Skin carotenoids status as a potential surrogate marker for cardiovascular disease risk determination in middle-aged and older adults.

BACKGROUND AND AIMS: Upon consumption, carotenoids, which may attenuate cardiovascular disease (CVD) risk, diffuse from the blood and accumulate in the skin. This study aimed to assess the associations between dietary, plasma, and skin carotenoids with CVD risk indicators and to examine the mediational role of plasma carotenoids in the relationship between skin carotenoids status (SCS) and CVD risk. METHODS AND RESULTS: Dietary, plasma, and skin carotenoids were assessed in a cross-sectional study from a community in Singapore (n = 103) aged 50 to 75 y. Multiple linear regression and binary logistics regression models were used to examine the associations between the carotenoids status with classical CVD risk factors and composite CVD risk indicators. After controlling for covariates, SCS and plasma carotenoids were inversely associated with systolic blood pressure (skin: P < 0.001; plasma: P < 0.05) and diastolic blood pressure (skin: P < 0.001; plasma: P < 0.005). Additionally, each increment of 1000 in SCS was associated with an odds ratio of 0.924 (P < 0.01) for metabolic syndrome diagnosis and 0.945 (P < 0.05) for moderate to high CVD risk classification. Associations between SCS and composite CVD risk indicators were null when adjusted for the corresponding plasma carotenoids, indicating complete mediation. Dietary carotenoids, however, showed no relationship with the CVD risk indicators. CONCLUSION: Carotenoids bioavailability may be important for cardiovascular protection. SCS, driven by the corresponding plasma carotenoids, could be a potential noninvasive surrogate marker for CVD risk determination in middle-aged and older adults. CLINICAL TRIAL REGISTRATION: NCT03554954, https://clinicaltrials.gov/. TRIAL REGISTRATION DATE: 13 June 2018.

Wolfberry (Lycium barbarum) Consumption with a Healthy Dietary Pattern Lowers Oxidative Stress in Middle-Aged and Older Adults: A Randomized Controlled Trial.

Incorporating zeaxanthin-rich wolfberry (Lycium barbarum) into a healthy dietary pattern may augment its antioxidant potential. The present 16-week, parallel design randomized controlled trial aimed to investigate the impact of adhering to a healthy dietary pattern, either with or without whole dried wolfberry (15 g/d) on oxidative stress status (plasma malondialdehyde and 8-iso-prostaglandin F2α) in middle-aged and older adults. Changes to carotenoids status (plasma and skin carotenoids) and body composition were further evaluated to explore potential mechanisms which underlie the antioxidant properties of wolfberry. Plasma 8-iso-prostaglandin F2α, plasma zeaxanthin and skin carotenoids status were significantly raised in the wolfberry consuming group (n = 22; p < 0.05) compared to the control group which showed no changes (n = 18). Likewise in the wolfberry group only, inverse association was observed between the change values of plasma zeaxanthin and plasma 8-iso-prostaglandin F2α (-0.21 (-0.43, 0.00) ng/µmol, regression coefficient (95% CI); p = 0.05). Wolfberry consumption with a healthy dietary pattern may serve as a dietary strategy to attenuate lipid peroxidation among middle-aged and older adults who are at a heightened risk of oxidative stress induced age-related disorders. The antioxidant properties of wolfberry may be attributed to its rich zeaxanthin content.

Analytical method development for directed enzyme evolution research: a high throughput high-performance liquid chromatography method for analysis of ribose and ribitol and a capillary electrophoresis method for the separation of ribose enantiomers.

Analytical methods were developed for a directed enzyme evolution research programme, which pursued high performance enzymes to produce high quality L-ribose using large scale biocatalytic reaction. A high throughput HPLC method with evaporative light-scattering detection was developed to test ribose and ribitol in the enzymatic reaction, a ß-cyclobond 2000 analytical column separated ribose and ribitol in 2.3 min, a C(18) guard column was used as an on-line filter to clean up the enzyme sample matrix and a short gradient was applied to wash the column, the enzymatic reaction solution can be directly injected after quenching. Total run time of each sample was approx. 4 min which provided capability of screening 4×96-well plates/day/instrument. Meanwhile, a capillary electrophoresis method was developed for the separation of ribose enantiomers, while 7-aminonaphthalene-1,3-disulfonic acid was used as derivatisation reagent and 25 mM tetraborate with 5 mM ß-cyclodextrin was used as electrolyte. 0.35%of D-ribose in L-ribose can be detected which can be translated into 99.3% ee of L-ribose. Derivatisation reagent and sample matrix did not interfere with the measurement.

