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 Table of Contents  
Year : 2021  |  Volume : 12  |  Issue : 4  |  Page : 508-516

Research design for a randomized control trial to assess the effects of almond supplementation on insulin resistance, glycemic markers, and inflammation among overweight Asian Indians

1 Department of Foods Nutrition and Dietetics Research, Madras Diabetes Research Foundation, Chennai, Tamil Nadu, India; Ph. D Scholar, Chennai, Tamil Nadu, India
2 Department of Foods Nutrition and Dietetics Research, Madras Diabetes Research Foundation, Chennai, Tamil Nadu, India
3 Department of Diabetology, Madras Diabetes Research Foundation, Chennai, Tamil Nadu, India
4 Department of Advanced Biochemistry, Madras Diabetes Research Foundation, Chennai, Tamil Nadu, India; Department of Neurochemistry, National Institute of Mental Health and Neurosciences (NIMHANS), Bengaluru, Karnataka, India
5 Department of Home Science, Women’s Christian College, Chennai, Tamil Nadu, India
6 Department of Advanced Biochemistry, Madras Diabetes Research Foundation, Chennai, Tamil Nadu, India
7 Department of Nutrition Science, College of Health and Human Sciences, Purdue University, West Lafayette, Indiana
8 Department of Biochemistry and Biotechnology, IISPV, School of Medicine, Rovira iVirgili University, Reus, Spain
9 Department of Nutrition, Harvard School of Public Health, Boston, USA; Department of Epidemiology, Harvard School of Public Health, Boston, Massachusetts, USA

Date of Submission28-Jun-2021
Date of Decision27-Aug-2021
Date of Acceptance03-Sep-2021
Date of Web Publication12-Jan-2022

Correspondence Address:
Mrs. Vasudevan Sudha
Department of Food, Nutrition and Dietetics Research, Madras Diabetes Research Foundation, ICMR Centre for Advanced Research on Diabetes, 98, Peters Road, Gopalapuram, Chennai 600086, Tamil Nadu.
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jod.jod_85_21

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Background: Fatty acids play an important role in health and well-being; almonds have the highest amount of monounsaturated fatty acids (MUFAs) among the nuts. Western studies have shown positive health effects of almonds. However, well-designed studies are sparse on Asian Indians who have a unique phenotype with higher predisposition to diabetes and cardiovascular disease (CVD). Hence, the present study describes the design and methods of a clinical trial to assess the effect of almond supplementation on insulin resistance, glycemic markers, and inflammation in overweight Asian Indians. Methods and Outcome Assessments: Parallel-arm open-labeled, randomized controlled trial was conducted in Chennai, India. The study included 400 overweight and obese volunteers of age 25–65 years with a body mass index ≥23 kg/m2 and with some having cardiometabolic risks. The participants in the intervention group received 43 g of almonds per day as recommended by the American Heart Association for 12 weeks, whereas the participants in the control arm followed their habitual dietary patterns and were advised not to consume any nuts. All other lifestyle habits were similar. The anthropometric, clinical, biochemical, and diet data of the participants were assessed periodically. Dietary 24-hour recalls and plasma percent fatty acid of the participants were assessed at the baseline and end of the study as a measure of participant compliance to protocol. This study also assessed gut hormone levels as a marker for satiety. The effects of almonds supplementation on anti-inflammatory and inflammatory markers such as adiponectin, monocyte chemoattractant protein-1, and tumor necrosis factor-α were also assessed. Discussion: The study findings, if benefits are found, would help to improve the MUFAs intake by a single supplementation of almonds daily to meet the dietary guidelines of 15% of total calories of MUFAs. In addition, it might aid in the prevention of obesity-related chronic diseases such as diabetes and CVDs by reducing the cardiometabolic risk factors. Trial Registration: The trial was registered in the clinical trial registry of India CTRI201710010251.

