Does Supplementation with Whey Protein Improve Post-prandial Glycemic Levels in Healthy Individuals and Type II Diabetics?

Transcription

1 Does Supplementation with Whey Protein Improve Post-prandial Glycemic Levels in Healthy Individuals and Type II Diabetics? Lauren Collier 2015

2 Table of Contents I. Introduction...2 Prevalence and Incidence of Diabetes...2 Etiology of Type II Diabetes...2 Medical Nutrition Therapy II. Search Strategy...4 III. Summary...5 IV. Conclusion...8 V. References...9 VI. Appendix...10 Search Plan and Results...10 Overview Table...14 Quality Criteria Checklists

3 I. Introduction Prevalence and Incidence of Diabetes According to the National Diabetes Association, 29.1 million Americans, or 9.3% of the population, had diabetes in Of those, 8.1 million individuals were undiagnosed. In addition, approximately 86 million Americans are living with pre-diabetes. Not surprisingly diabetes is a burden on our nation s health and financial wellness. The cost of diabetes is staggering, at $245 billion for diagnosed diabetes in the United States in One study examining the prevalence and incidence of diagnosed diabetes in adults aged 20 to 79 years of age in the United States from found that prevalence and incidence of diabetes doubled from Over recent years, data suggests a possible plateau in the number of newly diagnosed patients overall. Subgroups that continue to see an increase in cases include Hispanic and non-hispanic black adults and individuals with a high school education or less. The cause of this potential slowing of diabetes incidence is unknown but several possibilities have been identified including a plateau in obesity incidence in the US and a shift to Hemoglobin A1C as the gold standard for diagnosing diabetes. 2 Co-morbidities resulting from uncontrolled diabetes include renal disease, amputation, neuropathy, and blindness putting further strain on our health care system. The sheer number of individuals living with diabetes makes it a leading public health concern that demands solutions in medical management and prevention. Etiology of Type II Diabetes The etiologies of diabetes are not fully understood and ongoing research is being conducted to determine all causes of this condition. Researchers have concluded that lifestyle 2

4 factors and genetics contribute heavily to the onset of diabetes. In general, we understand that the pancreas increases insulin production in reaction to elevated serum glucose. Over time, insulin production decreases until an individual can no longer produce enough to sustain normal glucose levels. Some people experience insulin resistance, or the inability to utilize insulin properly. In insulin resistance, cells do not respond properly and cannot absorb serum glucose. This causes elevated glucose levels and increased insulin release. Insulin resistance can lead to a number of conditions including diabetes and obesity. Individuals with elevated glucose above normal range but below diagnostic criteria for diabetes are classified as pre-diabetics. Type II diabetes can be prevented or delayed in individuals with pre-diabetes thus reinforcing the importance of nutrition counseling and lifestyle change. Medical Nutrition Therapy The Nutrition Care Manual, available through the Academy of Nutrition and Dietetics, outlines current standards of care in both pre-diabetic and diabetic patients. Modest weight loss, a reduction in energy intake, and increased physical activity can significantly improve patient outcome in both conditions. 3 According to the position statement by the American Diabetic Association, individuals with diabetes should receive individualized medical nutrition therapy by a registered dietitian/nutritionist. The efficacy of medical nutrition therapy in the control of type II diabetes has been found to be significant through randomized control trials and Cochrane reviews. Many dietary patterns can be effective in the management of diabetes. Currently, there are no standards for the ideal combination of macronutrient consumption in diabetics. More specifically, evidence concerning the amount of protein to consume for best glycemic control is inconclusive. 4 3

5 Protein is a critical macronutrient that contributes to the body s function by acting as catalysts, messengers, structural elements, buffers, and immunoprotectors. During digestion, protein produces greater long-term satiety than carbohydrate or fat which may decrease appetite and overall food consumption. 5 In one study whey protein was found to have a greater satiating effect than casein. In addition, protein consumption appears to stimulate insulin secretion from the pancreas without raising serum glucose in type II diabetics. With this understanding, what does the research conclude concerning whey protein and its ability to assist individuals with glycemic control? As mentioned above, the American Diabetic Association concludes that the evidence concerning the amount of protein to consume for best glycemic control is inconclusive; however, the Academy of Nutrition and Dietetics has yet to make any formal statement in regards to this question. Does supplementation with whey protein improve post-prandial glycemic levels in healthy individuals and type II diabetics? II. Search Strategy The inclusion and exclusion criteria were developed before the acquisition of articles began. Inclusion criteria are as follows: participants must be 18 years or older, the study design should be a clinically controlled study, sample size > 8, subjects must be either healthy or with type II diabetes, the study must have been conducted in 2004 or sooner, articles must be published in English, and the only supplement being tested will be whey protein. Studies that included individuals less than 18 years of age, less than 8 participants, participants with any chronic condition beside type II diabetes, studies conducted prior to 2004, articles not published in English, and interventions containing whey protein in combination with an additional 4

6 supplement were excluded from the literature review. The PubMed database was used as the primary search engine. The search terms diabetes, whey protein, and glycemic control were used in several combinations to find applicable articles. Random control trials, clinical studies, observational studies, cohort and case-control studies were acceptable to include in this review. The search terms diabetes and whey protein generated 64 possible articles; 56 of which were excluded by title. The search terms whey protein and glycemic control gave 14 possible articles; 13 were excluded by title. One article was hand pulled from the references section of a selected study. Nine articles were identified for review from the online electronic database and one was hand selected. Four articles were excluded upon further investigation. For a list of excluded articles and the reason for exclusion please see appendix. For a justification of chosen articles please see appendix. III. Summary For the purpose of this project six clinical studies were reviewed and analyzed. Several trends and comparisons were found among the literature. Frid et. al. conducted a non-randomized control study in which 14 participants with diet controlled type II diabetes were asked to consume a breakfast and lunch meal on two separate occasions less than 1 week apart. One test day was designated as the treatment day with subjects consuming whey protein in place of ham and lactose in addition to breakfast and lunch. Blood samples were collected and researchers discovered a significantly lower blood glucose and higher insulin concentration after consumption of the lunch with whey protein as compared to the reference lunch containing ham and lactose. 6 5

7 In another study conducted by Ma et. al., diet controlled type II diabetics, 8 participants were asked to fast overnight on three separate occasions. On each test day subjects consumed a beef flavored soup 30 minutes before eating a mashed potato meal. On day one 55 g of whey protein was added to the beef soup, on day two 55 g of whey protein was added to the mashed potatoes, and on day three no whey protein was added. Researchers found that gastric emptying was significantly slowest when whey protein was added as a preload in the beef soup. There was a significant difference in increased insulin and decreased blood glucose in individuals when they consumed 55 g whey protein as both the preload and in the meal compared to no added whey protein. 7 The iauc for blood glucose was less after the whey preload ( / mmol min ) and whey in meal (406.3+/-85.9 mmol min ) compared with no whey ( / mmol min ) P+/ for both). Type II diabetics with well controlled diabetes were studied by Jakubowicz et. al. to examine if Glucagon-like Peptide-1(GLP-1) secretions stimulated by a whey protein preload would benefit from the glucose-lowering effects of these secretions. Subjects consumed 50 g of whey dissolved in 250 ml of water or a placebo followed by a high glycemic index breakfast on two separate occasions. Plasma glucose was found to be significantly lower in the whey preload group while insulin concentrations were increased 105%. Intact and total GLP were elevated postprandially in the whey preload group compared to the control group. 8 In addition to research conducted on type II diabetics, the glucose lowering effects of whey protein has been studied by Petersen et. al. using healthy individuals. In one study escalating doses of glycemic index lowering peptide (GILP) from whey protein was given in addition to 50 g of glucose. Post prandial serum glucose was found to decrease as GILP doses increased. 9 6

8 iaucs for the GILP plus glucose meals of 0, 5, 10 and 20 g of protein were 231 ± 23, 212 ± 23, 196 ± 23, and 138 ± 13 mmol/l min. In a positively rated, randomized crossover trial, Akhaven et. al. recruited healthy young adults to participate in a study which analyzed the effect of the pre-meal consumption of whey protein or its hydrolysate on food intake, pre and post-meal satiety, and blood glucose. Researchers found that whey protein suppressed food intake compared to the control. Whey protein, but not its hydrolysate, reduced post-meal blood glucose. 10 Finally, lean and healthy individuals were recruited by Acheson et. al. to participate in a study to analyze the effects of three proteins on energy metabolism, satiety, and glucose control. Twenty-three participants visited the clinic on six separate occasions after an overnight fast. Participants consumed one of four iso-caloric test meals, three of which consisted of 50% protein (whey, casein, or soy), 40% carbohydrate, and 10% fat. The fourth meal consisted of 95.5% carbohydrate. Researchers found that the thermic effect was greatest after the whey meal when compared to the other test meals. Glycemic response was attenuated 32% by the proteins and a greater insulin response was found after whey than casein and soy. 11 The results of each of these studies reveal a significant decrease in postprandial blood glucose and an increase in insulin. Such results suggest that both a whey protein preload or incorporation of whey to a meal may stimulate insulin secretion and help diet controlled type II diabetics and healthy individuals lower glycemic levels. The studies reviewed did not include participants with uncontrolled type II diabetes, insulin-dependent diabetics, or individuals with multiple chronic diseases. The studies outlined above generally controlled for age, sex, comorbidities, and Body Mass Index (BMI). All were clinical studies and were either non- 7

