Low Dose β-carotene Supplementation Diminishes Oxidative Stress in Type 2 Diabetics and Healthy Individuals

• Changes in oxidative status [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  

• Hemoglobin and hematocrit [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  
• Ferritin [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Enzyme linked immunosorbent assay (ELISA) with monoclonal antibodies
  
  
• Iron metabolism markers [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Serum iron, total iron binding capacity (TIBC) and unsaturated iron binding capacity (UIBC) were determined by the methods proposed by the International Committee of Standardization of Hematology.
  
  
• Blood Chemistry [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Glycemia, triglycerides, total cholesterol, LDL, and HDL were determined automatically in a Ciba Corning 550 Express autoanalizer, using classic enzymatic methods for the determination of these variables.
  
  
• Glycosylated Hemoglobin [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  It was determined using a commercial kit (Bioscience, Caracas, Venezuela),
  
  
• Oxidized LDL [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Analyzed by a solid phase two-site enzyme immunoassay from Mercodia (Sweden), which contains 2 monoclonal antibodies directed against separated antigenic determinants on the oxidized apolipoprotein B molecule.
  
  
• Thiobarbituric Acid Reactive substances (TBARS) [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Were detected by the quantification of malondialdehyde present in the sample, by reacting 2 molecules of thiobarbituric acid with 1 molecule of malondialdehyde, which produces an abduct that is detected at 535 mn.
  
  
• Ferric Reducing ability of Plasma (FRAP). [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Measured after 4 and 10 min incubation, was used to determine the ability of plasma to reduce iron from ferric to ferrous state, based on the formation of a triazine-Fe+3 complex, that when reduced to Fe+2, generate a change in color that is measured at 593 nm.
  
  
• Activities of the enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPx). [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Determined by commercial kits (Cayman Chemicals, Pittsburg) following the recommended protocols
  
  
• Serum zinc and copper. [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  By flame atomic absorption spectrophotometry
  
  
• β-carotene. [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  It was determined by HPLC, with a reverse fase C18 column.
  
  
• Serum retinol [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  It was determined by HPLC, with a reverse fase C18 column, as an indirect measure of betacarotene metabolism.
  
  
• Serum nitrites [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  Were determined as an indirect measure of the concentration of nitric oxide, since nitrites are the stable end products of its degradation. Nitrates were reduced to nitrites by activated cadmium. Then sulfanilamide and nitrites generate a chromophore that reacts with naftilethylenediamine, to generate a product visible at 540 nm.
  
  
• Serum and erythrocyte folates. [ Time Frame: Time 0, 45 days and 75 days after supplementation ] [ Designated as safety issue: No ]
  The method is based in the folate-dependent controlled growth of a Lactobacillus strain that is measured spectrophotometrically and quantified against a standard curve.
  
  

Type 2 diabetes is a chronic, multifactorial disease, and oxidative stress one of the pathophysiological mechanisms associated with its appearance and development. The objective was to determine the effect of supplementation with β-carotene to type 2 diabetics and healthy individuals, on iron metabolism, oxidative balance, and antioxidant plasma capacity, using doses similar to the daily nutritional requirement.

A total of 117 volunteers participated in the study. Type 2 diabetics (34) and healthy individuals (24), received 6 mg β-carotene for 45 d, and were compared to similar non-supplemented diabetic (33) and control (26) groups. Blood samples were taken at the beginning, end and 30 days after finishing supplementation, to determine hemoglobin, hematocrit unsaturated iron binding capacity, total iron binding capacity, transferrin saturation, ferritin, glycemia, glycosylated hemoglobin, cholesterol, triglycerides, HDL, LDL, oxidized LDL, copper, zinc, TBARS, FRAP, nitrites, GPx, SOD, folates, retinol and β-carotene.