Keywords: Almonds, insulin resistance, insulin sensitivity, MUFA, obesity

How to cite this article:
Gayathri R, Kalpana N, Manasa VS, Sudha V, Shobana S, Jeevan RG, Anjana RM, Unnikrishnan R, Gokulakrishnan K, Krishnaswamy K, Beatrice D A, Pradeepa R, Mattes R, Salas-Salvadó J, Willett W, Mohan V. Research design for a randomized control trial to assess the effects of almond supplementation on insulin resistance, glycemic markers, and inflammation among overweight Asian Indians. J Diabetol 2021;12:508-16

How to cite this URL:
Gayathri R, Kalpana N, Manasa VS, Sudha V, Shobana S, Jeevan RG, Anjana RM, Unnikrishnan R, Gokulakrishnan K, Krishnaswamy K, Beatrice D A, Pradeepa R, Mattes R, Salas-Salvadó J, Willett W, Mohan V. Research design for a randomized control trial to assess the effects of almond supplementation on insulin resistance, glycemic markers, and inflammation among overweight Asian Indians. J Diabetol [serial online] 2021 [cited 2023 Feb 4];12:508-16. Available from: https://www.journalofdiabetology.org/text.asp?2021/12/4/508/335601

  Background Top

Consumption of nuts, a rich source of monounsaturated fatty acids (MUFAs), has historically been low in India (8 g/day in 1975–1979 and 17 g/day in 1996–1997).[1] Various clinical trials on nut consumption have shown health benefits in improving the overall metabolic outcome. Mediterranean diets enriched with mixed nuts (30 g/day) have shown to be helpful in the management of metabolic syndrome in the Prevención con Dieta Mediterránea study.[2] Our recent randomized controlled trial (RCT) in Asian Indians with type 2 diabetes (T2DM) on cashew nut supplementation has shown improved systolic blood pressure and high-density lipoprotein cholesterol (HDL-c) levels with no significant deleterious effects on body weight, glycemic, or other lipid parameters.[3]

The prevalence of diabetes and prediabetes continues to be on the rise. Currently, 463 million people in the world have diabetes and of this 77 million people with diabetes reside in India[4] and 80 million people in India have prediabetes.[5] Asian Indian phenotype and thrifty genotype combined with altered lifestyles (poor dietary practices and physical inactivity) are the key drivers for the escalating prevalence of obesity further fuelling the diabetes epidemic in India.[6],[7] Suboptimal diet is attributed to more deaths globally even when compared to smoking. Good quality of a diet can prevent one in every five deaths.[8]

Diets high in refined grains showed a positive association with insulin resistance, low HDL levels, metabolic syndrome, and T2DM.[9],[10],[11] Replacement of refined grains with healthy fats such as MUFAs may have beneficial effects on HDL-c.[3],[12] MUFA-rich diet prevented central fat redistribution and the postprandial decrease in peripheral adiponectin gene expression and insulin resistance induced by a carbohydrate-rich diet.[13] Studies have also reported the beneficial role of MUFA intake in the prevention of T2DM.[14] The inclusion of natural whole nuts like almonds offers promise in terms of delivering MUFA in the healthiest form in addition to MUFA-rich cooking oils or as a standalone dietary supplement.

Almonds are good sources of protein (17%), dietary fiber (13%) and fat (58%), and magnesium (318 mg/100g), and remain to be a very rich source of MUFA (38%).[15],[16] Almonds have been well studied in the western population and are known to have positive health effects on satiety (appropriate changes in gut hormones), insulin sensitivity, improved glycemic, lipids, and inflammatory markers. Poor dietary practices of Asian Indians are associated with cardiometabolic risks and include a high intake of refined carbohydrates such as white rice, low intake of whole grains, fruits, vegetables, MUFA, and micronutrients.[17],[18] Therefore, MUFA-rich food supplements like almonds could offer a strategy to improve MUFA intake in Asian Indians. Very few studies in India have shown the beneficial effects of almonds on cardiometabolic risk factors, dyslipidemia, and glucose responses in T2DM. Almonds have also known to promote the growth of healthy microbiota.[19],[20],[21] However, the role of almonds on insulin resistance, inflammation, and satiety in Asian Indians especially in overweight and obese individuals, who are at risk for T2DM and cardiovascular disease (CVD), remains unexplored. Studies on the 24-hour glycemic response to nuts (a rich source of MUFA) supplementation adjusted to maintain total calorie intake may provide unique insights about postprandial glycemic excursions especially in “at risk population.” Therefore, there is an unmet need to evolve dietary strategies from well-conducted randomized clinical trials to mitigate the health and economic burden of diabetes. Thus, this article explains the study design and methods of a randomized trial to evaluate the effect of 43 g almond supplementation as per the guidelines of the American Heart Association (AHA, 2010) for 12 weeks among overweight and obese adults with the following objectives:

