to suffer from arrhythmias, chronic kidney diseases, endocrine disorders, heart failure, neurological diseases, and rheumatological diseases. Our findings highlight the importance of screening studies to evaluate the true epidemiological and biological implications of MGUS and suggest selection bias in prior studies. Figure 1 Figure 1. Disclosures Kampanis: The Binding Site: Current Employment. Hultcrantz: Daiichi Sankyo: Research Funding; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding; Curio Science LLC: Consultancy; Amgen: Research Funding; Intellisphere LLC: Consultancy. Durie: Amgen, Celgene/Bristol-Myers Squibb, Janssen, and Takeda: Consultancy; Amgen: Other: fees from non-CME/CE services . Harding: The Binding Site: Current Employment, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Landgren: Janssen: Honoraria; Janssen: Other: IDMC; Takeda: Other: IDMC; Janssen: Research Funding; Celgene: Research Funding; Amgen: Honoraria; Amgen: Research Funding; GSK: Honoraria. Kristinsson: Amgen: Research Funding; Celgene: Research Funding.

Food flavour ingredients are required by law to obtain prior approval from regulatory bodies, such as the U.S. Food and Drug Administration (FDA) or the European Food Safety Authority (EFSA) in terms of toxicological data and intended use levels. However, there are no regulations for labelling the type and concentration of flavour additives on the product, primarily due to their low concentration in food and generally recognised as safe (GRAS) status determined by the flavour and extract manufacturers’ association (FEMA). Their status for use in e-cigarettes and other vaping products challenges these fundamental assumptions, because their concentration can be over ten-thousand times higher than in food, and the method of administration is through inhalation, which is currently not evaluated by the FEMA expert panel. This work provides a review of some common flavour ingredients used in food and vaping products, their product concentrations, inhalation toxicity and aroma interactions reported with different biological substrates. We have identified several studies, which suggest that the high concentrations of flavour through inhalation may pose a serious health threat, especially in terms of their cytotoxicity. As a result of the wide range of possible protein-aroma interactions reported in our diet and metabolism, including links to several non-communicable diseases, we suggest that it is instrumental to update current flavour- labelling regulations, and support new strategies of understanding the effects of flavour uptake on the digestive and respiratory systems, in order to prevent the onset of future non-communicable diseases.

The influence of Viking-Age migrants to the British Isles is obvious in archaeological and place-names evidence, but their demographic impact has been unclear. Autosomal genetic analyses support Norse Viking contributions to parts of Britain, but show no signal corresponding to the Danelaw, the region under Scandinavian administrative control from the ninth to eleventh centuries. Y-chromosome haplogroup R1a1 has been considered as a possible marker for Viking migrations because of its high frequency in peninsular Scandinavia (Norway and Sweden). Here we select ten Y-SNPs to discriminate informatively among hg R1a1 sub-haplogroups in Europe, analyse these in 619 hg R1a1 Y chromosomes including 163 from the British Isles, and also type 23 short-tandem repeats (Y-STRs) to assess internal diversity. We find three specifically Western-European sub-haplogroups, two of which predominate in Norway and Sweden, and are also found in Britain; star-like features in the STR networks of these lineages indicate histories of expansion. We ask whether geographical distributions of hg R1a1 overall, and of the two sub-lineages in particular, correlate with regions of Scandinavian influence within Britain. Neither shows any frequency difference between regions that have higher (≥10%) or lower autosomal contributions from Norway and Sweden, but both are significantly overrepresented in the region corresponding to the Danelaw. These differences between autosomal and Y-chromosomal histories suggest either male-specific contribution, or the influence of patrilocality. Comparison of modern DNA with recently available ancient DNA data supports the interpretation that two sub-lineages of hg R1a1 spread with the Vikings from peninsular Scandinavia. ispartof: European Journal Of Human Genetics vol:29 issue:3 pages:512-523 ispartof: location:England status: published

Consumer sensory evaluation, aroma release analysis and biophysical protein analysis were used to investigate the effect of ethanol on the release and perception of flavour in beer (lager and stout) at different ethanol levels (0 and 5% ABV). Consumer study results showed no significant differences in orthonasal perception, yet retronasal results showed that 0% lager was perceived as maltier with reduced fruitiness, sweetness, fullness/body and alcohol warming sensation (p P, the presence of α-amylase selectively reduces the headspace concentration of hydrophobic compounds. It was found that ethanol has a subtle inhibitory effect on the binding of hydrophobic compounds to α-amylase, thereby increasing their headspace concentration in the 5% ABV as compared to the 0% beers. This synergistic ethanol * saliva effect is attributed to the changes in the conformation of α-amylase due to ethanol-induced denaturation. It is hypothesised that the partially unfolded protein structures have a lower number of hydrophobic pockets, leading to a lower capacity to entrap hydrophobic aroma compounds. This supports the hypothesis that ethanol * saliva interactions directly impact the sensory and flavour properties of beer, which would provide a basis for further investigations in reformulation of 0% ABV drinks.