9 randomized crossover trials, randomized crossover trials, or randomized control trials. Overall, sample sizes were small with some study designs being weaker than others. IV. Conclusion After careful analysis of the six research studies regarding whey protein and glycemic control, the grade assigned to the evidence is Grade II: Fair. All studies reviewed contained small sample sizes. In addition, recruitment for these studies were generally advertised and on a first come first serve basis. Some studies offered financial compensation while others may have some bias due to funding and employment sources. The researchers, however, are all in agreement with their results. Incorporating whey protein into a meal or consuming a pre-meal snack with whey protein may assist diet controlled type II diabetics in maintaining appropriate blood glucose ranges. In addition, with the staggering number of pre-diabetics in the United States, this recommendation may assist pre-diabetics in preventing the onset of type II diabetes through the insulin stimulation and satiating effects of whey. Additional research is needed to determine the amount of whey protein needed for optimal glycemic control. Diabetes is common among veterans served at the Memphis VA Medical Center. With our understanding of the costs and comorbidities associated with diabetes, it is prudent to explore potential medical nutrition therapies that may assist our veterans in controlling glycemic levels. Adding whey protein to a meal or consuming a pre-meal snack with whey protein may assist our diabetic and pre-diabetic patients in controlling blood glucose. Clinical judgment should be used when considering this recommendation for our patients. Comorbidities, compliance, and stage of change should be carefully deliberated. 8

10 V. References 1) American Diabetes Association. (2010). American Diabetes Association. Retrieved from 2) Geiss LS, Wang J, Cheng YJ, et al. Prevalence and Incidence Trends for Diagnosed Diabetes Among Adults Aged 20 to 79 Years, United States, JAMA. 2014;312(12): doi: /jama ) American Dietetic Association. Nutrition Care Manual. Accessed [January 25, 2015]. 4) Evert AB, Boucher JL, Cypress M, Dunbar SA, Franz MJ, Mayer-Davis EJ, Neumiller JJ, Nwankwo R, Verdi CL, Urbanski P, Yancy Jr WS. Nutrition therapy recommendations for the management of adults with diabetes. Diabetes Care. 2013;36: ) Padden-Jones D, Westman E, Mattes RD, et. al. Protein, weight management, and satiety. Am J Clin Nutr. 2008;87(5): 1558S-1561S. 6) Frid A, Nilsson M, Holst JJ, Bjorck I. Effect of whey on blood glucose and insulin responses to composite breakfast and lunch meals in type 2 diabetic subjects. Am J Clin Nutr. 2005;82: ) Ma J, Stevens JE, Cukier K, et. al. Effects of a protein preload on gastric emptying, glycemia, and gut hormones after a carbohydrate meal in diet-controlled type 2 diabetes. Diabetes Care. 2009;32(9): ) Jakubowicz D, Froy O, Ahren B, et. Al. Incretin, insulinotropic and glucose-lowering effects of whey protein pre-load in type 2 diabetes: a randomized clinical trial. Diabetologia. 2014;57(9): DOI: /s x 9) Petersen BL, Ward LS, Bastian ED, Jenkins AL, Campbell J, Vuksan V. A whey protein supplement decreases post-prandial glycemia. Nutr J. 2009;8:47. DOI: / ) Akhavan T, Luhovyy BL, Brown PH, Cho CE, Anderson GH. Effect of premeal consumption of whey protein and its hydrolysate on food intake and postmeal glycemia and insulin responses in young adults. Am J Clin Nutr.2010;91(4): DOI: /ajcn ) Acheson KJ, Blondel-Lubrano A, Oguey-Araymon S, et. al. Protein choices targeting thermogenesis and metabolism. Am J Clin Nutr. 2011;93:

11 VI. Appendix Search Plan and Results Question: Does the supplementation of whey protein in the diet affect post-prandial glycemic levels? Date of Literature Review for the Evidence Analysis: November 10, 2014 Inclusion Criteria: Age: 18 years or older Study Design Preference: Clinical controlled studies Study Group Size: 8 or more individuals Nutrition related condition: Human subjects, healthy or with type 2 diabetes mellitus Year Range: 2004-current Language: Articles published in English Supplement/preload: Whey protein Exclusion Criteria: Age: <18 years Study Group Size: <8 individuals Nutrition related condition: Type 1 Diabetes, any additional disease state or condition Year Range: Prior to 2004 Language: Articles not published in English Supplement/preload: Whey protein combination supplements Search Terms: Diabetes, Whey Protein, Glycemic Control Health Condition: Type 2 Diabetes and healthy individuals Intervention: Whey Protein supplementation or preload Type of Study Design: RCTs, Clinical Studies, Observational Studies, Cohort and Case-Control Studies Electronic Databases: Pubmed Database: Pubmed Search Terms: Diabetes Whey Protein Hits: 64 Articles to Review: 8 Excluded by title Jakubowicz D, Froy O, Ahren B, et. Al. Incretin, insulinotropic and glucose-lowering effects of whey protein pre-load in type 2 diabetes: a randomized clinical trial. Diabetologia. 2014;57(9): DOI: /s x 10

12 Clifton PM, Galbraith C, Coles L. Effect of a low dose whey/guar preload on glycemic control in people with type 2 diabetes-a randomized control trial. Nutr J. 2014;13(1):103. DOI: / Ang M, Muller AS, Wagenlehner F, Pilatz A, Linn T. Combining protein and carbohydrate increases postprandial insulin levels but does not improve glucose response inpatients with type 2 diabetes. Metabolism. 2012;61(12): DOI: /j.metabol Mortensen LS, Homer-Jensen J, Hartvigsen ML, et. Al. Effects of different fractions of whey protein on postprandial lipid and hormone responses in type 2 diabetes. Eur J Clin Nutr. 2012;66(7): DOI: /ejcn Esteves de Oliveira FC, Pinheiro Volp AC, Alfenas RC. Impact of different protein sources in the glycemic and insulinemic responses. Nutr Hosp. 2011;26(4): DOI: /S Petersen BL, Ward LS, Bastian ED, Jenkins AL, Campbell J, Vuksan V. A whey protein supplement decreases post-prandial glycemia. Nutr J. 2009;8:47. DOI: / Ma J, Stevens JE, Cukier K, et. al. Effects of a protein preload on gastric emptying, glycemia, and gut hormones after a carbohydrate meal in diet-controlled type 2 diabetes. Diabetes Care. 2009;32(9): Frid A, Nilsson M, Holst JJ, Bjorck I. Effect of whey on blood glucose and insulin responses to composite breakfast and lunch meals in type 2 diabetic subjects. Am J Clin Nutr. 2005;82: Database: Pubmed Search Terms: Whey Protein, Glycemic Control Hits: 14 Articles to Review: 1 Excluded by title Acheson KJ, Blondel-Lubrano A, Oguey-Araymon S, et. al. Protein choices targeting thermogenesis and metabolism. Am J Clin Nutr. 2011;93: Hand Pull: Esteves de Oliveira FC, Pinheiro Volp AC, Alfenas RC. Impact of different protein sources in the glycemic and insulinemic responses. Nutr Hosp. 2011;26(4): DOI: /S Articles to Review: 1 Akhavan T, Luhovyy BL, Brown PH, Cho CE, Anderson GH. Effect of premeal consumption of whey protein and its hydrolysate on food intake and postmeal glycemia and insulin responses in young adults. Am J Clin Nutr.2010;91(4): DOI: /ajcn Total articles identified to review from electronic databases: 9 11