  1. Primary: To determine the effects of almond supplementation on Homeostasis Model Assessment-insulin resistance (HOMA-IR) and beta-cell function assessed by oral disposition index (DIo), C-peptide levels, and adiposity.

  2. Secondary: To study the effect on the biomarkers of glucose metabolism (i.e., fasting glucose and hemoglobin A1c), lipid profile (i.e., triglycerides, total cholesterol, low-density lipoprotein cholesterol (LDL-c), and HDL-cholesterol), plasma fatty acid profile, satiety-related gut hormones using a multiplex panel (glucagon-like peptide-1 [GLP-1], gastric inhibitory polypeptide [GIP], peptide YY [PYY], and ghrelin), adiponectin, and selected inflammatory markers (i.e., hs-C reactive protein [hs-CRP], tumor necrosis factor-α [TNF-α], monocyte chemoattractant protein-1 [MCP-1]).

  Materials and Methods Top

Study design and ethics approval

The study was a randomized, parallel-arm 12 weeks trial. The participants recruited for the study were randomized either into the control group or the almond (43 g/day) intervention group. The study was approved by the Institutional Ethics Committee at Madras Diabetes Research Foundation (MDRF) and registered in the clinical trial registry of India, reference number CTRI201710010251. The study was conducted in accordance with guidelines in the Declaration of Helsinki.

Study population

Sample size

The sample size was calculated with the primary outcome measure of HOMA-IR with the mean difference of 0.72 considered from the reported studies in literature since data on almond supplementation in Asian Indians were unavailable. The study was planned with 80% power and an alpha of 0.05 to detect a difference of 20% change in HOMA-IR with 200 participants per group and a dropout rate of 20%.

Participant recruitment and consent

The present study was conducted among 400 overweight/obese Asian Indian adults selected from the volunteer registry maintained at our research center, the MDRF. Potential participants were invited for screening at our center and interviewed by trained study staff who explained the background, objectives, and procedures of the trial, sought and obtained written informed consent, confirmed eligibility, and completed the baseline assessments. Participants then completed a baseline glucose tolerance test to rule out any new diabetes. Eligible participants meeting the inclusion and exclusion criteria were recruited for the study.

Inclusion and exclusion criteria employed to screen participants were as follows:

Inclusion criteria

  • 25–60 years (men and women)

  • Body mass index (BMI) ≥23 kg/m2

  • Central obesity (waist circumference ≥90 cm for men and ≥80 cm for women)

  • Dyslipidemia – low HDL (≤40 mg/dl men and 50 mg/dl women) and (or) high triglyceride (≥150 mg/dl)

  • Positive family history of diabetes

  • Hypertension (≥140/90 mmHg)

  • Snacking (mid-morning)

  • Exclusion criteria

  • Self-reported diabetes

  • History of allergy to nuts

  • Use of nutritional supplements

  • Bad dentures

  • Weight loss diets

  • Pregnant/lactating women

  • Liver, kidney, thyroid, or other endocrine diseases

  • Any condition that hinders compliance to the intervention

  • Possible eating disorder

  • Randomization

    After screening, randomization was done centrally by an independent biostatistician using a computer-generated random number sequence. Participants were randomized with stratification by day-to-day screening based on a 1:1 allocation ratio to either the intervention or to the control group.