Aroma compounds are diverse low molecular weight organic molecules responsible for the flavour of food, medicines or cosmetics. Natural and artificial aroma compounds are manufactured and used by the industry to enhance the flavour and fragrance of products. While the low concentrations of aroma compounds present in food may leave no effect on the structural integrity of the mucosa, the effect of concentrated aroma volatiles is not well understood. At high concentrations, like those found in some flavoured products such as e-cigarettes, some aroma compounds are suggested to elicit a certain degree of change in the mucin glycoprotein network, depending on their functional group. These effects are particularly associated with carbonyl compounds such as aldehydes and ketones, but also phenols which may interact with mucin and other glycoproteins through other interaction mechanisms. This study demonstrates the formation of such interactions in vitro through the use of molecular hydrodynamics. Sedimentation velocity studies reveal that the strength of the carbonyl compound interaction is influenced by compound hydrophobicity, in which the more reactive short chain compounds show the largest increase in mucin-aroma sedimentation coefficients. By contrast, the presence of groups that increases the steric hindrance of the carbonyl group, such as ketones, produced a milder effect. The interaction effects were further demonstrated for hexanal using size exclusion chromatography light scattering (SEC-MALS) and intrinsic viscosity. In addition, phenolic aroma compounds were identified to reduce the sedimentation coefficient of mucin, which is consistent with interactions in the non-glycosylated mucin region.

Cucurbits are plants that have been used frequently as functional foods. This study includes the extraction, isolation, and characterisation of the mesocarp polysaccharide of Cucurbita moschata. The polysaccharide component was purified by gel filtration into three fractions (NJBTF1, NJBTF2, and NJBTF3) of different molecular weights. Characterisation includes the hydrodynamic properties, identification of monosaccharide composition, and bioactivity. Sedimentation velocity also indicated the presence of small amounts of additional discrete higher molecular weight components even after fractionation. Sedimentation equilibrium revealed respective weight average molecular weights of 90, 31, and 19 kDa, with the higher fractions (NJBTF1 and NJBTF2) indicating a tendency to self-associate. Based on the limited amount of data (combinations of 3 sets of viscosity and sedimentation data corresponding to the 3 fractions), HYDFIT indicates an extended, semi-flexible coil conformation. Of all the fractions obtained, NJBTF1 showed the highest bioactivity. All fractions contained galacturonic acid and variable amounts of neutral sugars. To probe further, the extent of glycosidic linkages in NJBTF1 was estimated using gas chromatography&ndash, mass spectrometry (GCMS), yielding a high galacturonic acid content (for pectin polysaccharide) and the presence of fructans&mdash, the first evidence of fructans (levan) in the mesocarp. Our understanding of the size and structural flexibility together with the high bioactivity suggests that the polysaccharide obtained from C. moschata has the potential to be developed into a therapeutic agent.

Polysaccharides are promising macromolecular platforms for use in the life sciences. Here, bioactive cellulose, pullulan, and dextran valproates are characterized hydrodynamically by sedimentation velocity and thermodynamically by sedimentation equilibrium analytical ultracentrifugation. Using sedimentation-diffusion analysis of sedimentation velocity experiments by numerical solution of the Lamm equation enabled the calculation of sedimentation and diffusion coefficients, and consequently molar masses. Sedimentation equilibrium experiments were then also used to determine the average molar masses. The corresponding set of data, with independently performed self-diffusion measurements by nuclear magnetic resonance spectroscopy, and together with size exclusion chromatography molar masses by coupling to refractive index-, viscometric-, and multi-angle laser light scattering detection, were subsequently correlated to each other by the hydrodynamic invariant and sedimentation parameter. We assess statistically most relevant average values of the molar masses of these polysaccharide valproates with relevant macromolecular conformational characteristics.

This study concerns glulisine, a rapid-acting insulin analogue that plays a fundamental role in diabetes management. We have applied a combination of methods namely X-ray crystallography, and biophysical characterisation to provide a detailed insight into the structure and function of glulisine. X-ray data provided structural information to a resolution of 1.26 Å. Crystals belonged to the H3 space group with hexagonal (centred trigonal) cell dimensions a = b = 82.44 and c = 33.65 Å with two molecules in the asymmetric unit. A unique position of D21Glu, not present in other fast-acting analogues, pointing inwards rather than to the outside surface was observed. This reduces interactions with neighbouring molecules thereby increasing preference of the dimer form. Sedimentation velocity/equilibrium studies revealed a trinary system of dimers and hexamers/dihexamers in dynamic equilibrium. This new information may lead to better understanding of the pharmacokinetic and pharmacodynamic behaviour of glulisine which might aid in improving formulation regarding its fast-acting role and reducing side effects of this drug.