13 Inclusion List: 6 List of Articles Included from Electronic Databases: 5 Article Frid A, Nilsson M, Holst JJ, Bjorck I. Effect of whey on blood glucose and insulin responses to composite breakfast and lunch meals in type 2 diabetic subjects. Am J Clin Nutr. 2005;82: Ma J, Stevens JE, Cukier K, et. al. Effects of a protein preload on gastric emptying, glycemia, and gut hormones after a carbohydrate meal in diet-controlled type 2 diabetes. Diabetes Care. 2009;32(9): Jakubowicz D, Froy O, Ahren B, et. Al. Incretin, insulinotropic and glucose-lowering effects of whey protein pre-load in type 2 diabetes: a randomized clinical trial. Diabetologia. 2014;57(9): DOI: /s x Petersen BL, Ward LS, Bastian ED, Jenkins AL, Campbell J, Vuksan V. A whey protein supplement decreases post-prandial glycemia. Nutr J. 2009;8:47. DOI: / Akhavan T, Luhovyy BL, Brown PH, Cho CE, Anderson GH. Effect of premeal consumption of whey protein and its hydrolysate on food intake and postmeal glycemia and insulin responses in young adults. Am J Clin Nutr.2010;91(4): DOI: /ajcn Acheson KJ, Blondel-Lubrano A, Oguey-Araymon S, et. al. Protein choices targeting thermogenesis and metabolism. Am J Clin Nutr. 2011;93: Reason Met all inclusion criteria Met all inclusion criteria Met all inclusion criteria Met all inclusion criteria Met all inclusion criteria Met all inclusion criteria List of Excluded Articles with Reason: Article Ang M, Muller AS, Wagenlehner F, Pilatz A, Linn T. Combining protein and carbohydrate increases postprandial insulin levels but does not improve glucose response inpatients with type 2 diabetes. Metabolism. 2012;61(12): DOI: /j.metabol Clifton PM, Galbraith C, Coles L. Effect of a low dose whey/guar preload on glycemic control in people with type 2 diabetes-a randomized control trial. Nutr J. 2014;13(1):103. DOI: / Esteves de Oliveira FC, Pinheiro Volp AC, Alfenas RC. Impact of different protein sources in the glycemic and insulinemic responses. Nutr Hosp. 2011;26(4): DOI: /S Mortensen LS, Homer-Jensen J, Hartvigsen ML, et. Al. Effects of different fractions of whey protein on postprandial lipid and hormone responses in type 2 diabetes. Eur J Clin Nutr. 2012;66(7): DOI: /ejcn Reason Whey protein supplement was combined with Soy Whey protein supplement was combined with fiber Review paper Whey protein fractions 12

14 Summary of Articles Identified to Review: Insulin response is higher when whey protein is added to high carbohydrate meals in both healthy individuals and those with Type 2 Diabetes. Whey protein consumed before a carbohydrate meal can stimulate insulin and incretin hormone secretion and slow gastric emptying, leading to marked reduction in postprandial glycemia. Number of Primary Articles Identified: 9 Number of Review Articles Identified: 1 Total Number of Articles Identified: 10 Number of Articles Reviewed but Excluded: 4 13

15 Overview Table Author, Year, Study Design, Class, Rating Frid A, Nilsson M, Holst JJ, Bjorck I 2005 Non-Randomized Crossover Trial C Neutral Ma J, Stevens JE, Cukier K, et. al Non-Randomized Crossover Trial C Positive Jakubowicz D, Froy O, Ahren B, et. al Randomized Crossover Clinical Trial A Neutral Study Purpose Evaluate if whey supplementation would increase insulin secretion in type 2 diabetic individuals after consumption of a high GI meal Determine if a whey protein preload could slow gastric emptying, stimulate incretin hormones, and attenuate postprandial glycemia in type 2 diabetics. stimulate GLP-1 secretion with a whey protein preload in order to benefit type 2 diabetics from the glucose-lowering effects of these secretions. Study Population Participants with Type 2 diabetes Diet controlled Type 2 Diabetic patients Patients with well-controlled type 2 diabetes who were not taking any medications except for sulfonylurea or metformin. Intervention Outcomes Conclusion Limitations Breakfast whey meal consists of the following: Bread(44.7 g CHO, 11.6 g protein), whey (5.3 g CHO, 18.2 g protein). Lunch whey meal consists of the following: Mashed potatoes (35.6 g CHO, 3.9 g protein), meatballs(5.0 g CHO, 6.5 g protein), whey (5.3 g CHO, 18.2 g protein). Standard meal with 55 g whey protein added to beef soup, standard meal with 55 g whey protein added to mashed potatoes 50 g whey in 250 ml water followed by a high glycemic index breakfast There was a significant difference in postprandial insulin concentrations with the whey protein meal exhibiting a greater insulin response than the reference meal. Significantly lower blood glucose responses and insulin responses were observed when whey was included in the lunch than with the reference meal. Gastric emptying was significantly slowest when whey protein was added as a preload in the beef soup than when ingested with the potato meal or not ingested at all. The iauc for blood glucose was less after the whey preload ( / mmol min ) and whey in meal (406.3+/-85.9 mmol min ) compared with no whey ( / mmol min ) P+/ for both). Plasma glucose was significantly lower in the whey preload group during time intervals 0-30 minutes, minutes, and minutes postprandial than in the control group and was reduced overall 28% after whey preload. Insulin levels were increased 105% in individuals receiving the whey preload. The ingestion of whey protein causes an insulinotropic effect. This could potentially assist in the management of postprandial blood glucose. There is a significant difference in postprandial blood glucose control in diet-controlled type 2 diabetics when whey is consumed before or during a high carbohydrate meal. A whey protein preload before a high glycemic meal increases insulin secretion, enhanced GLP-1 responses, and reduced postprandial glycemia in type 2 diabetic patients. Small number of participants, Non- Randomized Small number of participants, Non- Randomized Possible bias related to funding source, small number of participants, recruitment is unknown 14

16 Petersen BL, Ward LS, Bastian ED, Jenkins AL, Campbell J, Vuksan V 2009 Randomized Control Trial A Negative Akhavan T, luhovyy BL, Brown PH, Cho CE, Anderson GH 2009 Randomized Crossover Trial A Positive Acheson KJ, Blondel-Lubrano A, Oguey-Araymon S, et. al Randomized Crossover Control Trial A Positive Examine the impact of adding escalating doses of a glycemic index lowering peptide fraction (GILP) from whey to a glucose drink to identify food components that may decrease postprandial glycemia. Analyze the effect of the premeal consumption whey protein or its hydrolysate on food intake, pre and post meal satiety, and blood glucose in healthy young adults. Analyze the effects of three proteins on energy metabolism, satiety, and glucose control. Healthy subjects Healthy individuals, normal weight (BMI ) Sedentary, lean, and healthy individuals with normal clinical signs and lab values. Zero, 5, 10, and 20 g of protein from GILP were added to a 50 g glucose drink. The control meal was repeated twice Experiment 1: 10 g, 20 g, 30 g, and 40 g whey protein preload. Experiment 2: 5 g, 10 g, 20 g, and 40 g whey protein preload, and 10 g whey protein hydrolysate preload. All preloads were isovolumetric and served in a 300 ml chilled beverage. Three of the test meals consisted of 50% protein (whey, casein, or soy), 40% carbohydrate, and 10% fat and the fourth meal consisted of 95.5% carbohydrate. These were compared with a glucose meal that provided the same glucose load as the protein meals. Average reduction in the glucose iauc was 4.6 ± 1.4 mmol.min/l per gram of GILP. iaucs for the GILP plus glucose meals of 0, 5, 10 and 20 g of protein were 231 ± 23, 212 ± 23, 196 ± 23, and 138 ± 13 mmol/l min g whey protein suppressed food intake compared with control g whey protein reduced post meal blood glucose and the area under the curve (AUC) (P, 0.05) g whey protein, but not whey protein hydrolysate, reduced post meal blood glucose AUC and insulin AUC with greater reduction occurring with higher doses of whey protein. The glycemic response to glucose was attenuated 32% by the proteins (P, 0.001) at the expense of a greater insulin response after whey than after glucose (154%; P = 0.02), casein (143%; P = 0.07), and soy (151%; P = 0.03). Addition of whey based protein significantly lowers the glycemic response to 50g of glucose. Addition of GILP to carbohydrate food may be an effective way of lowering the glycemic effects of these foods. A whey protein preload reduces food intake, post meal blood glucose and insulin, and the ratio of blood glucose to insulin with greater doses exerting greater effects. Intact whey contributes to blood glucose control by both insulin-dependent and insulin-independent mechanisms. This was not found with the whey protein hydrolysate. Different protein sources could be used to modulate metabolism and energy balance. Possible bias related to employment Small sample size, findings can only be applied to healthy, young adults Small sample size 15

17 Academy of Nutrition and Dietetics Evidence Analysis Library Worksheet Template and Quality Criteria Checklist: Primary Research Citation Study Design Class Frid A, Nilsson M, Holst JJ, Bjorck I. Effect of whey on blood glucose and insulin responses to composite breakfast and lunch meals in type 2 diabetic subjects. Am J Clin Nutr. 2005;82: Non-Randomized Crossover Trial C Quality Rating + (Positive) - (Negative) (Neutral) Research Purpose Inclusion Criteria Exclusion Criteria The purpose of the study is to evaluate if whey protein supplementation would increase insulin secretion in type 2 diabetic individuals after consumption of a high GI meal. Participants with Type 2 diabetes Not clearly defined Recruitment: Individuals were recruited through advertising in a local paper. Of the people who responded, the first 14 to meet all inclusion criteria were admitted to the study. Design: Non-Randomized Crossover Trial Blinding used (if applicable): N/A Intervention (if applicable): Breakfast reference meal consists of the following: Bread (44.7 g CHO, 11.6 g protein), ham (18.2 g protein), lactose (5.3 g CHO). Lunch reference meal consists of the following: Mashed potatoes (35.6 g CHO, 3.9 Description of Study Protocol g protein), meatballs (5 g CHO, 6.5 g protein), ham(18.2 g protein), lactose (5.3 g CHO). Breakfast whey meal consists of the following: Bread(44.7 g CHO, 11.6 g protein), whey (5.3 g CHO, 18.2 g protein). Lunch whey meal consists of the following: Mashed potatoes (35.6 g CHO, 3.9 g protein), meatballs(5.0 g CHO, 6.5 g protein), whey (5.3 g CHO, 18.2 g protein). Statistical Analysis: The GRAPH PAD PRISM was used to calculate incremental areas under the curve (AUCs) for glucose, insulin, GIP, and GLP-1. This was done for each subject and each meal. Significant differences were assessed using analysis of variance. The differences were then assessed using Tukey's multiple comparisons test. P values <0.05 were considered significant. The difference 16