    Intervention and control group

    Participants assigned to the intervention group were given sachets containing ≈43 g of raw almonds daily for 12 weeks. A 1-week run-in period was conducted before intervention to ensure participants’ acquiescence to the study protocol. The participants in the intervention group were advised to adjust the almond calories for either carbohydrate or fat calories in their routine standard Indian diets by trained research dietitians. Further, the participants were instructed not to share the almonds with family members, and to ensure this, per month 200 g of almonds were provided for their family members. To preserve the quality and freshness of almonds, sufficient sachets of almonds were provided to the participants monthly once. [Table 1] depicts the nutrient composition of almonds. Almonds are rich in MUFAs (38 g/100 g), dietary fiber (13 g/100 g), and calcium (228 mg/100 g), magnesium (318 mg/100g), vitamin E (13 g/100 g) and have minimal carbohydrate (3 g/100 g) content. This is especially important in the Indian context wherein the diets are high in carbohydrates and deficient in MUFA and fiber. The present study was carried out using the nonpareil almond variety which is the most common and popular form of almonds known for its versatility and texture. The nutrient composition of nonpareil versus other common almond varieties is given in [Table 2]. The nutrient content of nonpareil almonds was more or less similar to the other almond varieties.
    Table 1: Selected nutrition profile of almond

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    Table 2: Nutrient profile of nonpareil almond vs other almond varieties

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    Participants in the control group continued with their usual diet and were further instructed to refrain from intake of nuts in any form during the 12-week study period. All the participants recruited for the study were advised to refrain from fasting and feasting and also requested not to make any major lifestyle modifications during the study period.

    Data collection and follow-up

    Anthropometric and clinical assessments

    Bodyweight (kg) (electronic OMRON machine; Omron HBF 212, Tokyo, Japan), and waist circumference (cm) were measured using standard protocols. BMI was calculated as weight (kg) divided by height squared (m2). Blood pressure was measured twice on each occasion with an interval of 5 minutes using an electronic OMRON machine (Omron HEM 7120, Tokyo, Japan). Anthropometric and clinical assessments were done at baseline and followed up monthly once during the study period.

    Biochemical assessments

    Glucose tolerance test (0, 30, and 120), HbA1c, fasting insulin, hs-CRP, and lipid profile were assessed on the first day of biochemical assessments. In addition, after 2 days interval another blood sample was collected at fasting for assessment of inflammatory markers such as TNF-α, MCP-1, adiponectin, gut hormones such as GLP-1, GIP, PYY, and ghrelin and post-standardized breakfast blood samples were collected for assessment of GLP-1 (four pointers: 0, 30, 60, and 120 min), and postprandial C-peptide. The plasma fatty acid profiles of the participants were also measured as a marker of compliance at baseline and post-intervention with almonds for 12 weeks. All the specified biochemical assessments were done at baseline and end of 12 weeks study period. In addition, fasting blood glucose and fasting insulin were assessed at midpoint of the study (45 days). The blood samples were collected by a trained phlebotomist.

    Plasma glucose was estimated using the glucose oxidase–peroxidase method (Roche Diagnostics, Basel, Switzerland). Glycemic and lipid profiles were assessed using a Hitachi 912 Autoanalyzer (Hitachi, Mannheim, Germany). HbA1c was estimated by high-pressure liquid chromatography using the Variant machine (Bio-Rad, Hercules, and CA), and serum insulin by electrochemiluminescence assay (Roche Diagnostics, Basel, Switzerland). The β cell function has been evaluated using DIo, which is a measure of ratio of the difference in fasting to 30-min insulin (I) to glucose (G) values = (δI0–30/δG0–30), and insulin resistance calculated by the HOMA-IR using the formula: fasting insulin (uIU/ml) × fasting glucose (mmol/l)/22.5.