The 6-deoxy-6-aminocelluloses—or “aminocelluloses”—are a class of synthetic natural cellulose derivatives which are mostly aqueous soluble and have excellent film-forming properties. Recent studies have connected these properties at the molecular level with protein-like self-associative behaviour for a range of aminocelluloses including a 6-deoxy-6-(ω-aminoethyl) aminocellulose AEA-1 with the association being a two-stage process—a reversible oligomerisation followed by further (semi-reversible) aggregation into larger structures. Here, we synthesise and compare a new 6-deoxy-6-(ω-aminoethyl) aminocellulose AEA-1′ with different degree of substitution with one with further alkyl derivatisation, namely 6-deoxy-6-(ω-hydroxyethyl) aminocellulose HEA-1′. As with AEA-1, sedimentation velocity and sedimentation equilibrium in the analytical ultracentrifuge still show a two-stage process for both AEA-1′ and HEA-1′, with the latter giving higher molar masses. The consequences of these properties for use as consolidants for archaeological wood are considered.

Quorum‐sensing mechanisms regulate gene expression in response to changing cell‐population density detected through pheromones. In Enterococcus faecalis, Fsr quorum sensing produces and responds to the gelatinase biosynthesis‐activating pheromone (GBAP). Here we establish that the enterococcal FsrB membrane protein has a direct role connected with GBAP by showing that GBAP binds to purified FsrB. Far‐UV CD measurements demonstrated a predominantly α‐helical protein exhibiting a small level of conformational flexibility. Fivefold (400 μm) GBAP stabilised FsrB (80 μm) secondary structure. FsrB thermal denaturation in the presence and absence of GBAP revealed melting temperatures of 70.1 and 60.8 °C, respectively, demonstrating GBAP interactions and increased thermal stability conferred by GBAP. Addition of GBAP also resulted in tertiary structural changes, confirming GBAP binding.

Vancomycin, a branched tricyclic glycosylated peptide antibiotic, is a last-line defence against serious infections caused by staphylococci, enterococci and other Gram-positive bacteria. Orally-administered vancomycin is the drug of choice to treat pseudomembranous enterocolitis in the gastrointestinal tract. However, the risk of vancomycin-resistant enterococcal infection or colonization is significantly associated with oral vancomycin. Using the powerful matrix-free assay of co-sedimentation analytical ultracentrifugation, reinforced by dynamic light scattering and environmental scanning electron microscopy, and with porcine mucin as the model mucin system, this is the first study to demonstrate strong interactions between vancomycin and gastric and intestinal mucins, resulting in very large aggregates and depletion of macromolecular mucin and occurring at concentrations relevant to oral dosing. In the case of another mucin which has a much lower degree of glycosylation (~60%) – bovine submaxillary mucin - a weaker but still demonstrable interaction is observed. Our demonstration - for the first time - of complexation/depletion interactions for model mucin systems with vancomycin provides the basis for further study on the implications of complexation on glycopeptide transit in humans, antibiotic bioavailability for target inhibition, in situ generation of resistance and future development strategies for absorption of the antibiotic across the mucus barrier., The authors are grateful for a grant from the Independent Diabetes Trust (G.G.A. and S.E.H.) and the Engineering and Physical Sciences Research Council, grant number EP/L015633/1 (I.F., S.E.H., G.G.A., V.D.).

In the associated published article Kremmyda et al. (2021), the C1 region of the NMR spectra were examined in detail, as the C1 carbon was common to the crucial linkages in the glucan, and the most clearly separated region in the spectra. We provide the original measurement data here in topspin format, in order that different authors can examine the regions corresponding to the other carbons and in addition repeat our processing of the FID's using different parameters. Mushroom, Cereal, Reference and other samples, such as salts and salt hydrates have been measured. A series of different experiments on individual samples is also given. All of the samples and experiments available in the database are summarised in the Table below. We also provide an FTIR database in the form of an excel workbook with spectra in frequency/absorbence pairs.

We recall the experimental approaches involved in the discovery of hydrogen bonds in deoxyribonucleic acid (DNA) made 70 years ago by a team of scientists at University College Nottingham led by J.M. Gulland, and in relation to previous studies. This discovery proved an important step in the elucidation of the correct structure for DNA made by J.D. Watson and F.H.C. Crick, as acknowledged in ‘The Double Helix’. At that time of the discovery, however, it was impossible to delineate between inter- and intra-chain hydrogen bonds. We also consider in the light of more recent hydrodynamic theory a tentative model for DNA proposed by Gulland's and D.O. Jordan's PhD student J.M. Creeth in his PhD thesis of 1948, with the correct prediction of two chains with a sugar-phosphate backbone on the exterior and hydrogen-bonded bases between the nucleotide bases of opposite chains in the interior. Our analysis shows that his incorporation of alternating breaks in the two-chain structure was not necessary to explain the viscosity data on scission of hydrogen bonds after titrating to high or low pH. Although Creeth's model is a depiction of DNA structure alone, he could not know whether the hydrogen bonding was intermolecular, although this was subsequently proved correct by others. The mechanisms by which replicative processes occurred were of course unknown at that time, and so, he could not have realised how closely his tentative model resembled steps in some viral replicative mechanisms involving the molecule of life that he was working on.