18 between the products at all time points were analyzed using a mixed model with repeated measures and an autoregressive covariance structure. Timing of Measurements: Blood samples were drawn before breakfast (time 0) and at 10, 20, 30, 40, 60, 120, 180, and 240 minutes after breakfast. Immediately Data Collection Summary Description of Actual Data Sample Summary of Results following the 240 minute blood draw the subjects began eating lunch and data was collected at 10, 20, 30, 40, 60, 120, 180, and 240 minutes after lunch. Dependent Variables: Blood glucose, plasma GIP, plasma GLP-1, and serum insulin Independent Variables: Whey protein meal, reference meal Control Variables: Every subject was their own control Initial: 14 (8 Males 6 Females) Attrition (final N): 14 Age: Ethnicity: Unknown Other relevant demographics: Hb A1C (4.3%-7.7%), mean fasting plasma glucose mmol/l Anthropometrics: Mean (+/-SD) body mass index (in kg/m2) of /- 3.1 Location: Unknown Key Findings: There was no significant difference in fasting blood glucose and insulin concentration between the reference meal and the whey breakfast meal days before consumption of the meals. There was no significant difference in the blood glucose response after consuming both the reference meal and the whey protein meal. There was a significant difference in postprandial insulin concentrations with the whey protein meal exhibiting a greater insulin response than the reference meal. There was no significant difference in blood glucose and serum insulin immediately before lunch (240 minutes after breakfast) between the reference meal day and the whey protein meal day. Significantly lower blood glucose responses and insulin responses were observed when whey was included in the lunch than with the reference meal. Other Findings: The GIP responses after lunch were significantly higher after the 17

19 whey meal but no significant difference was observed in the GLP-1 response. The ingestion of whey protein causes an insulinotropic effect. This could potentially assist in the management of postprandial blood glucose. Additional Author Conclusion research is underway to explore the benefits of supplemental protein on blood glucose management. This method of control may have fewer adverse effects than currently used therapeutic agents. Reviewer Comments Funding Source The crossover design which allowed each individual to serve as their own control was one strength in this study. The subjects were also required to be dietcontrolled type 2 diabetics. Unknown Quality Criteria Checklist: Primary Research Symbols Used Explanation + Positive Indicates that the report has clearly addressed issues of inclusion/exclusion, bias, generalizability, and data collection and analysis -- Negative Indicates that these issues have not been adequately addressed. Relevance Questions Neutral indicates that the report is neither exceptionally strong nor exceptionally week Select a rating from the drop-down menu 1. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (NA for some Epi studies) 2. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? 3. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dietetics practice? 1 Yes 2 Yes 3 Yes 4. Is the intervention or procedure feasible? (NA for some epidemiological studies) 4 Yes If the answers to all of the above relevance questions are Yes, the report is eligible for designation with a plus (+) on the Evidence Quality Worksheet, depending on answers to the following validity questions. Validity Questions 1. Was the research question clearly stated? 1.1. Was the specific intervention(s) or procedure (independent variable(s)) identified? 1.2. Was the outcome(s) (dependent variable(s)) clearly indicated? 1.3. Were the target population and setting specified? 1 Yes 1.1 Yes 1.2 Yes 1.3 Yes 18

20 2. Was the selection of study subjects/patients free from bias? 2.1. Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? 2.2. Were criteria applied equally to all study groups? 2.3. Were health, demographics, and other characteristics of subjects described? 2.4. Were the subjects/patients a representative sample of the relevant population? 3. Were study groups comparable? 3.1. Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) 3.2. Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? 3.3. Were concurrent controls used? (Concurrent preferred over historical controls.) 3.4. If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? 3.5. If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable. Criterion may not be applicable in some crosssectional studies.) 3.6. If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., gold standard )? 2 Unclear 2.1 No 2.2 Unclear 2.3 No 2.4 No 3 Yes 3.1 Yes 3.2 Unclear 3.3 N/A 3.4 N/A 3.5 N/A 3.6 N/A 4. Was method of handling withdrawals described? 4.1. Were follow up methods described and the same for all groups? 4.2. Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) 4.3. Were all enrolled subjects/patients (in the original sample) accounted for? 4.4. Were reasons for withdrawals similar across groups 4.5. If diagnostic test, was decision to perform reference test not dependent on results of test under study? 5. Was blinding used to prevent introduction of bias? 5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? 5.2. Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) 5.3. In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? 5.4. In case control study, was case definition explicit and case ascertainment not influenced by exposure status? 5.5. In diagnostic study, were test results blinded to patient history and other test results? 4 N/A 4.1 Yes 4.2 Yes 4.3 Yes 4.4 N/A 4.5 N/A 5 No 5.1 No 5.2 No 5.3 N/A 5.4 N/A 5.5 N/A 6. Were intervention/therapeutic regimens/exposure factor or procedure and any 6 Unclear 19

21 comparison(s) described in detail? Were intervening factors described? 6.1. In RCT or other intervention trial, were protocols described for all regimens studied? 6.2. In observational study, were interventions, study settings, and clinicians/provider described? 6.3. Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? 6.4. Was the amount of exposure and, if relevant, subject/patient compliance measured? 6.5. Were co-interventions (e.g., ancillary treatments, other therapies) described? 6.6. Were extra or unplanned treatments described? 6.7. Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? 6.8. In diagnostic study, were details of test administration and replication sufficient? 7. Were outcomes clearly defined and the measurements valid and reliable? 7.1. Were primary and secondary endpoints described and relevant to the question? 7.2. Were nutrition measures appropriate to question and outcomes of concern? 7.3. Was the period of follow-up long enough for important outcome(s) to occur? 7.4. Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? 7.5. Was the measurement of effect at an appropriate level of precision? 7.6. Were other factors accounted for (measured) that could affect outcomes? 7.7. Were the measurements conducted consistently across groups? 6.1 Yes 6.2 Unclear 6.3 Yes 6.4 Yes 6.5 No 6.6 No 6.7 Unclear 6.8 N/A 7 Yes 7.1 Yes 7.2 Yes 7.3 Yes 7.4 Yes 7.5 Yes 7.6 Yes 7.7 Yes 8. Was the statistical analysis appropriate for the study design and type of outcome indicators? 8.1. Were statistical analyses adequately described the results reported appropriately? 8.2. Were correct statistical tests used and assumptions of test not violated? 8.3. Were statistics reported with levels of significance and/or confidence intervals? 8.4. Was intent to treat analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? 8.5. Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? 8.6. Was clinical significance as well as statistical significance reported? 8.7. If negative findings, was a power calculation reported to address type 2 error? 9. Are conclusions supported by results with biases and limitations taken into consideration? 9.1. Is there a discussion of findings? 9.2. Are biases and study limitations identified and discussed? 10. Is bias due to study s funding or sponsorship unlikely? Were sources of funding and investigators affiliations described? Was there no apparent conflict of interest? 20 8 Yes 8.1 Yes 8.2 Yes 8.3 Yes 8.4 N/A 8.5 No 8.6 No 8.7 N/A 9 Yes 9.1 Yes 9.2 No 10 Unclear 10.1 No 10.2 Unclear MINUS/NEGATIVE (-) If most (six or more) of the answers to the above validity questions are No, the report should be designated with a minus (-) symbol on the Evidence Worksheet. NEUTRAL ( )

22 If the answers to validity criteria questions 2, 3, 6, and 7 do not indicate that the study is exceptionally strong, the report should be designated with a neutral ( ) symbol on the Evidence Worksheet. PLUS/POSITIVE (+) If most of the answers to the above validity questions are Yes (including criteria 2, 3, 6, 7 and at least one additional Yes ), the report should be designated with a plus symbol (+) on the Evidence Worksheet. 21

