Oxidation levels of North American over-the-counter n-3 (omega-3) supplements and the influence of supplement formulation and delivery form on evaluating oxidative safety.

The aim of the present study was to evaluate the oxidation status of North American n-3 (omega-3) PUFA nutritional supplements commercially available in Canada and evaluate the influence of product formulation and delivery form on oxidative safety. A total of 171 North American over-the-counter n-3 PUFA nutritional supplements were analysed for oxidation safety. Primary and secondary oxidation and total oxidation (TOTOX) were determined using the American Oil Chemists' Society (AOCS) procedures. Comparisons between supplements' final forms, oil source and n-3 PUFA concentration quartiles, as measures of product formulations and delivery forms, were compared using ANOVA. Of the products successfully tested, 50 % exceeded the voluntary recommended levels for markers of oxidation. Another 18 % of products were approaching the limits with 1-3 years before expiration. Encapsulated products without flavour additives had significantly lower secondary and TOTOX levels than bulk oils and flavoured products (P < 0·05). Children's products had significantly higher primary, secondary and TOTOX levels compared with all other products (P < 0·05). Markers of oxidation did not differ between oil sources (P > 0·05), with the exception of krill oil products having higher secondary oxidation levels than plant-based products (P > 0·05). Markers of oxidation did not differ between n-3 PUFA supplement concentration quartiles. Consumers may be at risk of exposure to higher levels of oxidative products. New regulatory mandates need to be introduced to ensure that all n-3 PUFA products, used as nutritional supplements, regardless of their formulation or delivery form, can be tested for oxidative safety and compliance.

Efficacy of a unique omega-3 formulation on the correction of nutritional deficiency and its effects on cardiovascular disease risk factors in a randomized controlled VASCAZEN(®) REVEAL Trial.

Low blood levels of long chain omega-3 polyunsaturated fatty acids (LC n-3 PUFA) have been reported to be associated with increased risk for cardiovascular disease (CVD) deaths. Systematic studies measuring LC n-3 PUFA blood levels (pre and post-treatment) in defined subjects, and monitoring the correction of nutritional deficiency with a pure LC n-3 PUFA formulation in sufficient doses, while monitoring CVD risk factors are lacking. We tested the efficacy of a novel LC n-3 PUFA Medical Food formulation (VASCAZEN(®), > 90 % pure with a 6:1 eicosapentaenoic acid-(EPA):docosahexaenoic acid-(DHA) ratio; 6:1-OM3), to correct such deficiency and determine the concomitant effects on lipid profiles. Of 655 subjects screened, 89 % were LC n-3 PUFA deficient (Omega-Score, (OS) = blood EPA + DHA + Docosapentaenoic acid < 6.1 %). From these, a study was conducted on 110 ambulatory cardiovascular subjects. Placebo: corn oil. Primary endpoint: change in OS. Secondary endpoint: changes in blood lipid profiles. At 8 weeks of treatment with 6:1-OM3 (4 g/day), placebo-adjusted median OS levels (n = 56) significantly improved (132 %, P < 0.0001) with a decrease in AA (arachidonic acid): EPA ratio (82 %, P < 0.0001). In hypertriglyceridemic subjects (TG 2.26-5.65 mmol/L), HDL-C improved (9 %, P = 0.0069), TG-reduced (48 %, P < 0.0001), and VLDL-C reduced (30 %, P = 0.0023), without significantly affecting LDL-C levels. This study confirms that LC n-3 PUFA deficiency is prevalent in the US population, and its correction with 6:1-OM3 in CVD subjects improves lipid profiles. The purity, EPA:DHA ratio and dose are determinant factors for optimal efficacy of a formulation in reducing CVD risk factors.

Human serum proteins preseparated by electrophoresis or chromatography followed by tandem mass spectrometry.

Electrophoretic and chromatographic sample preparations were compared and together detected the presence of some 600 types of protein products in human serum. Proteins from crude serum preseparated by ionic electrophoresis, chromatography, or a combination of both were analyzed. Proteins were digested with trypsin or chymotrypsin. Naturally occurring peptides were also collected by reversed-phase chromatography. The resulting peptides were identified by tandem mass spectrometry. The peptides were either desorbed by a laser from a metal chip into a quadrupole-time-of-flight mass spectrometer or ionized as an electro-spray from reversed-phase chromatography via a metal needle under voltage into an ion-trap mass spectrometer. All of the commonly known proteins associated with serum were detected, and the two mass spectrometers agreed on the identity of abundant serum proteins. Preseparation of serum proteins prior to digestion markedly enhanced the capacity to detect un-common proteins from blood. Electrophoretic- and chromatography-based experiments were found to be complementary. Many novel cellular proteins not previously associated with serum were recorded.