The analytical ultracentrifuge (AUC) invented by T. Svedberg has now become an extremely versatile and diverse tool in Biochemistry and Polymer Science for the characterisation of the sizes, shapes and interactions of particles ranging in size from a few nanometres to tens of microns, or in molecular weight, M (molar mass) terms from a few hundred daltons to hundreds of megadaltons. We illustrate this diversity by reviewing recent work on (1) small lignin-like isoeugenols of M ~ 0.4–0.9 kDa for archaeological wood conservation, (2) protein-like association of a functional amino-cellulose M = 3.25 kDa, (3) a small glycopeptide antibiotic (M ~ 1.5 kDa) and its association with a protein involved in antibiotic resistance (M ~ 47 kDa), (4) tetanus toxoid protein TTP (M ~ 150 kDa) and (5) the incorporation of TTP into two huge glycoconjugates considered in glycovaccine development with molecular weight species in a broad distribution appearing to reach 100 MDa. In illustrating the diversity, we will highlight developments in hydrodynamic analysis which have made the AUC such an exciting and important instrument, and point to a potential future development for extending its capability to highly concentrated systems.

The major challenges in biophysical characterization of human protein-carbohydrate interactions are obtaining monodispersed preparations of human proteins that are often post-translationally modified and lack of detection of carbohydrates by traditional detection systems. Light scattering (dynamic and static) techniques offer detection of biomolecules and their complexes based on their size and shape, and do not rely on chromophore groups (such as aromatic amino acid sidechains). In this study, we utilized dynamic light scattering, analytical ultracentrifugation and small-angle X-ray scattering techniques to investigate the solution properties of a complex resulting from the interaction between a 15 kDa heparin preparation and miniagrin, a miniaturized version of agrin. Results from dynamic light scattering, sedimentation equilibrium, and sedimentation velocity experiments signify the formation of a monodisperse complex with 1:1 stoichiometry, and low-resolution structures derived from the small-angle X-ray scattering measurements implicate an extended conformation for a side-by-side miniagrin‒heparin complex.