23 Academy of Nutrition and Dietetics Evidence Analysis Library Worksheet Template and Quality Criteria Checklist: Primary Research Citation Study Design Class Ma J, Stevens JE, Cukier K, et. al. Effects of a protein preload on gastric emptying, glycemia, and gut hormones after a carbohydrate meal in diet-controlled type 2 diabetes. Diabetes Care. 2009;32(9): Non-randomized Crossover Trial C Quality Rating + (Positive) - (Negative) (Neutral) Research Purpose Inclusion Criteria Exclusion Criteria The purpose of the research was to determine if a whey protein preload could slow gastric emptying, stimulate incretin hormones, and attenuate postprandial glycemia in type 2 diabetics. Diet-controlled Type 2 Diabetes patients Not clearly defined Recruitment: Not clearly defined Design: The 8 participants fasted overnight (14 hours for solids and 12 hours for liquids) a total of three times. On each occasion the subject consumed the following: beef-flavored soup (3.8 g beef flavoring dissolved in 350 ml water) 30 min before a mashed potato meal containing 65 g instant potatoes with 20 g glucose. The total macronutrient composition of the meal was 59.1 g carbohydrate, 4.3 g fat, 5.2 g protein, and1,276.5 kj. On day one 55g whey protein was added to the soup. On day two 55 grams of whey protein was added Description of Study Protocol to the potatoes. On day three no whey protein was added to the meal. Gastric emptying was assessed using scintigraphy, blood glucose was assessed using a glucometer, plasma insulin was assessed using enzymelinked immunosorbent assay, and total GLP-1, GIP and CCK-8 were measured by radioimmunoassay. Finally cardiovascular autonomic function was measured using the variation in R-R interval during deep breathing and the systolic blood pressure changes while subject was standing. Blinding used (if applicable): No Intervention (if applicable): The first intervention was the standard meal with 55 g of whey protein added to the beef soup. The second intervention consisted of the standard meal with 55 g of whey protein added to the mashed potatoes. 22

24 Statistical Analysis: Repeated-measures ANOVA with treatment and time as factors. P<0.05 was considered significant. Timing of Measurements: -30, 0, 30, 60, 90, 120, 150, 180, 210, 240, 270, 300 (minutes before and after meal). Data Collection Summary Description of Actual Data Sample Summary of Results Dependent Variables: Gastric emptying, blood glucose, plasma insulin, GLP, GIP- 1, CCK, and cardiovascular autonomic function. Independent Variables: Whey protein preload (55g) and whey protein meal supplement (55g). Control Variables: Fasting procedures and timing of measurement. Initial: 8 (7 Males 1 Females) Attrition (final N): 8 Age: years Ethnicity: Unknown Other relevant demographics: Duration of known diabetes 5.4 +/- 1.1 years, A1C 6.5 +/- 0.2% Anthropometrics: BMI /- 1.3 kg/m2 Location: Adelaide, Australia Key Findings: Gastric emptying was significantly slowest when whey protein was added as a preload in the beef soup than when ingested with the potato meal or not ingested at all (T50: /- 5.4 min; P = ). There were no differences in baseline blood glucose, plasma insulin, GLP-1, GIP, or CCK values. The iauc for blood glucose was less after the whey preload meal and the whey in meal than when no whey was consumed. The iaucs for insulin,glp-1, GIP, and CCK were greater for the whey preload meal and whey in meal than when no whey was consumed. Author Conclusion Other Findings: GLP-1 was greater between 15 minutes and 90 minutes with the whey preload meal compared with the whey in meal but there was no significant difference in iauc overall. There is a significant difference in postprandial blood glucose control in dietcontrolled type 2 diabetics when whey is consumed before or during a high 23

25 carbohydrate meal. The results were similar to those of many pharmacological therapies. Reviewer Comments Funding Source This study consists of a small number of participants with well controlled and uncomplicated diabetes.this was addressed in the conclusion section of the research. National Health and Medical Research Council (NHMRC) of Australia Quality Criteria Checklist: Primary Research Symbols Used Explanation + Positive Indicates that the report has clearly addressed issues of inclusion/exclusion, bias, generalizability, and data collection and analysis -- Negative Indicates that these issues have not been adequately addressed. Relevance Questions Neutral indicates that the report is neither exceptionally strong nor exceptionally week Select a rating from the drop-down menu 5. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (NA for some Epi studies) 6. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? 7. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dietetics practice? 1 Yes 2 Yes 3 Yes 8. Is the intervention or procedure feasible? (NA for some epidemiological studies) 4 Yes If the answers to all of the above relevance questions are Yes, the report is eligible for designation with a plus (+) on the Evidence Quality Worksheet, depending on answers to the following validity questions. Validity Questions 11. Was the research question clearly stated? Was the specific intervention(s) or procedure (independent variable(s)) identified? Was the outcome(s) (dependent variable(s)) clearly indicated? Were the target population and setting specified? 12. Was the selection of study subjects/patients free from bias? Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Were criteria applied equally to all study groups? Were health, demographics, and other characteristics of subjects described? Were the subjects/patients a representative sample of the relevant population? 1 Yes 1.1 Yes 1.2 Yes 1.3 Yes 2 Yes 2.1 Unclear 2.2 Yes 2.3 Yes 2.4 No 24

26 13. Were study groups comparable? Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Were concurrent controls used? (Concurrent preferred over historical controls.) If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable. Criterion may not be applicable in some crosssectional studies.) If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., gold standard )? 3 Yes 3.1 Unclear 3.2 Yes 3.3 Yes 3.4 N/A 3.5 N/A 3.6 N/A 14. Was method of handling withdrawals described? Were follow up methods described and the same for all groups? Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) Were all enrolled subjects/patients (in the original sample) accounted for? Were reasons for withdrawals similar across groups If diagnostic test, was decision to perform reference test not dependent on results of test under study? 15. Was blinding used to prevent introduction of bias? In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? In case control study, was case definition explicit and case ascertainment not influenced by exposure status? In diagnostic study, were test results blinded to patient history and other test results? 16. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were intervening factors described? In RCT or other intervention trial, were protocols described for all regimens studied? In observational study, were interventions, study settings, and clinicians/provider described? Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Was the amount of exposure and, if relevant, subject/patient compliance measured? Were co-interventions (e.g., ancillary treatments, other therapies) described? 4 N/A 4.1 N/A 4.2 Yes 4.3 Yes 4.4 N/A 4.5 N/A 5 Unclear 5.1 No 5.2 Unclear 5.3 N/A 5.4 N/A 5.5 N/A 6 Yes 6.1 Yes 6.2 N/A 6.3 Yes 6.4 Yes 6.5 Yes 6.6 N/A 25

27 16.6. Were extra or unplanned treatments described? Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? In diagnostic study, were details of test administration and replication sufficient? 17. Were outcomes clearly defined and the measurements valid and reliable? Were primary and secondary endpoints described and relevant to the question? Were nutrition measures appropriate to question and outcomes of concern? Was the period of follow-up long enough for important outcome(s) to occur? Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Was the measurement of effect at an appropriate level of precision? Were other factors accounted for (measured) that could affect outcomes? Were the measurements conducted consistently across groups? 6.7 Yes 6.8 N/A 7 Yes 7.1 Yes 7.2 Yes 7.3 Yes 7.4 Yes 7.5 Yes 7.6 Yes 7.7 Yes 18. Was the statistical analysis appropriate for the study design and type of outcome indicators? Were statistical analyses adequately described the results reported appropriately? Were correct statistical tests used and assumptions of test not violated? Were statistics reported with levels of significance and/or confidence intervals? Was intent to treat analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? Was clinical significance as well as statistical significance reported? If negative findings, was a power calculation reported to address type 2 error? 19. Are conclusions supported by results with biases and limitations taken into consideration? Is there a discussion of findings? Are biases and study limitations identified and discussed? 20. Is bias due to study s funding or sponsorship unlikely? Were sources of funding and investigators affiliations described? Was there no apparent conflict of interest? 8 Yes 8.1 Yes 8.2 Yes 8.3 Yes 8.4 N/A 8.5 Unclear 8.6 Yes 8.7 N/A 9 Yes 9.1 Yes 9.2 Yes 10 Yes 10.1 Yes 10.2 Yes MINUS/NEGATIVE (-) If most (six or more) of the answers to the above validity questions are No, the report should be designated with a minus (-) symbol on the Evidence Worksheet. NEUTRAL ( ) If the answers to validity criteria questions 2, 3, 6, and 7 do not indicate that the study is exceptionally strong, the report should be designated with a neutral ( ) symbol on the Evidence Worksheet. PLUS/POSITIVE (+) If most of the answers to the above validity questions are Yes (including criteria 2, 3, 6, 7 and at least one additional Yes ), the report should be designated with a plus symbol (+) on the Evidence Worksheet. 26