    Serum cholesterol (CHOD-PAP method), serum triglycerides (GPO-PAP method), and HDL cholesterol (Direct method–polyethylene glycol-pre-treated enzymes) have been measured, and LDL cholesterol calculated using the Friedwald formula. The coefficients of variation for the biochemical assays ranged from 3.1% to 7.6%. Gut hormones and inflammatory markers assessments were done using Enzyme-Linked Immunosorbent Assays. Plasma fatty acid was assessed using gas chromatography, lipids were extracted in chloroform–methanol mixture and converted to fatty acid methyl esters for quantification of plasma fatty acids using gas chromatography. Appropriate fatty acid methyl ester standards were used for the identification and quantification of peaks.

    Diet and lifestyle

    Data collection was done by trained research staff and baseline demographics, medical history, and lifestyle factors of the participants were assessed using a screening questionnaire. Participants’ physical activity (PA) level was assessed at baseline and end of every month using a validated PA questionnaire. Dietary 24-hour recalls were administered during screening, run-in period, at baseline blood collection, and monthly twice (1 weekday and 1 weekend) during the study. Further, adverse event and satiety score questionnaires were assessed at baseline and at the end of every month.

    Participants’ compliance to the study protocol was assessed at three levels: (1) return of empty sachet of almonds collected at end of every month, (2) change in the nutrient profile of the participants from 24-hour dietary recall data, and (3) change in plasma fatty acid levels

    The study parameters and timeline of the assessments are shown in [Table 3] and [Figure 1].
    Table 3: Study protocol assessments and schedule

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    Figure 1: Study design.

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    Statistical analysis

    Statistical analysis was performed using SAS software, version 9.0 (SAS Institute Inc., Cary, NC). Between-group differences for change in health measurements and dietary intake between baseline and 12 weeks will be analyzed using generalized linear models. The outcome data will be controlled for baseline outcome as a sensitivity analysis. The baseline demographic, biochemical, clinical, and dietary data of participants recruited for the study are reported as mean ± SD.

      Results Top

    The proposed trial was designed as an intervention trial with almonds that are of late commonly consumed in different parts of India.

    Participants (n = 635) willing to participate in the study were screened and underwent an oral glucose tolerance test. Participants with newly diagnosed diabetes, not willing to participate, refused to give a blood sample and wear flash glucose monitors, etc (n = 235) were excluded. Participants (n = 400) were then assigned to intervention group (n = 200) and control group (n = 200) using computer-generated randomized numbers.

    The baseline characteristics of the participants recruited for the study are given in [Table 4]. The mean age of the participants recruited for the study was 37± 9 years. About 68% of participants in the control group and 57% in the intervention group were women. The systolic blood pressure, 30 min and 120 min insulin assays were observed to be higher in the intervention group compared with the control group.
    Table 4: Demographic, anthropometric, and clinical characteristics of the study participants at baseline (n = 400)

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    The baseline dietary characteristics of the study participants based on an average of three dietary recalls (24 hours) collected are shown in [Table 5]. The total energy, carbohydrate, protein, fat, and dietary fiber in the intervention group were higher, whereas the calorie from carbohydrate was lower compared with control group.
    Table 5: Dietary characteristics of the study participants at baseline (n = 400)

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      Discussion Top

    This study successfully recruited 400 participants suggesting that the approach developed can efficiently and rapidly recruit participants at risk for developing cardiometabolic disorders from the urban population in India to evaluate the effect of almond supplementation on insulin resistance, glycemic markers, and inflammation among overweight Asian Indian adults. The evaluation of the acceptability and adherence to long-term almond supplementation depended on the complete, unbiased follow-up, and outcome assessment of participants during the study period.