Background Multiple myeloma (MM) evolves from monoclonal gammopathy of undetermined significance (MGUS), a clinically detectable but asymptomatic premalignant phase seen in ~3% of the general population 50 years of age or older. The prevalence of MGUS has not been described in a population at high risk of developing MM, specifically Black/African American (AA) individuals or first-degree relatives of patients with hematologic malignancies (HM). In 2019, we launched the first nationwide US screening study for individuals at high risk of MM to help better identify what population would benefit most from screening and early intervention for precursor MM stages. We aim to assess the prevalence of MGUS in a population at high risk of MM and characterize clinical variables of individuals who screen positive. Here, we report interim screening data on the first 2,960 participants. Methods Individuals aged 40 or older with an additional MM risk factor are eligible to be screened in the PROMISE Study. High-risk individuals include Black/AAs and those with a first-degree relative diagnosed with a hematologic malignancy or a precursor condition to MM. Blood from all participants was analyzed via serum protein electrophoresis, immunofixation, and Optilite® to measure the serum free light chains (sFLC), IgG, IgA and IgM. Results were returned to all participants, and those who tested positive for a monoclonal gammopathy (MGUS/SMM) were referred to a hematologist for clinical follow-up and invited to periodically complete epidemiologic exposure and psychosocial questionaries, including a 4-item cancer worry questionnaire and the RAND 36-item Short Form Survey (SF-36). To investigate the use of the higher-sensitivity matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) with the Optilite® IgG, IgA, IgM and sFLC results as a screening test for all participants, we rescreened 1,092 samples from PROMISE. The Binding Site Group proprietary software was used for the analysis of the combined MS/Optilite® results, allowing for the detection and quantification of M-protein. Heavy-Chain MGUS (HC-MGUS) was defined by the presence of one or more paired heavy and light chain monoclonal peaks detected by MS. Pairing was based on mass to charge ratio of identified peaks. To enrich the PROMISE cohort with Black/AA individuals, we identified and screened 1,868 Black/AA additional individuals from the Mass General Brigham (MGB) Biobank who met the PROMISE enrollment criteria. Screening was performed by MS/Optilite®, and results have not been returned to participants. Results We screened 2,960 participants with the combined MS/Optilite® approach. We report here the prevalence of HC-MGUS and plan on presenting the estimated rate of light chain MGUS in our cohort, at the meeting. We detected HC-MGUS in 9.6% (95% CI: 8.6-11%) of our cohort, with a prevalence of 10% (95% CI: 8.3-12%) in the PROMISE cohort and 9.4% (95% CI: 8.1-11%) in the MGB cohort (Table 1 and Figure 1). HC-MGUS prevalence increased with age in high-risk individuals from 4.9% (CI: 3.3-6.9%) for participants aged 40-49 to 13% (CI: 10-17%) in the 70-79 range (P < 1.2E-5). Among monoclonal HC-MGUS, we found 65% IgG, 18% IgM, and 18% IgA. M-spike was quantified in 97% of samples. Median M-spike concentration was, 0.058g/dL (max. 2.6g/dL) for IgG, 0.0043g/dL (max. 0.6g/dL) for IgM, and 0.067g/dL (max. 0.8g/dL) for IgA. In the Promise cohort, no significant change in cancer worry was observed across the pre- and post-screening interval among participants who screened positive (P = 0.52). Health-related quality of life, as measured by the SF-36, was not significantly different in screen-positive vs. screen-negative individuals for any of the eight subscales (all P > 0.20). Conclusions We present the largest dataset on monoclonal gammopathy prevalence and screening in individuals at high risk for MM, and more specifically the largest cohort of Black/AA, using a novel high-sensitivity testing approach. Our results confirm that older adults who are Black/AA or have a first-degree relative with an HM have a high prevalence MGUS and may benefit from precision screening approaches to allow for early detection and clinical intervention. Preliminary data on cancer worry and quality of life indicates that the psychosocial burden of screening in this population is likely minimal. Figure 1 Figure 1. Disclosures Sakrikar: The Binding Site: Current Employment. Krause: The Binding Site: Current Employment. Barnidge: The Binding Site: Current Employment. Bustoros: Takeda: Consultancy, Honoraria; Janssen, Bristol Myers Squibb: Honoraria, Speakers Bureau. Perkins: The Binding Site: Current Employment. Harding: The Binding Site: Current Employment, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Kapoor: Ichnos Sciences: Research Funding; Amgen: Research Funding; Pharmacyclics: Consultancy; Takeda: Research Funding; Sanofi: Consultancy; Regeneron Pharmaceuticals: Research Funding; AbbVie: Research Funding; BeiGene: Consultancy; Sanofi: Research Funding; Karyopharm: Research Funding; Glaxo SmithKline: Research Funding; Karyopharm: Consultancy; Cellectar: Consultancy. Mo: Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees; GSK: Consultancy, Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Epizyme: Consultancy; Eli Lilly: Consultancy; Janssen: Honoraria; Karyopharm: Honoraria, Membership on an entity's Board of Directors or advisory committees. Murray: The Binding Site: Patents & Royalties: Potential Royalties for use of mass spectrometry in M-protein detection. Getz: Scorpion Therapeutics: Consultancy, Current holder of individual stocks in a privately-held company, Membership on an entity's Board of Directors or advisory committees; IBM, Pharmacyclics: Research Funding. Marinac: JBF Legal: Consultancy; GRAIL Inc: Research Funding. Ghobrial: AbbVie, Adaptive, Aptitude Health, BMS, Cellectar, Curio Science, Genetch, Janssen, Janssen Central American and Caribbean, Karyopharm, Medscape, Oncopeptides, Sanofi, Takeda, The Binding Site, GNS, GSK: Consultancy.

