A double-blinded, randomized, placebo-controlled study evaluating the impact of dates vinegar consumption on blood biochemical and hematological parameters in patients with type 2 diabetes

Purpose: To determine the effects of dates vinegar on blood biochemical and hematological parameters in type 2 diabetic subjects. Methods: Current research focused on fifty-five subjects having blood sugar more than126 mg/dL. Participants ingested dates vinegar (20 mL) daily into their normal diets for a period of 10 weeks. Glycated hemoglobin (HbA1c), fasting blood sugar (FBS), total cholesterol, high-density lipoprotein (HDL), low-density lipoprotein (LDL), creatinine (Cr), urea, complete blood count (CBC), alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), potassium and folate levels were analyzed before, after 5 weeks and after the experiment Results: Dates vinegar improved the blood concentrations of HbA1c (6.80 ±2.34 to 6.17 ± 2.14 (%)), FBS (171.43 ±36.74 to 147.56 ± 38.86 mg/dL,p=0.001), TC (218.10 ± 16.9 to 191.14 ± 14.23 mg/dL, p<0.001), ALT (24.94 ± 5.03 to 21.88±5.08 IU/L, p=0.002) and ALP (264.32± 45.26 to 257.30 ±44.21 IU/L) and folate (34.6 ± 6.6 to 41.7 ± 6.5 nmol/ L,p<0.001). Conclusion: Dates vinegar significantly improved the total cholesterol.The other blood biochemical and hematological factors were also improved however; the improvements were not significant.


INTRODUCTION
Diabetes mellitus (DM) is a universally emergent health complication and is one of the five main causes of mortality and morbidity in many parts of the world [1].Globally, more than 171 million people were affected by diabetes in 2000 and this figure is projected to reach 366 million by 2030 [2].Impaired Insulin resistance, insulin secretion and hepatic glucose over-creation are possibly related causes of diabetes [3].Insulin resistance do not merely play a critical role in diabetic problems, but also in dyslipidemia, atherosclerosis and hypertension [4].Generally, diabetes mellitus is involved in various organ impairment linked disorders including cataract, atherosclerosis, retinopathy, nephropathy and wound restorative impedance [5].Because of cost of therapy with the various hypoglycemic reagents and insulin commonly used to treat the disease and the unwanted side effects associated with these medications, dietary control is recommended among one of various approaches and techniques required to prevent the diseases.Results of different studies revealed that some plants and their byproducts are useful in treating chronic diseases such as diabetes, cardiovascular disease and cancer [6].Acetic acid is considered as main component of vinegar and it is responsible for its biting odor, tart flavor and pungent smell [7,8].Acetate thiokinase, changes the acetic acid to acetyl-coA and is consequently assimilated as citrate.In vitro, the activity of phosphofructokinase is inhibited by citrate but in vivo, citrate increase the repletion of glycogen after meticulous exercise [9].In 1988, it was revealed for first time that vinegar can be used to cure diabetes.Different animal trials showed that consumption of different doses of acetic acid or vinegar produced fluctuating effects from reduction of hypertension and improvement of glycogen repletion to activation of calcium captivation [10,11].One study showed that white vinegar decreased both insulin levels and postprandial blood glucose [12].Similarly, another study revealed that intake of apple cider vinegar enhances postprandial insulin sensitivity [13].Dates vinegar has a variety of flavones and other polyphenols.Dates contain high quantity of anthocyanins, carotenoids and polyphenolic compounds [14].Extracts of the fruit are used in the prevention of hyperglycemia, hypercholesterolemia, hypertension, and lipoproteins oxidation by reducing the oxidative stress and inflammation on vascular system.The health potentials of dates fruits have been credited to their bioactive compounds, trace elements, vitamin C, tannins, lignins, flavones and other polyphenols especially phenolic acids [15][16][17].However, no study has explored the beneficial outcomes of dates vinegar consumption against type 2 diabetes.
In this study, the objective was to evaluate the effects of dates vinegar on blood biochemical and hematological parameters in type 2 diabetic subjects.