    This study was designed to encourage future RCT that can be conducted for longer terms and in greater strengths of cohorts that will test the effect of almond supplementation in a more diverse population with respect to socio-demographical differences. Evidence-based medicine seeks the best use of available evidence in making sound clinical decisions while caring for patients. Only RCTs offer the kind of confirmation for dietary suggestion better than observational study designs.[22] RCTs have proven that medical nutrition therapy is beneficial and cost-effective in improving the metabolic outcomes of patients.[23] There is a growing recognition that nutrition science will benefit from more adequately powered and well-conducted RCTs to evaluate the health effects of nutrients and foods on clinical outcomes, which can complement findings from prospective cohorts and metabolic studies. These research approaches have different strengths and weaknesses, and the consistency of findings can significantly enhance the development of dietary guidelines and their effective implementation.[24] In a country like India where CVD and diabetes are known to increase disability-adjusted life years,[25] it is important to establish nutrition interventions, which will aid in the prevention of the disease especially through its effects on the predominant modifiable risk factor dyslipidemia and nuts consumption falls in line with this need of the hour.[26]

    Dietary guidelines have been developed for Asian Indians for the prevention of obesity and dyslipidemia and include reduction in the intake of carbohydrates, preferential intake of complex carbohydrates and low glycemic index foods, higher intake of fiber, lower intake of saturated fats, the optimal ratio of essential fatty acids, and reduction of trans fatty acids.[27],[28] Besides this, the Chennai Urban Rural Epidemiological Studies identified the combination of obesity, physical inactivity, unfavorable diet risk score, hypertriglyceridemia, and low-HDL cholesterol could explain 80.7% of all incident diabetes and concluded that modifying these easily identifiable risk factors could therefore prevent the majority of cases of non-communicable diseases beginning with diabetes.[9] Therefore, almonds which are a rich source of minerals like calcium, copper, iron, magnesium, phosphorus, potassium, zinc, manganese, and vitamins such as thiamine, riboflavin, niacin, and vitamin E with their healthy fatty acid composition are greatly beneficial in mitigating the risks of CVDs by reducing the levels of LDL-C and triglycerides while improving HDL levels.[26] Besides these, omega-3 fatty acids, especially alpha-linolenic acid, which are present in almonds reduce CVD risk, by changing vascular inflammation and improving endothelial dysfunction[29] and the authors also pointed out that sufficient studies to prove the efficacy of nuts especially almonds and walnuts were sparse. Therefore, design and execution of this study seem to contribute science-based pieces of evidence for the prevention of non-communicable disease through simple lifestyle changes like consuming almonds as a potent dietary supplement.

    There are several notable strengths of the study. First, the study 2-arm RCT allowed testing for a potential effect of almond consumption. Second, the sample size of the study provides a piece of stronger evidence compared with previously conducted RCT on almond supplementation. Third, using inflammatory markers such as hs-CRP, TNF-α, MCP-1; adiponectin, gut hormones- GLP-1, GIP, PYY, and ghrelin and plasma fatty acids to assess compliance minimized the risk of measurement bias and errors associated with self-reported dietary intake.

    The limitations of the study include

    1. It was not possible to blind the participants to their randomized group.

    2. Long-term effects may not be evaluated as this study is up to 12 weeks of supplementation in a community trial.

    3. The baseline plasma fatty acid and gut hormones are yet to be analyzed

    4. Though we advised the participants to adhere to the study protocol and not to make major lifestyle and dietary changes, in a community-based trial such as this, family and environmental influences (like festivals, marriage feasts, and parties) may have had some impact on the outcomes.

    5. These effects would be likely to occur in both groups, biasing results toward the null.

      Conclusion Top

    The study results if found to be positive data could help to devise dietary strategies which can potentially reduce insulin resistance even among overweight and obese adults and thereby help prevent diabetes. Public health message to improve the MUFA intake by a single supplementation of almonds daily which would help Asian Indians not only to meet the dietary guidelines of 15% of total calories of MUFA, but also aids in prevention of obesity-related chronic diseases such as diabetes and CVDs.


    The authors thank Almond Board of California for sponsoring the study.

    Authors contribution

    GR initiated the article, KN supported in data collection and editing the article. VSM assisted in writing article. VS, SS, RGJ, RMA, RU, GK, KK, AB, GP, LMJ, JSS, WW, and VM reviewed and edited the article.

    Financial support and sponsorship


    Conflicts of interest

    There are no conflicts of interest.

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      [Figure 1]

      [Table 1], [Table 2], [Table 3], [Table 4], [Table 5]


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