Mining biomarkers in human sera using proteomic tools.

One of the major difficulties in mining low abundance biomarkers from serum or plasma is due to the fact that a small number of proteins such as albumin, alpha2-macroglobulin, transferrin, and immunoglobulins, may represent as much as 80% of the total serum protein. The large quantity of these proteins makes it difficult to identify low abundance proteins in serum using traditional 2-dimensional electrophoresis. We recently used a combination of multidimensional liquid chromatography and gel electrophoresis coupled to matrix-assisted laser desorption/ionization-quadrupole-time of flight and Ion Trap liquid chromatography-tandem mass spectrometry to identify protein markers in sera of Alzheimer's disease (AD), insulin resistance/type-2 diabetes (IR/D2), and congestive heart failure (CHF) patients. We identified 8 proteins that exhibit higher levels in control sera and 36 proteins that exhibit higher levels in disease sera. For example, haptoglobin and hemoglobin are elevated in sera of AD, IR/D2, and CHF patients. The levels of several other proteins including fibrinogen and its fragments, alpha 2-macroglobulin, transthyretin, pro-platelet basic protein, protease inhibitors clade A and C, as well as proteins involved in the classical complement pathway such as complement C3, C4, and C1 inhibitor, were found to differ between IR/D2 and control sera. The sera levels of proteins, such as the 10 kDa subunit of vitronectin, alpha 1-acid glycoprotein, apolipoprotein B100, fragment of factor H, and histidine-rich glycoprotein were observed to be different between AD and controls. The differences observed in these biomarker candidates were confirmed by Western blot and the enzyme-linked immunosorbent assay. The biological meaning of the proteomic changes in the disease states and the potential use of these changes as diagnostic tools or for therapeutic intervention will be discussed.

Processing of serum proteins underlies the mass spectral fingerprinting of myocardial infarction.

The MALDI-TOF spectra of peptides from the sera of normal and myocardial infarction patients produced patterns that provided an accurate diagnostic of MI. In myocardial infarction, the spectral pattern originated from the cleavage of complement C3 alpha chain to release the C3f peptide and cleavage of fibrinogen to release peptide A. The fibrinogen peptide A and complement C3f peptide were in turn progressively truncated by aminopeptidases to produce two families of fragments that formed the characteristic spectral pattern of MI. Time course and inhibitor studies demonstrated that the peptide patterns in the serum reflect the balance of disease-specific-protease and aminopeptidase activity ex vivo.

Autoimmune islet destruction in spontaneous type 1 diabetes is not beta-cell exclusive.

Pancreatic islets of Langerhans are enveloped by peri-islet Schwann cells (pSC), which express glial fibrillary acidic protein (GFAP) and S100beta. pSC-autoreactive T- and B-cell responses arise in 3- to 4-week-old diabetes-prone non-obese diabetic (NOD) mice, followed by progressive pSC destruction before detectable beta-cell death. Humans with probable prediabetes generate similar autoreactivities, and autoantibodies in islet-cell autoantibody (lCA) -positive sera co-localize to pSC. Moreover, GFAP-specific NOD T-cell lines transferred pathogenic peri-insulitis to NOD/severe combined immunodeficient (NOD/SCID) mice, and immunotherapy with GFAP or S100beta prevented diabetes. pSC survived in rat insulin promoter Iymphocytic choriomeningitis virus (rip-LCMV) glycoprotein/CD8+ T-cell receptor(gp) double-transgenic mice with virus-induced diabetes, suggesting that pSC death is not an obligate consequence of local inflammation and beta-cell destruction. However, pSC were deleted in spontaneously diabetic NOD mice carrying the CD8+/8.3 T-cell receptor transgene, a T cell receptor commonly expressed in earliest islet infiltrates. Autoimmune targeting of pancreatic nervous system tissue elements seems to be an integral, early part of natural type 1 diabetes.