Background: Cancer screening is performed worldwide for several malignancies. Monoclonal gammopathy of undetermined significance (MGUS) precedes multiple myeloma (MM) and related lymphoproliferative disorders (LP). However, less than 5% of all MM patients are diagnosed during their precursor state and individuals who develop MM while being monitored for MGUS have better overall survival and fewer complications, compared to MM patients diagnosed without knowledge of MGUS. Thus, population-based screening for MGUS could identify candidates for early treatment of MM/LPs. To evaluate whether systematic screening is beneficial, we performed the Iceland Screens, Treats, or Prevents Multiple Myeloma (iStopMM) study, the first population-based screening study for MGUS that includes a randomized clinical trial (RCT) of follow-up and treatment strategies. Methods: All living residents of Iceland on September 9th, 2016 who were born before 1976 (N=148,708) were invited to participate. Of those, 80,759 (54.3%) provided informed consent for screening. Serum samples were collected from participants alongside clinical blood sampling in the Icelandic health service between September 2016 and the end of 2020. All samples were shipped to the Binding Site in Birmingham, UK, for screening. Samples were tested for M-proteins by capillary zone electrophoresis and immunofixation electrophoresis performed to confirm and characterize suspected M-proteins. Free light chains (FLCs) were measured using the FreeLite® assay. Individuals with a previous diagnosis of MM/LPs/MGUS (N=237) were excluded. Per protocol and informed consent, participants with MGUS were randomized to one of the three study arms: Arm 1 where participants are not contacted; Arm 2 where individuals are followed based on current guidelines; and Arm 3 where individuals are followed with a more intensive diagnostic and monitoring strategy. Participants who progress are offered early treatment. All participants repeatedly answered questionnaires on quality of life and mental health. Results: A total of 75,422 participants (93.4%) provided a serum sample for screening. Of those, 3,725 (4.9%) had MGUS. The prevalence of MGUS was dependent on age with 2.3%, 6.2%, and 12.9% diagnosed in age groups 40-59, 60-79, and 80-103 years, respectively. The prevalence of MGUS was higher in males, 5.9% vs 4.1% (p The RCT includes 3,487 newly diagnosed MGUS individuals with 1164, 1159, and 1164 individuals in arms 1, 2 and 3, respectively (Table). The median age at diagnosis was 69 years in arms 1 and 2, and 70 years in arm 3. Females constituted 45.9% and the isotypes were IgG (50%), IgA (10%), IgM (18%) and biclonal (8%). The median M-protein concentration was 0.34 g/dL. A total of 428 light-chain MGUS cases were randomized. The demographic distribution was well balanced between the three arms. After a median follow-up of 3 years, 194 patients in the RCT have been diagnosed with any LP: 9 in arm 1, 92 in arm 2, and 133 in arm 3 (p Conclusion: In this large prospective population-based screening study including >75,000 screened persons, we have identified 3,725 individuals with monoclonal gammopathy. In the RCT, after 3 years of follow-up, we show that active screening identifies significantly higher number of individuals with full-blown malignancy and smoldering disease, illustrating the fact that early detection and intervention is achievable. Although our findings are encouraging, until final results of the iStopMM study become available, including data on survival and quality of life, we advise against systematic MGUS screening in healthy individuals. Figure 1 Figure 1. Disclosures Kristinsson: Amgen: Research Funding; Celgene: Research Funding. Kampanis: The Binding Site: Current Employment. Hultcrantz: Curio Science LLC: Consultancy; Daiichi Sankyo: Research Funding; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Research Funding; Intellisphere LLC: Consultancy. Durie: Amgen: Other: fees from non-CME/CE services ; Amgen, Celgene/Bristol-Myers Squibb, Janssen, and Takeda: Consultancy. Harding: The Binding Site: Current Employment, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties. Landgren: Janssen: Research Funding; Celgene: Research Funding; Janssen: Honoraria; Janssen: Other: IDMC; Amgen: Honoraria; Takeda: Other: IDMC; Amgen: Research Funding; GSK: Honoraria.

Analytical ultracentrifugation is a versatile approach for analysing the molecular mass, molecular integrity (degradation/aggregation), oligomeric state and association/dissociation constants for self-association, and assay of ligand binding of kinase related membrane proteins and glycans. It has the great property of being matrix free—providing separation and analysis of macromolecular species without the need of a separation matrix or membrane or immobilisation onto a surface. This short review—designed for the non-hydrodynamic expert—examines the potential of modern sedimentation velocity and sedimentation equilibrium and the challenges posed for these molecules particularly those which have significant cytoplasmic or extracellular domains in addition to the transmembrane region. These different regions can generate different optimal requirements in terms of choice of the appropriate solvent (aqueous/detergent). We compare how analytical ultracentrifugation has contributed to our understanding of two kinase related cellular or bacterial protein/glycan systems (i) the membrane erythrocyte band 3 protein system—studied in aqueous and detergent based solvent systems—and (ii) what it has contributed so far to our understanding of the enterococcal VanS, the glycan ligand vancomycin and interactions of vancomycin with mucins from the gastrointestinal tract.

β-D-glucans are proposed to have many health benefits. It is therefore important to have methods which can distinguish these from other carbohydrates present in natural products, as well as giving glucan content and structural information. Correlations between features in the CP/MAS spectra of β-D-glucans and enzyme assay determined β-D-glucan content were generally found to be poor. The β-D-glucan in dry and hydrated forms of the mushroom Ganoderma lucidum was investigated in detail by spectral peak fitting to the anomeric carbon C1 region in CP/MAS NMR spectra. Hydrated samples gave spectra with enhanced resolution and suggested that a clear distinction between β-D-glucans and other carbohydrates could be possible in the anomeric carbon C1 region. Chemical shift values for a range of carbohydrate polymers, which can be found alongside β-D-glucans, as well as the values for various linkages are given. Contamination by other carbohydrates and buffer salts is discussed.