Subjects and study design
Following approval by the Institutional Review Board of Minhag University Lahore (Pakistan), 60type 2 diabetic patients (fasting blood sugar (FBS)>126 mg/dL, 1 antidiabetic drug/daily), aged 30-60 years, 29 females and 26 males weighing between 55 to 75 kg, who visited an Al-Rashid Clinic Lahore in Pakistan and volunteered to participate in this study were recruited.Those with hepatic, renal, asthmatic or cardiovascular diseases, vinegar intolerance or consumed alcohol were excluded.The patients were randomized into a group (30 patients) that consumed20 ml of dates vinegar with their normal diet preferably in the morning (or before bedtime) or the placebo group (30 patients) that received20 ml of honey diluted with water using a randomization scheme with blocks of 5 patients for 10 weeks.Throughout the trial, the participants consumed foods with similar energy levels, carbohydrates, cholesterol, proteins and fats from the beginning to the end.

Statistical analysis
The Statistical Package for the Social Sciences (SPSS) version 21.0.0 (SPSS Inc., IBM) was used to analyze the data using one-way analysis of variance (ANOVA).Variations in experimental and placebo groups were determined by the paired t-test.All statistical outcomes with a p value of less than 0.05 were deliberated as statistically significant.The values were expressed as mean ± SD.

Subject characteristics and diet monitoring
Of the 60 patients recruited for this study, five participants (2 in the date vinegar group and 3 in the placebo group) were excluded from the data analyzed because of protocol variance (n=3) or consent withdrawal (n=2) Figure 1.No serious side effects were observed during the trial.The baseline data for the patients are provided Table 1.

Effect of dates vinegar consumption on various blood biochemical factors
As from the (Table 3), it can be seen that fasting blood sugar (FBS), cholesterol and LDL showed statistically significant reduction (P<0.05) in dates vinegar group while folate and HDL significantly increased., however no significant difference was observed in placebo group.MCH and MCV values decreased, while PLT (P<0.005)enhanced in date vinegar group.Slight variations were also observed in RBC, WBC, hemoglobin, MCHC and HCT (Table 4).

DISCUSSION
This is first evidence based study that showed that continuous intake of dates vinegar reduces the HbA1c and FBS levels in patients of type 2 diabetes but increased folate levels in blood.In agreement with our findings, Mahmoodi et al.
(2011) revealed that white vinegar consumption by type 2 diabetic patients reduced HbA1c and improves various biochemical factors like total cholesterol and LDL as compared to placebo.
Similarly, one researcher showed that regular consumption of vinegar reduced the HbA1c (0.16%) concentration in experimental subjects as compared to control [21].In the small intestine, acetic acid, major component of dates vinegar, stalled the starch molecules digestion completely via blocking the activity of disaccharides and also reduces uptake of glucose through muscle performance.Intake of 0.2 per 100 grams of acetic acid decreased accretion of xylulose-5-phosphate (Xu-5-P) in liver and activity of phosphofructokinase-1 (PFK-1) in skeletal muscles [9].Acetic acid also slows down the rate of gastric emptying [22].Enhancement of free fatty acids in subjects having impaired glucose tolerance, presupposes that insulin resistance connected with enhanced free fatty acids befalls earlier the inception of hyperglycemia [23].Results of one trial conducted in non-diabetic subjects reveled that reduced glucose exploitation in muscles was connected with severe raise of free fatty acids [24].Some experiments also showed association among insulin resistance and levels of free fatty acids [25,26].During the manifestation of insulin resistance, free fatty acids in the form of triglycerides are stored in heart, liver, pancreas and muscle.Particularly, agents that reduced enhanced levels of free fatty acids, like thiazolidinedione's, have been exposed to upregulate insulin sensitivity in liver, adipose tissue and muscle.As, outcomes of our study showed that regular consumption of dates vinegar reduces lipid profile parameters hence, we can say that dates vinegar might be involved in improving insulin sensivity by reducing the free fatty acids levels.
High level of fructose and fiber content in dates might be interconnected with reduced level of glycemic index (GI).The results of one study revealed that regular consumption of rutab dates enhanced the GI and suggested that high content of rutab dates hydration affects gastric emptying, insulin secretion or intestinal absorption [27].Similarly, results of various studies showed that ingestion of dietary fibers applies physiological effects on the small intestine and stomach that curb postprandial glycemic responses, comprising adjourning small bowel passage and modifying myoelectric commotion of gastrointestinal way, suspending gastric emptying lessening the availability of αamylase to its substrate because of enhanced gut contents viscosity, and decreasing diffusion of glucose via callous water layer gel forming and enhanced viscosity chattels of fibers are primarily accountable for its glycemic effect.Because of reformed levels of incretin, soluble fibers prolonged the carbohydrates absorption in intestine along with enhanced levels of glucagonlike peptide 1 (GLP1) [28][29][30].Soluble dietary fibers also affect the uptake mechanisms of glucose via periphery, comprising enhanced skeletal muscle manifestation of the insulinresponsive glucose transporter type 4 (GLUT-4), which increases skeletal muscle upregulation, enhances sensitivity of insulin and regularizes blood glucose [31].Several fatty acids in humans stimulate the manifestation of peroxisome proliferator-activated receptor-γ (PPARG), which enhances levels of adipocyte .
During metabolic disorders the levels of hepatic enzymes like ALT and AST increased and thus resulted in atherosclerosis [33].In our study the levels of ALT, AST and ALP reduced at the end of trial in the experimental group.While, the results of one study showed that the activity of liver AST and ALP in the animals treated with the vinegar decreased and elevated, respectively [34].
Current results showed slight variations in the number of RBC, WBC count and its allied indices like Hb, MCHC, MCV, MCH and platelets.Total RBC and Hb along with platelets increased at the end of study.While, results of one study showed that no substantial variations were observed in these variables after ingestion of vinegar [35].
The increase in total RBC and Hb might be due to high content of folate in dates.
In summary, this study indicates that date vinegar improves the concentration of HbA1c, FBS and lipid profile, and/or potentiates the capacity of anti-glycemic remedies in type 2 diabetic subjects.The limitations of this study include several proposed molecular mechanisms through which date vinegar may exert its effect were not measured directly.More studies are required to substantiate these preliminary outcomes.During the trial, neither deformity of kidney or liver functions nor adversative outcomes were noted.As regular consumption of dates vinegar (20 mL) for 10 weeks in diabetic patients was testified not to cause side effects, dates vinegar consumption is supposed to be safe in type 2 diabetic patients.