Void exclusion of antibodies by grafted-ligand porous particle anion exchangers.

We describe a new variant of anion exchange chromatography in columns packed with porous particles that embody charged low-density polymer zones supported by a higher density polymer skeleton. IgG defies the norms of anion exchange and is excluded to the void volume at pH 3-10 and 0-4M NaCl. Void exclusion also occurs with Fab, F(ab')2, and IgM. Host cell protein contaminants mostly follow the usual norms of anion exchange and bind more strongly with increasing pH and decreasing conductivity. Sample buffer composition has no impact on partitioning so long as applied sample volume does not exceed the interparticle void volume of the column. Void-excluded antibody elutes in equilibration buffer. This seemingly conflicted collection of behaviors is reconciled by a variable size exclusion function mediated through the low-density polymer zones, the charge properties of the antibody species, and the pH and conductivity of the equilibration buffer. Current-generation porous particle anion exchangers that employ grafting techniques to achieve high charge density mediate void exclusion to varying degrees, with the best-suited achieving complete exclusion, and others as little as 65%. Perfusive and non-grafted particle-based exchangers mediate as little as 50% exclusion. Monoliths mediate no exclusion, due to their lack of an interparticle void volume. On qualified exchangers, the technique supports greater than 99% reduction of host proteins, DNA, and endotoxin. Virus is reduced more than 99.9%, and aggregates are reduced to less than 0.05%. The method supports better process control than other anion exchange formats because pH excursions in conjunction with changes in salt concentration do not occur until after the antibody has eluted from the column.

Characterization and removal of aggregates formed by nonspecific interaction of IgM monoclonal antibodies with chromatin catabolites during cell culture production.

We observed that IgM monoclonal antibodies and aggregates in mammalian cell culture supernatants were associated nonspecifically with nucleosomes, DNA, and histone proteins derived from nuclei of host cells that died during antibody production. A series of multimodal sample treatments were evaluated for their ability to selectively remove these contaminants without significant antibody loss. The first consisted of adding 2,5-dioxo-4-imidazolidinyl urea (allantoin) and the DNA intercalating agent 7-ethoxyacridine-3,9-diamine (ethacridine), then flowing the supernatant through a column of mixed porous particles bearing metal affinity, anion exchange, and cation exchange functionalities. A one-step variant of the method was to mix chromatography particles with the allantoin-ethacridine-treated supernatant. An alternative one-step treatment consisted of passing untreated cell supernatant through a chelating monolith in tandem with an anion exchange monolith. All methods eliminated high molecular weight aggregates, and reduced smaller aggregates to 2-4%. They also achieved 98% DNA reduction, 99% reduction of nucleosomes and histones, 30-70% reduction of general host proteins, and 98% IgM recovery. Size exclusion chromatography analysis indicated that IgG monoclonal antibodies benefit similarly from treatment. Subsequent IgM purification reduced DNA levels beneath the level of detectability by fluorescent dye intercalation, histones to less than 10 parts per million by ELISA, and aggregates to less than 0.05% by size exclusion chromatography. The results point to chromatin catabolites as promoters of antibody aggregate formation.

Principles and applications of steric exclusion chromatography.

We introduce a chromatography method for purification of large proteins and viruses that works by capturing them at a non-reactive hydrophilic surface by their mutual steric exclusion of polyethylene glycol (PEG). No direct chemical interaction between the surface and the target species is required. We refer to the technique as steric exclusion chromatography. Hydroxyl-substituted polymethacrylate monoliths provide a hydrophilic surface and support convective mass transport that is unaffected by the viscosity of the PEG. Elution is achieved by reducing PEG concentration. Selectivity correlates with molecular size, with larger species retained more strongly than smaller species. Retention increases with PEG size and concentration. Salts weaken retention in proportion to their concentration and Hofmeister ranking. Retention is enhanced near the isoelectric point of the target species. Virus binding capacity was measured at 9.9×10(12) plaque forming units per mL of monolith. 99.8% of host cell proteins and 93% of DNA were eliminated. Mass recovery exceeded 90%. IgM capacity was greater than 60 mg/mL. 95% of host cell proteins were eliminated from IgM produced in protein-free media, and mass recovery was up to 90%. Bioactivity was fully conserved by both viruses and antibodies. Process time ranged from less than 30 min to 2 h depending on the product concentration in the feed stream.