28 Academy of Nutrition and Dietetics Evidence Analysis Library Worksheet Template and Quality Criteria Checklist: Primary Research Citation Study Design Class Jakubowicz D, Froy O, Ahren B, et. al. Incretin, insulinotropic and glucose-lowering effects of whey protein pre-load in type 2 diabetes: A randomized clinical trial. Diabetologia. 2014;57(9): DOI: /s x Randomized Crossover Clinical Trial A Quality Rating + (Positive) - (Negative) (Neutral) Research Purpose Inclusion Criteria Exclusion Criteria Description of Study Protocol The objective is to stimulate GLP-1 secretion with a whey protein preload in order to benefit type 2 diabetics from the glucose-lowering effects of these secretions. Patients with well-controlled type 2 diabetes who were not taking any medications except for sulfonylurea or metformin. Patients taking anorectic drugs, steroids, medications known to affect gastric motility or oral hypoglycaemic agents other than sulfonylurea or metformin were excluded. Recruitment: Randomized using a coin flip Design: Subjects consumed on 50 g of whey in 250 ml water or a placebo of 250 ml water followed by a high glycemic index breakfast on two separate days that were seperated by a period of two weeks. A fasting blood sample at 30 minutes prior to meal consumption was taken and the premeal drinks were consumed. Blood samples were taken at t=0 minutes and a high glycemic index breakfast (353 calories) was consumed. Blood was collected at 30, 60, 90, 120, 150, and 180 minutes. Glucose was analyzed using an analyzer, insulin and C-peptide were analyzed using electrochemiluminescence immunoassay, total GLP-1 and intact GLP-1 were quantified by ELISA, and DPP4 was also assessed. On the second meal, participants were crossed over to the opposite pre meal drink. Blinding used (if applicable): Single blind Intervention (if applicable): The intervention was a premeal drink given 30 minutes prior to a high glysemic index breakfast. The premeal drink consisted of 50g of whey dissolved in 250 ml water. Statistical Analysis: The areas under the curve (AUCs) for glucose, intact GLP-1 and insulin during the 30 min following meal ingestion were the primary endpoints for this study and were analyzed using a two-way ANOVA for the time 27

29 series and a least-significant difference t test post hoc analysis for a comparison between the groups at each time point. To assess the fasting values with post prandial time points Dunnet's post hoc test was used and a t-test was used to compare AUCs at different time intervals. Finally a multivariate ANOVA for repeated measurements was used to assess between and within subject effects for diet and time. The AUCs for C-peptide, total GLP-1 and DPP-4 were secondary endpoints in the data analysis. Timing of Measurements: Measurements were taken at -30, 0, 30, 60, 90, 120, Data Collection Summary Description of Actual Data Sample Summary of Results 150, 180 minutes. Dependent Variables: Plasma glucose, intact GLP-1, and insulin, C-Peptide, total GLP-1, and DPP-4 Independent Variables: Timing of measurements Control Variables: Whey preload and high glycemic breakfast Initial: 19 (Unknown Males Unknown Females) Attrition (final N): 15 Age: Unknown Ethnicity: Unknown Other relevant demographics: Unknown Anthropometrics: Unknown Location: Israel Key Findings: Fasting glucose, insulin, and C-peptide showed no significant difference between the two test days or groups. Plasma glucose was significantly lower in the whey preload group during time intervals 0-30 minutes, minutes, and minutes postprandial than in the control group and was reduced overall 28% after whey preload. Insulin levels were increased 105% in individuals receiving the whey preload overall with significantly higher concentrations of insulin at 0-30 minutes, minutes, and minutes postprandial. Other Findings: C-Peptide plasma concentrations were significantly higher in the whey preload group than in the control group at timer interval 0-30 minutes, 60-28

30 120 minutes, and minutes. Intact and total GLP-1 were observed to be higher at all time intervals postprandially in the whey preload group compared to the control group. There was no significant difference in DPP4 after breakfast between the placebo and whey pre-loads. A whey protein preload befor a high glycemic meal increases insulin secretion, Author Conclusion Reviewer Comments Funding Source enhanced GLP-1 responses, and reduced postprandial glycemia in type 2 diabetic patients. Whey protein has potential to be an additional method of managing type 2 diabetes. There could be possible bias through the funding source of this study. This study is small and does not address how individuals were recruited to the study. Anthropometrics and subject data is lacking. The Israeli Ministry of Health and Milk Council Quality Criteria Checklist: Primary Research Symbols Used Explanation + Positive Indicates that the report has clearly addressed issues of inclusion/exclusion, bias, generalizability, and data collection and analysis -- Negative Indicates that these issues have not been adequately addressed. Relevance Questions Neutral indicates that the report is neither exceptionally strong nor exceptionally week Select a rating from the drop-down menu 9. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (NA for some Epi studies) 10. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? 11. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dietetics practice? 1 Yes 2 Yes 3 Yes 12. Is the intervention or procedure feasible? (NA for some epidemiological studies) 4 Yes If the answers to all of the above relevance questions are Yes, the report is eligible for designation with a plus (+) on the Evidence Quality Worksheet, depending on answers to the following validity questions. Validity Questions 21. Was the research question clearly stated? Was the specific intervention(s) or procedure (independent variable(s)) identified? 1 Yes 1.1 Yes 1.2 Yes 29

31 21.2. Was the outcome(s) (dependent variable(s)) clearly indicated? Were the target population and setting specified? 22. Was the selection of study subjects/patients free from bias? Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Were criteria applied equally to all study groups? Were health, demographics, and other characteristics of subjects described? Were the subjects/patients a representative sample of the relevant population? 23. Were study groups comparable? Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Were concurrent controls used? (Concurrent preferred over historical controls.) If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable. Criterion may not be applicable in some crosssectional studies.) If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., gold standard )? 1.3 Yes 2 Unclear 2.1 Yes 2.2 Yes 2.3 No 2.4 Unclear 3 Yes 3.1 Yes 3.2 Unclear 3.3 Yes 3.4 N/A 3.5 N/A 3.6 N/A 24. Was method of handling withdrawals described? Were follow up methods described and the same for all groups? Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) Were all enrolled subjects/patients (in the original sample) accounted for? Were reasons for withdrawals similar across groups If diagnostic test, was decision to perform reference test not dependent on results of test under study? 25. Was blinding used to prevent introduction of bias? In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? In case control study, was case definition explicit and case ascertainment not influenced by exposure status? In diagnostic study, were test results blinded to patient history and other test results? 4 No 4.1 Unclear 4.2 No 4.3 No 4.4 Unclear 4.5 N/A 5 Yes 5.1 Yes 5.2 Unclear 5.3 N/A 5.4 N/A 5.5 N/A 26. Were intervention/therapeutic regimens/exposure factor or procedure and any 6 Yes 30

32 comparison(s) described in detail? Were intervening factors described? In RCT or other intervention trial, were protocols described for all regimens studied? In observational study, were interventions, study settings, and clinicians/provider described? Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Was the amount of exposure and, if relevant, subject/patient compliance measured? Were co-interventions (e.g., ancillary treatments, other therapies) described? Were extra or unplanned treatments described? Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? In diagnostic study, were details of test administration and replication sufficient? 27. Were outcomes clearly defined and the measurements valid and reliable? Were primary and secondary endpoints described and relevant to the question? Were nutrition measures appropriate to question and outcomes of concern? Was the period of follow-up long enough for important outcome(s) to occur? Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Was the measurement of effect at an appropriate level of precision? Were other factors accounted for (measured) that could affect outcomes? Were the measurements conducted consistently across groups? 6.1 Yes 6.2 N/A 6.3 Yes 6.4 Yes 6.5 N/A 6.6 N/A 6.7 N/A 6.8 N/A 7 Yes 7.1 Yes 7.2 Yes 7.3 Yes 7.4 Yes 7.5 Yes 7.6 Yes 7.7 Yes 28. Was the statistical analysis appropriate for the study design and type of outcome indicators? Were statistical analyses adequately described the results reported appropriately? Were correct statistical tests used and assumptions of test not violated? Were statistics reported with levels of significance and/or confidence intervals? Was intent to treat analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? Was clinical significance as well as statistical significance reported? If negative findings, was a power calculation reported to address type 2 error? 29. Are conclusions supported by results with biases and limitations taken into consideration? Is there a discussion of findings? Are biases and study limitations identified and discussed? 30. Is bias due to study s funding or sponsorship unlikely? Were sources of funding and investigators affiliations described? Was there no apparent conflict of interest? 31 8 Yes 8.1 Yes 8.2 Yes 8.3 No 8.4 Unclear 8.5 Unclear 8.6 Yes 8.7 N/A 9 Yes 9.1 Yes 9.2 Yes 10 Unclear 10.1 Yes 10.2 Unclear MINUS/NEGATIVE (-) If most (six or more) of the answers to the above validity questions are No, the report should be designated with a minus

33 (-) symbol on the Evidence Worksheet. NEUTRAL ( ) If the answers to validity criteria questions 2, 3, 6, and 7 do not indicate that the study is exceptionally strong, the report should be designated with a neutral ( ) symbol on the Evidence Worksheet. PLUS/POSITIVE (+) If most of the answers to the above validity questions are Yes (including criteria 2, 3, 6, 7 and at least one additional Yes ), the report should be designated with a plus symbol (+) on the Evidence Worksheet. 32