IgG2 is the most efficient subclass for providing protection against pneumococcal pathogens. We hypothesised that some individuals may be unable to mount an effective pneumococcal capsular polysaccharide (PCP) IgG2 response despite having a normal PCP IgG concentration (PCP IgG2 deficient). The median pre-vaccination PCP IgG2 concentration was significantly lower in individuals referred for immunological investigation compared to healthy controls (2.8 mg/L range, 95% CI 1.1-88 vs. 29.5mg/L, 95% CI 13.5-90, p = 0.0002). PCP IgG:IgG2 ratios were significantly higher for the referral population than for healthy controls suggesting the increased production of PCP specific subclasses other than IgG2. The percentage of individuals with PCP IgG2 deficiency was significantly higher in referral groups compared to controls (31% vs. 5%; p = 0.0009) and in an individual with PCP IgG2 deficiency, the balance of PCP specific IgG subclass antibodies post vaccination changed from IgG2>IgG1>IgG3>IgG4 to IgG1>IgG3>IgG2>IgG4. The median PCP IgG2 concentration in those with PCP IgG2 deficiency was significantly lower in the referral groups compared to controls (7.8 mg/L, 95% CI 1.1-12 vs. 12.7 mg/L, 95% CI 11.8-13.1; p = 0.006). The data suggests a defect in the production PCP IgG2 may be present in individuals with normal PCP IgG referred for immunological investigation.

The role of antibody structure (conformation) in solution rheology is probed. It is demonstrated here that pH-dependent changes in the tertiary structure of 2 mAb solutions lead to viscoelasticity and not merely a shear viscosity (η) increase. Steady shear flow curves on mAb solutions are reported over broad pH (3.0 ≤ pH ≤ 8.7) and concentration (2 mg/mL ≤ c ≤ 120 mg/mL) ranges to comprehensively characterize their rheology. Results are interpreted using size exclusion chromatography, differential scanning calorimetry, analytical ultracentrifugation, near-UV circular dichroism, and dynamic light scattering. Changes in tertiary structure with concentration lead to elastic yield stress and increased solution viscosity in solution of “mAb1.” These findings are supported by dynamic light scattering and differential scanning calorimetry, which show increased hydrodynamic radius of mAb1 at low pH and a reduced melting temperature Tm, respectively. Conversely, another molecule at 120 mg/mL solution concentration is a strong viscoelastic gel due to perturbed tertiary structure (seen in circular dichroism) at pH 3.0, but the same molecule responds as a viscous liquid due to reversible self-association at pH 7.4 (verified by analytical ultracentrifugation). Both protein–protein interactions and structural perturbations govern pH-dependent viscoelasticity of mAb solutions.

Okra extract is known to have potential application as a bio-flocculant for wastewater treatment. However, no research to date has given insight into the components responsible for the flocculating ability of okra extract or its flocculating mechanism. The work presented here addresses this knowledge gap showing that pectin, especially pectin homogalacturonan (HGA) regions, appear to be the polysaccharides responsible for the flocculating ability of okra extract. The way pectin works in flocculation may be best explained by a polymer bridging mechanism. Specifically, a linear relationship between okra bio-flocculating ability and pectin homogalacturonan region to rhamnogalacturonan-I region weight ratio (HGA/RG-I) was found (y = 2.0x+47.6, R2 = 0.93, when GalA content > 300 mg/g extract), which was also validated using commercial citrus peel pectin.

We report the novel application of Analytical Ultracentrifugation (AUCF) to characterise the polymeric proanthocyanidin fraction of hops. Extraction of hop samples with 70% acetone (aq) followed by a C-18 Solid Phase Extraction yielded polyphenolic fractions for AUCF analysis. Sedimentation velocity experiments demonstrated the presence of discrete molecular weight bands of proanthocyanidins, as opposed to a continuous distribution of molecular weights. There were 4 such bands for Saaz hop (0.15, 1.1, 2.7 and 4.4S) and 3 bands for Magnum (0.15, 1.6 and 3.0S). The method resulted in a reproducible size (sedimentation coefficient) distribution for replicate runs of the same extract and for extracts prepared from different samples of the same hop variety. Sedimentation equilibrium experiments were then used to fit molecular weight distributions using the new SEDFIT-MSTAR method for the same samples. Thus we report for the first time polymeric proanthocyanidins in hops with molecular weights of up to 100 kDa in Saaz hop (or up to 56 kDa in Magnum). This represents the first application of AUCF to characterise complex fractions of polyphenolics extracted from botanical sources and the methodology developed should find wider application in the study of this diverse and bioactive class of compounds.

Background We sought to compare Pneumovax®23 responses in adults with subnormal IgG subclass concentrations. We studied adults with normal total IgG, frequent/severe respiratory infection, and subnormal IgG1, IgG3, or IgG1 + IgG3 before and after Pneumovax®23. We defined response as serotype-specific IgG > 1.3 μg/mL and aggregate response as IgG > 1.3 μg/mL for ≥70% of all serotypes tested. We compared patients with and without serotype-specific responses and performed logistic regression on aggregate responses using: age; male sex; body mass index; autoimmune condition(s); atopy; other allergies; subnormal IgGSc immunophenotypes; IgA; and IgM. Results There were 59 patients (mean age 44 ± 13 (SD) years; 83.1% women). Median days between pre- and post-Pneumovax®23 testing was 33 (range 19–158). The median post-vaccination summated concentration of serotype-specific IgG was higher in patients with subnormal IgG1 than subnormal IgG3 (responders and non-responders). All subnormal IgG1 + IgG3 non-responders responded to serotypes 8, 9 and 26, unlike other non-responders. Subnormal IgG3 responders had lower responses to serotypes 1, 4, 12, 23, 26, and 51. Subnormal IgG3 non-responders had higher responses to serotypes 1, 3, 8, 9, 12, 14, 19, 51, and 56. Response rates decreased with increasing age. Aggregate responders were: subnormal IgG1, 54%; IgG3, 46%; and IgG1 + IgG3, 46%. Regression on aggregate response revealed lower response with male sex (odds ratio 0.09 [95% CI 0.01, 0.77]) and atopy (0.17 [0.03, 0.83]). Conclusions Serotype-specific IgG responses to Pneumovax®23 were greater in patients with subnormal IgG1 than subnormal IgG3. Male sex and atopy were associated with lower aggregate responses.