CONCLUSION
Conclusively based on the present outcomes, it can be decided that dates vinegar is beneficial as a therapeutic agent in diabetes, without producing side effects.Intake of 20 mL of dates vinegar on daily basis was adequate to attain these effects.Dates vinegar might be considered helpful for precluding metabolic syndrome by reducing diabetes and its related complications.However, more experiments are required to discover both its disadvantages and advantages as a tool for directing type 2 diabetes.
authors listed in this manuscript contributed equally, and read and approved the manuscript.

Table 1 :
Baseline characteristics of subjects who completed the intervention trial Values are expressed as mean ± SD.No significant difference as compared with the initial (p<0.05).BMI; body mass index; BP, blood pressure; HDL-C high-density lipoprotein cholesterol; LDL-C low-density lipoprotein cholesterol VLDL-C very low-density lipoprotein cholesterol.

Table 2 :
Mean daily intake of energy and selected nutrients at baseline and after intervention Intakes of test products were included in the analysis and estimated from three-day food records.Values are expressed as mean±SD.*Significant difference as compared with placebo group.

Table 3 :
Effects of dates vinegar consumption on various biochemical factorsVariables are expressed as mean ± SD.

Table 4 :
Effects of dates vinegar on various hematological factors Bickerton AS, McQuaid SE.The contribution of splanchnic fat to VLDL triglyceride is greater in insulin-resistant than insulin-sensitive men and women: studies in the postprandial state.Diabetes 2007;56: 2433-2441.27.Miller C, Dunn E, Hashim I.The glycaemic index of dates and date/yoghurt mixed meals.Are dates 'the candy that grows on trees'?Eur J Clin Nutr 2003;57: 427.28.Schulze MB, Liu S, Rimm EB, Manson JE, Willett WC, Hu FB.Glycemic index, glycemic load, and dietary fiber intake and incidence of type 2 diabetes in younger and middle-aged women.Am J Clin Nutr 2004;80: 348-356.29.Brownlee IA.The physiological roles of dietary fibre.