34 Academy of Nutrition and Dietetics Evidence Analysis Library Worksheet Template and Quality Criteria Checklist: Primary Research Citation Study Design Class Petersen BL, Ward LS, Bastian ED, Jenkins AL, Campbell J, Vuksan V. A whey protein supplement decreases post-prandial glycemia. Nutr J. 2009;8:47. DOI: / Randomized Control Trial A Quality Rating + (Positive) - (Negative) (Neutral) Research Purpose Inclusion Criteria Exclusion Criteria The objective of this study is to examine the impact of adding escalating doses of a glycemic index lowering peptide fraction (GILP) from whey to a glucose drink to identify food components that may decrease postprandial glycemia. Healthy subject Not clearly defined Recruitment: Subjects were recruited through the Glycemic Index Laboratories clinic volunteer roster. Subjects received financial compensation for participation. Design: Participants would perform an overnight fast for hours prior to each test day. They were weighed and the subjects blood was collected. Within 10 minutes the subject consumed the test meal and further blood was collected at 15, 30, 45, 60, 90, and 120 minutes after the beginning of the test meal. A beverage of either one cup of water, two cups of water, tea, or coffee with or without milk was offered to each subject at the first test meal and that beverage Description of Study Protocol choice was offered at every test meal following. Blinding used (if applicable): Not clearly defined Intervention (if applicable): Zero, 5, 10, and 20 g of protein from GILP were added to a 50 g glucose drink. The control meal was repeated twice Statistical Analysis: The incremental area under the curve (iauc) for each test meal per subject was assessed with the trapezoid rule. The mean of the two control meals was used for analysis. Differences between meals was analyzed using repeated ANOVA and adjusted for multiple pair-wise comparisons using the Tukey-Kramer process. Pearson's correlation and linear regression were used to assess the relationship between glucose iauc andgilp doseage. All data are expressed as Mean ± SEM. 33

35 Timing of Measurements: Blood samples were collected at 0, 15, 30, 45, 60, 90, and 120 minutes after the start of the meal. Dependent Variables: Blood glucose Data Collection Summary Description of Actual Data Sample Summary of Results Independent Variables: Timing of measurements Control Variables: Test meals which consisted of 250 ml of water blended with 50 g anhydrous glucose (control), 50 g ofglucose g GILP powder (5 g GILP protein), 50 g of glucose g GILP powder (10 g GILP protein) and 50g of glucose g GILP powder (20 g GILP protein). Doses were calculated to contain 5,10, and 20 g of protein. Initial: 10 (3 Males 7 Females) Attrition (final N): 10 Age: 44.4 ± 9.3 years Ethnicity: Unknown Other relevant demographics: None Anthropometrics: BMI 33.6 ± 4.8 kg/m2 Location: Unknown Key Findings: The incremental areas under the glucose curve decreased as GILP doses increased (Pearson's r = 0.48, p = 0.002). The incremental areas (iauc) under the glucose curve for the 0, 5, 10, and 20 g of protein from GILP were 231 ± 23, 212 ± 23, 196 ± 23, and 138 ± 13 mmol min/l. The iauc of the 20 g GILP was significantly different from control, 5 g GILP and 10 g GILP (p < 0.001). Average reduction in the glucose iauc was 4.6 ± 1.4 mmol.min/l per gram of GILP Other Findings: None Addition of whey based protein significantly lowers the glycemic response to 50g Author Conclusion Reviewer Comments Funding Source of glucose. Addition of GILP to carbohydrate food may be an effective way of lowering the glycemic effects of these foods. This study is a small study done on healthy individuals. Three of the six researchers are emplyees of the company that provided funding for this study. Glanbia Nutritionals.Three of the six researchers are emplyees of Glanbia 34

36 Nutritionals. Quality Criteria Checklist: Primary Research Symbols Used Explanation Positive Indicates that the report has clearly addressed issues of inclusion/exclusion, + bias, generalizability, and data collection and analysis -- Negative Indicates that these issues have not been adequately addressed. Neutral indicates that the report is neither exceptionally strong nor exceptionally week Select a rating from the drop-down menu Relevance Questions 13. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (NA for some Epi studies) 14. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? 15. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dietetics practice? 1 Yes 2 Yes 3 Yes 16. Is the intervention or procedure feasible? (NA for some epidemiological studies) 4 Yes If the answers to all of the above relevance questions are Yes, the report is eligible for designation with a plus (+) on the Evidence Quality Worksheet, depending on answers to the following validity questions. Validity Questions 31. Was the research question clearly stated? Was the specific intervention(s) or procedure (independent variable(s)) identified? Was the outcome(s) (dependent variable(s)) clearly indicated? Were the target population and setting specified? 32. Was the selection of study subjects/patients free from bias? Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Were criteria applied equally to all study groups? Were health, demographics, and other characteristics of subjects described? Were the subjects/patients a representative sample of the relevant population? 33. Were study groups comparable? Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Were concurrent controls used? (Concurrent preferred over historical 1 Yes 1.1 Yes 1.2 Yes 1.3 Unclear 2 No 2.1 No 2.2 Unclear 2.3 Yes 2.4 Unclear 3 Yes 3.1 Yes 3.2 Yes 35

37 controls.) If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable. Criterion may not be applicable in some crosssectional studies.) If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., gold standard )? 3.3 Yes 3.4 N/A 3.5 N/A 3.6 N/A 34. Was method of handling withdrawals described? Were follow up methods described and the same for all groups? Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) Were all enrolled subjects/patients (in the original sample) accounted for? Were reasons for withdrawals similar across groups If diagnostic test, was decision to perform reference test not dependent on results of test under study? 35. Was blinding used to prevent introduction of bias? In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? In case control study, was case definition explicit and case ascertainment not influenced by exposure status? In diagnostic study, were test results blinded to patient history and other test results? 36. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were intervening factors described? In RCT or other intervention trial, were protocols described for all regimens studied? In observational study, were interventions, study settings, and clinicians/provider described? Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Was the amount of exposure and, if relevant, subject/patient compliance measured? Were co-interventions (e.g., ancillary treatments, other therapies) described? Were extra or unplanned treatments described? Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? In diagnostic study, were details of test administration and replication sufficient? 4 Unclear 4.1 Unclear 4.2 Unclear 4.3 Yes 4.4 Unclear 4.5 N/A 5 Unclear 5.1 Unclear 5.2 Unclear 5.3 N/A 5.4 N/A 5.5 N/A 6 Yes 6.1 Yes 6.2 Yes 6.3 Yes 6.4 Yes 6.5 Unclear 6.6 Unclear 6.7 Unclear 6.8 N/A 37. Were outcomes clearly defined and the measurements valid and reliable? 7 Yes 36

38 37.1. Were primary and secondary endpoints described and relevant to the question? Were nutrition measures appropriate to question and outcomes of concern? Was the period of follow-up long enough for important outcome(s) to occur? Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Was the measurement of effect at an appropriate level of precision? Were other factors accounted for (measured) that could affect outcomes? Were the measurements conducted consistently across groups? 7.1 Yes 7.2 Yes 7.3 Yes 7.4 Yes 7.5 Yes 7.6 Unclear 7.7 Yes 38. Was the statistical analysis appropriate for the study design and type of outcome indicators? Were statistical analyses adequately described the results reported appropriately? Were correct statistical tests used and assumptions of test not violated? Were statistics reported with levels of significance and/or confidence intervals? Was intent to treat analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? Was clinical significance as well as statistical significance reported? If negative findings, was a power calculation reported to address type 2 error? 39. Are conclusions supported by results with biases and limitations taken into consideration? Is there a discussion of findings? Are biases and study limitations identified and discussed? 40. Is bias due to study s funding or sponsorship unlikely? Were sources of funding and investigators affiliations described? Was there no apparent conflict of interest? 8 Yes 8.1 Yes 8.2 Yes 8.3 No 8.4 Unclear 8.5 Unclear 8.6 Yes 8.7 N/A 9 Unclear 9.1 Yes 9.2 Yes 10 No 10.1 Yes 10.2 No MINUS/NEGATIVE (-) If most (six or more) of the answers to the above validity questions are No, the report should be designated with a minus (-) symbol on the Evidence Worksheet. NEUTRAL ( ) If the answers to validity criteria questions 2, 3, 6, and 7 do not indicate that the study is exceptionally strong, the report should be designated with a neutral ( ) symbol on the Evidence Worksheet. PLUS/POSITIVE (+) If most of the answers to the above validity questions are Yes (including criteria 2, 3, 6, 7 and at least one additional Yes ), the report should be designated with a plus symbol (+) on the Evidence Worksheet. 37