Water-in-oil-in-water emulsions containing an internalised salt solution were stabilised with non-chemically modified waxy rice starch (WRS), and octinyl succinic anhydride (OSA) as reference, to release salt during oral processing due to amylase-induced destabilisation. Salt levels were 1.5 g salt and 0.47 g salt per 100 g external and internal aqueous phases, respectively. Variables were the starch content (2, 3, 4 g per 100 g emulsion; 20 g oil per 100 g emulsion), level of polyglycerol polyricinoleate (PGPR) as a lipophilic emulsifier (0.29, 0.57 g per 100 g emulsion) and ambient-pressure processing temperature for WRS gelatinisation, the non-chemical modification process, (75 ± 3, 88 ± 5 °C). OSA starch was used under previously applied conditions (2, 3, 4 g starch, 0.57 g PGPR per 100 g emulsion, 25 ± 5 °C). Emulsions were stable for three months, except OSA and lower level PGPR low temperature processed WRS emulsions lost salt into the external emulsion phase. One day after processing, encapsulation efficiency (EE) was as predicted from the composition for OSA emulsions, while at the same PGPR content an external aqueous phase was incorporated into the oil droplets of the WRS emulsion increasing EE. Salt release was assessed in vitro and through sensory evaluation using paired comparison testing. The results revealed that the efficacy of this salt reduction approach was enhanced for gelatinised WRS compared to OSA starch stabilised emulsions. Consumer tests on a tomato soup, to validate this salt reduction approach for a real food, revealed a possible 25% salt reduction, compared to current UK products.

While a good mucoadhesive biopolymer must adhere to a mucus membrane, it must also have a good unloading ability. Here, we demonstrate that the biopolymer pullulan is partially digested by human salivary α-amylase, thus acting as a controlled release system, in which the enzyme triggers an increased release of flavour. Our oral processing simulations have confirmed an increase in the bioavailability of aroma and salt compounds as a function of oral pullulan degradation, although the release kinetics suggest a rather slow process. One of the greatest challenges in flavour science is to retain and rapidly unload the bioactive aroma and taste compounds in the oral cavity before they are ingested. By developing a cationic pullulan analogue we have, in theory, addressed the “loss through ingestion” issue by facilitating the adhesion of the modified polymer to the oral mucus, to retain more of the flavour in the oral cavity. Dimethylaminoethyl pullulan (DMAE-pullulan) was synthesised for the first time, and shown to bind submaxillary mucin, while still retaining its susceptibility to α-amylase hydrolysis. Although DMAE-pullulan is not currently food grade, we suggest that the synthesis of a sustainable food grade alternative would be a next generation mucoadhesive targeted for the oral cavity.

Highlights • Conventional and microwave-assisted extraction of “hairy” pectin from sugar beet. • Determined effect of heating method, temperature, time & pH on yield & composition. • No difference between microwave and conventional extraction under conditions tested. • Strong alkaline is favoured in rhamnogalacturonan-I “hairy” pectin extraction. • Hydrothermal water extraction can be an alternative to strong alkaline extraction., Sugar beet pectin is rich in rhamnogalacturonan-I (RG-I) region, which is a potential source of prebiotics. RG-I pectin cannot be extracted the same way as commercial homogalacturan-rich pectin using hot acid. Therefore, this study has explored several alternative methods, including microwave-assisted extraction (MAE) and conventional-solvent extraction (CSE) at atmospheric pressure using different solvents, and microwave-assisted hydrothermal extraction (MAHE) under pressure using water. No conclusive differences in microwave and conventional heating were found with heating rate controlled. The optimum treatment times of both MAE and CSE at 90 °C atmospheric pressure and regardless of the solvents used were 120 min; however, MAHE at 130 °C under pressure can dramatically reduce the time to 10 min. Alcohol-insoluble solids (AIS) extracted using pH13 solvent by MAE had both the highest RG-I yield at 25.3% and purity at 260.2 mg/g AIS, followed by AIS extracts using water by MAHE with 7.5% and 166.7 mg/g AIS respectively.