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

Study Overview
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Brief Summary
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Since diabetes has multiple etiologies and oxidative stress one of the proposed mechanisms, the objective is 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.

Detailed Description
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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.

Official Title
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Effect of the Supplementation With β-carotene to Type 2 Diabetic Patients and Healthy Controls on the Iron Status and Antioxidant Capacity of Plasma

Conditions
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Diabetes Mellitus, Oxidative Stress, Iron Metabolism Disorders

Intervention / Treatment
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* Dietary Supplement: Betacarotene
* Dietary Supplement: Controls. No treatment


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Patients with a diagnose of Type 2 diabetes mellitus of at least 5 years of diagnosis, in treatment with oral hypoglycemics Patients in regular control (once a month) in the Hospital Exclusion Criteria: Hospitalized patient Diabetic patient with diabetes related acute complications (ketoacidosis, hyperosmolar coma)in the 3 months previous to the study. Individuals with infections that required antibiotics in the 3 weeks previous to the study. Individuals with antibodies anti-insulin, autoimmune diseases or in treatment with immunosuppressive drugs. Individuals with viral infections such as hepatitis B, hematological, renal or hepatic diseases. Pregnancy

Ages Eligible for Study
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Minimum Age: 18 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Basic Science
Allocation: Randomized
Intervention Model: Parallel Assignment
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: Supplemented Diabetics (DB)<br>Diabetics supplemented with betacarotene for 45 days | Dietary Supplement: Betacarotene<br>* 6 mg betacarotene in caplets for 45 days (daily)and reevaluate parameters 30 days after finishing supplementation<br>* Other names: β-carotene as a soft gel capsule (GNC, Pennsylvania-USA);|
| Experimental: Unsupplemented Diabetics (DS)<br>Diabetics without betacarotene supplementation | Dietary Supplement: Controls. No treatment<br>* Evaluate at time 0, 45 days and 75 days, but without receiving betacarotene supplements<br>* Other names: No supplements;|
| Active Comparator: Supplemented Controls (CB)<br>Controls supplemented with betacarotene for 45 days | Dietary Supplement: Betacarotene<br>* 6 mg betacarotene in caplets for 45 days (daily)and reevaluate parameters 30 days after finishing supplementation<br>* Other names: β-carotene as a soft gel capsule (GNC, Pennsylvania-USA);|
| Active Comparator: Unsupplemented Controls (CS)<br>Controls without betacarotene supplementation | Dietary Supplement: Controls. No treatment<br>* Evaluate at time 0, 45 days and 75 days, but without receiving betacarotene supplements<br>* Other names: No supplements;|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Changes in oxidative status |  | Time 0, 45 days and 75 days after supplementation | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Hemoglobin and hematocrit |  | Time 0, 45 days and 75 days after supplementation | 
| Ferritin | Enzyme linked immunosorbent assay (ELISA) with monoclonal antibodies | Time 0, 45 days and 75 days after supplementation | 
| Iron metabolism markers | 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. | Time 0, 45 days and 75 days after supplementation | 
| Blood Chemistry | 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. | Time 0, 45 days and 75 days after supplementation | 
| Glycosylated Hemoglobin | It was determined using a commercial kit (Bioscience, Caracas, Venezuela), | Time 0, 45 days and 75 days after supplementation | 
| Oxidized LDL | 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. | Time 0, 45 days and 75 days after supplementation | 
| Thiobarbituric Acid Reactive substances (TBARS) | 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. | Time 0, 45 days and 75 days after supplementation | 
| Ferric Reducing ability of Plasma (FRAP). | 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. | Time 0, 45 days and 75 days after supplementation | 
| Activities of the enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPx). | Determined by commercial kits (Cayman Chemicals, Pittsburg) following the recommended protocols | Time 0, 45 days and 75 days after supplementation | 
| Serum zinc and copper. | By flame atomic absorption spectrophotometry | Time 0, 45 days and 75 days after supplementation | 
| β-carotene. | It was determined by HPLC, with a reverse fase C18 column. | Time 0, 45 days and 75 days after supplementation | 
| Serum retinol | It was determined by HPLC, with a reverse fase C18 column, as an indirect measure of betacarotene metabolism. | Time 0, 45 days and 75 days after supplementation | 
| Serum nitrites | 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. | Time 0, 45 days and 75 days after supplementation | 
| Serum and erythrocyte folates. | The method is based in the folate-dependent controlled growth of a Lactobacillus strain that is measured spectrophotometrically and quantified against a standard curve. | Time 0, 45 days and 75 days after supplementation | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Type 2 diabetes, β-carotene, Oxidative balance, Oxidants, Antioxidants