Marta Bianchi

Rapid Fluorimetric Method to Detect Total Plasma Malondialdehyde with Mild Derivatization Conditions

Malondialdehyde (MDA), an oxidation product of polyunsaturated fatty acids, is used as an in vivo marker to assess lipid peroxidation in diseases such as atherosclerosis and diabetes (1)(2)(3)(4). In biological matrixes, MDA is measured after derivatization with thiobarbituric acid (TBA) (5). Because TBA reacts with many other aldehydes (6), results are expressed as TBA-reactive substances (TBARS). Several problems are associated with TBARS analysis, in particular, low reproducibility and a lack of specificity that leads to overestimations. To overcome these difficulties, more specific methods have been proposed that require sample pretreatment to precipitate proteins and extract MDA-reactant adducts (6)(7)(8)(9). This additional step is time-consuming and adversely affects precision. The aim of the present study was to develop a rapid and sensitive method to measure MDA in plasma, avoiding sample pretreatment. Tetraethoxypropane (TEP), TBA, and bilirubin were obtained from Fluka. Fatty acid-free bovine serum albumin (BSA), Total Protein Reagent, and Protein Standard were from Sigma, and 2,2′-azobis(2-amidinopropane) (ABAP) was from Wako. We prepared an aqueous stock solution of 1 mmol/L TEP. A 10 μmol/L MDA solution was obtained by diluting TEP in 0.1 mol/L HCl. A 0.025 mol/L TBA solution was prepared daily by dissolving TBA in water. BSA solutions were prepared in 0.1 mol/L HCl. To remove protein-bound MDA, BSA solutions were heated at 80 °C for 1 h and dialyzed for 3 days against 0.1 mol/L HCl in a 3500-Da cutoff dialysis membrane (Spectrapore; Spectrum Medical Industries). The actual protein concentration was verified using the Total Protein Reagent Kit. A 50 mmol/L ABAP solution was prepared in water. Six MDA solutions ranging from 0.05 to 0.5 μmol/L were prepared by diluting the 10 μmol/L stock solution with 0.1 mol/L HCl. Triplicate solutions were used to obtain a dose–response …

A fluorescence-based method for the detection of adhesive properties of lactic acid bacteria to Caco-2 cells

Aims:  The ability of probiotic micro‐organisms to adhere to the intestinal surface is regarded as a substantial advantage in terms of bacteria persistence in the gastrointestinal tract. The aim of the present study was the development of a method based on fluorescent staining of bacteria and subsequent spectrofluorimetric detection to quantify the adhesion of several strains of Lactobacillus and Bifidobacterium to Caco‐2 cells.

In Vitro Bioaccessibility of Phenolics and Vitamins from Durum Wheat Aleurone Fractions

Durum wheat aleurone, thanks to its nutrient-rich composition, might be of potential use as a functional ingredient in cereal-based foods provided nutrients can be made available for absorption. We evaluated the in vitro bioaccessibility of thiamine, niacin, and phenolic acids in different aleurone fractions obtained with an industrial processing aimed to obtain material of different composition and particle size. Results indicate that the main phenolic compounds and vitamins investigated have a higher bioaccessibility when present in the inner part of the aleurone layer compared to the outer part of aleurone or the unfractionated bran. Moreover, an ultramicronization treatment employed to reduce particle size does not further improve the bioaccessibility of these compounds. We conclude that aleurone fractions from durum wheat bran could represent a nutritionally relevant ingredient, bringing together a high fiber content and an excellent bioaccessibility of vitamins and phytochemicals generally associated with nutritional benefits.

Ability of a high-total antioxidant capacity diet to increase stool weight and bowel antioxidant status in human subjects

There is limited knowledge about the possible effect of unabsorbed dietary antioxidants that reach the large intestine on bowel habits. The aim of the present study was to investigate whether a dietary recommendation directed to increase diet total antioxidant capacity (TAC) is able to affect gut function in human subjects. In this cross-over intervention, nineteen subjects followed a high-TAC (HT) and a low-TAC (LT) diet for 2 weeks, which were comparable for energy, macronutrient, total dietary fibre and alcohol contents. At the end of each intervention period, the 48 h stool output was recorded. In the faecal samples obtained from a subset of nine subjects, moisture, pH, ammonia content, Lactobacillus and Bifidobacterium counts, faecal water antioxidants and genotoxicity were measured. A 3 d weighed food record was used to assess the diet composition during HT and LT diet intake. Significant increases in the intake of TAC, vitamins E and C and phenolic compounds were observed during the HT diet intake. The higher intake of antioxidants led to increased 48 h stool output (324 (SD 38) g in HT v. 218 (SD 22) g in LT), and to higher TAC and total phenolic concentrations in faecal water. No significant variation in the other measured parameters was observed between the diets. In conclusion, a diet selected to raise the intake of dietary antioxidants is able to increase stool bulk and antioxidant content of faeces.

Intervention study with a high or low antioxidant capacity diet: effects on circulating β-carotene.

Background and objectives:A cross-sectional observation suggests that total antioxidant capacity (TAC) of the diet positively affects plasma concentrations of β-carotene independent of β-carotene intake. This study was carried out to investigate the effect of two dietary strategies, designed to be comparable in fruits, vegetables, fibre, alcohol and β-carotene intake but substantially different in their TAC, on changes in antioxidant intake and antioxidant status, and in particular in circulating β-carotene concentrations.Subjects:A randomized cross-over intervention trial involving 33 healthy participants and consisting of two 14-day dietary periods (high TAC diet, HT; low TAC diet, LT) with a 14-day washout in between was conducted.Results:Energy, macronutrient, dietary fibre, alcohol and β-carotene intake was not significantly different between LT and HT, whereas intake of other carotenoids and dietary TAC was significantly higher in the HT than in the LT (P<0.001). Circulating carotenoids (with the exception of α-carotene, which followed an inverse trend) and α-tocopherol decreased significantly during the LT and increased during the HT period. Among these, β-carotene almost doubled its concentration in plasma after the HT diet.Conclusions:The increase in circulating β-carotene along with the increase in dietary TAC suggests that plasma β-carotene could be a marker of TAC intake rather than of β-carotene intake itself. This may explain, in part, why β-carotene supplementation alone has shown no benefit in chronic disease prevention and adds to a putative beneficial role of high dietary TAC diets, which merits further investigation.

Wheat aleurone polyphenols increase plasma eicosapentaenoic acid in rats

Methods These studies were designed to assess whether wheat polyphenols (mainly ferulic acid [FA]) increased the very-long-chain omega-3 fatty acids (VLC n-3) [eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] in rats. Wheat aleurone (WA) was used as a dietary source of wheat polyphenols. Two experiments were performed; in the first one, the rats were fed WA or control pellets (CP) in presence of linseed oil (LO) to provide alpha-linolenic acid (ALA), the precursor of VLC n-3. In the second one, the rats were fed WA or CP in presence of control oil (CO) without ALA. The concentrations of phenolic acid metabolites in urine were also investigated. Results The urinary concentration of conjugated FA increased with WA ingestion (p<0.05). Plasma EPA increased by 25% (p<0.05) with WA in the CO group but not in the LO group. In contrast, there was no effect of WA on plasma DHA and omega-6 fatty acids (n-6). Finally, both n-3 and n-6 in the liver remained unchanged by the WA. Conclusion These results suggest that WA consumption has a significant effect on EPA in plasma without affecting n-6. Subsequent studies are required to examine whether these effects may explain partly the health benefits associated with whole wheat consumption.