39 Academy of Nutrition and Dietetics Evidence Analysis Library Worksheet Template and Quality Criteria Checklist: Primary Research Citation Study Design Class Akhavan T, Luhovyy BL, Brown PH, Cho CE, Anderson GH. Effect of premeal consumption of whey protein and its hydrolysate on food intake and postmeal glycemia and insulin responses in young adults. Am J Clin Nutr.2010;91(4): DOI: /ajcn Randomized Crossover Trial A Quality Rating + (Positive) - (Negative) (Neutral) Research Purpose The purpose of this study was to analyze the effect of the premeal consumption whey protein or its hydrolysate on food intake, pre and post meal satiety, and blood glucose in healthy young adults. Inclusion Criteria Healthy individuals, normal weight (BMI ) Exclusion Criteria Description of Study Protocol Breakfast skippers, smokers, dieters, or individuals with diabetes (fasting blood glucose greater than or equal to 7.0 mmol/l) or other metabolic diseases, restrained eaters (identified by a score of greater than or equal to 11 on the Eating Habits Questionnaire), and those taking medication were excluded. Recruitment: Recruited through advertisements posted on the University of Toronto campus. Participants were compensated financially. Design: Experiement 1: A standard breakfast of a single serving cheerios, 250 ml box of 2% milk, and a 250 ml box of orange juice (300 kcal) was consumed by each participant 4 hours prior to scheduled study session and after a 10 hour overnight fast. Participants were instructed to refrain from alcohol and unusual exercise and activity the night before each study. On arrival the subjects completed questionnaires concerning Sleep Habits, Stress Factors, Food Intake and Activity Level, Feelings of Fatigue, and Motivation to Eat. A baseline blood sample was collected to measure insulin and glucose concentration. Men were provided the preloads in random order once per week. They were instructed to drink the preloads within 5 minutes at a constant pace and remained seated throughout the experimental session. At 30 minutes from the time the subject began drinking the preload the participant was fed an ad libitum pizza meal. Subjects were provided 3 types of pizza based on preference and were asked to eat until they were "comfortably full". They were allowed 20 minutes to eat. Immediately after the meal and at 50, 65, 80, and 95 minutes 38

40 subjective appetite and blood glucose were measured. Experiment 2: A standard breakfast of a single serving cheerios, 250 ml box of 2% milk, and a 250 ml box of orange juice (300 kcal) was consumed by each participant 4 hours prior to scheduled study session and after a 10 hour overnight fast. Participants were instructed to refrain from alcohol and unusual exercise and activity the night before each study. On arrival the subjects completed questionnaires concerning Sleep Habits, Stress Factors, Food Intake and Activity Level, Feelings of Fatigue, and Motivation to Eat. A baseline blood sample was collected to measure insulin and glucose concentration. after measurement of blood glucose,300 ll capillary blood was collected into blood collection tubes. Insulin was measured by enzyme immunoassay. Men were provided the preloads in random order twice weekly and women in the second experiment were studied twice a week in the first 2 weeks of their menstrual cycle. They were instructed to drink the preloads within 5 minutes at a constant pace and remained seated throughout the experimental session. At 30 minutes from the time the subject began drinking the preload the participant was fed a fixed quantity of Deluxe pizza with a bottle of 500 ml of spring water. They were allowed 20 minutes to eat. Immediately after the meal and at 65, 80, 95, 110, 140, and 170 minutes subjective appetite and blood glucose were measured. Insulin was measured at 0, 30, 50, 95, 110, 140, and 170 minutes. Blinding used (if applicable): Single Intervention (if applicable): Experiment 1: 10 g, 20 g, 30 g, and 40 g whey protein preload. Experiment 2: 5 g, 10 g, 20 g, and 40 g whey protein preload, and 10 g whey protein hydrolysate preload. All preloads were isovolumetric and served in a 300 ml chilled beverage. Statistical Analysis: Two and 3-factor repeated-measures ANOVA were used to assess time, sex, preload, and their effect on outcome variables. Average appetite scores, blood glucose, and insulin responses were also analyzed. When a preload and time was statistically significant, a one-factor ANOVA was followed by Tukey s post hoc test to analyze the effect of the preload on the baseline for blood glucose and insulin at each time of measurement. Premeal changes from 39

41 baseline were calculated from 0 min and postmeal changes from 30 min before meal consumption. The effect of preload on food intake, cumulative energy intake, and caloric compensation (in experiment 1) and on premeal, postmeal, and AUC for appetite and blood glucose (in both experiments) and insulin (in experiment 2) were tested by one factor ANOVA followed by Tukey s post hoc test to identify differences between preloads. Pearson s correlation coefficients were used to detect associations between dependent measures. Significance was set at P, Data are presented as means 6 SEMs. The average appetite score of the 4 appetite queations in the "Motivation to Eat" survey was calculated for statistical analysis. Data Collection Summary Description of Actual Data Sample Timing of Measurements: Experiment 1: Immediately after the meal and at 50, 65, 80, and 95 minutes subjective appetite and blood glucose were measured. Experiment 2: Immediately after the meal and at 65, 80, 95, 110, 140, and 170 minutes subjective appetite and blood glucose were measured. Insulin was measured at 0, 30, 50, 95, 110, 140, and 170 minutes. Dependent Variables: Blood glucose, subjective appetite, insulin Independent Variables: Timing of measurements, standard breakfast of a single serving cheerios, 250 ml box of 2% milk, and a 250 ml box of orange juice (300 kcal), lunch meals of an ad libitum pizza meal (experiment 1) and a fixed quantity of Deluxe pizza with a bottle of 500 ml of spring water (experiment 2). Control Variables: Whey protein preload. Experiment 1: 10 g, 20 g, 30 g, and 40 g whey protein preload. Experiment 2: 5 g, 10 g, 20 g, and 40 g whey protein preload, and 10 g whey protein hydrolysate preload. Initial: Experiement 1: 16 men Experiment 2: 22 (12 Males 10 Females) Attrition (final N): Experiment 1: 16 Experiment 2: 21 ( 12 Males 9 Females) Age: years Ethnicity: Unknown Other relevant demographics: None Anthropometrics: Experiment 1: Men (n = 16) had a mean body weight of /- 40

42 1.6 kg, height of 1.8 +/- 0.0 m, and body mass index of / Experiment 2: Men (n = 12) and women (n = 9) with a mean age body weight of /- 2.1 and /- 2.2 kg, height of 1.8 +/- 0.0 and /- 0.0 m, and body mass index of /- 0.5 and / Location: Toronto, Canada Summary of Results Key Findings: Experiment 1: g whey protein suppressed food intake (P, ) compared with control g whey protein reduced postmeal blood glucose and the area under the curve (AUC) (P, 0.05). Experiment 2: g whey protein, but not whey protein hydrolysate, reduced postmeal blood glucose AUC and insulin AUC with greater reduction occurring with higher doses of whey protein (P, 0.05). The ratio blood glucose to insulin AUCs (0 170 min) was reduced by greater than or equal to 10 g whey protein but not by 10 g whey protein hydrolysate. Other Findings: Experiment 1: Average appetite was not affected by preload or time interval. Experiment 2: Average appetite was affected by preload (P, 0.01) and sex (P, ). A whey protein preload reduces food intake, postmeal blood glucose and insulin, and the ratio of blood glucose to insulin with greater doses exerting greater Author Conclusion Reviewer Comments Funding Source effects. Intact whey contributes to blood glucose control by both insulindependent and insulin-independent mechanisms. This was not found with the whey protein hydrolysate. In experiment 2, whey protein hydrolysate was administered in a 10g dose. With higher doses there may illicit greater responses to insulin and glucose blood concentrations. Both experiments 1 and 2 had small sample sizes and all participants were healthy young adults. As a result the findings can only be applied to healthy, young adults.bias in these studies is unlikely. Supported by a grant from the Natural Sciences and Engineering Research Council of Canada Quality Criteria Checklist: Primary Research 41

43 Symbols Used Explanation + Positive Indicates that the report has clearly addressed issues of inclusion/exclusion, bias, generalizability, and data collection and analysis -- Negative Indicates that these issues have not been adequately addressed. Relevance Questions Neutral indicates that the report is neither exceptionally strong nor exceptionally week Select a rating from the drop-down menu 17. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (NA for some Epi studies) 18. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? 19. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dietetics practice? 1 Yes 2 Yes 3 Yes 20. Is the intervention or procedure feasible? (NA for some epidemiological studies) 4 Yes If the answers to all of the above relevance questions are Yes, the report is eligible for designation with a plus (+) on the Evidence Quality Worksheet, depending on answers to the following validity questions. Validity Questions 41. Was the research question clearly stated? Was the specific intervention(s) or procedure (independent variable(s)) identified? Was the outcome(s) (dependent variable(s)) clearly indicated? Were the target population and setting specified? 42. Was the selection of study subjects/patients free from bias? Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Were criteria applied equally to all study groups? Were health, demographics, and other characteristics of subjects described? Were the subjects/patients a representative sample of the relevant population? 43. Were study groups comparable? Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Were concurrent controls used? (Concurrent preferred over historical controls.) If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable. Criterion may not be applicable in some cross- 1 Yes 1.1 Yes 1.2 Yes 1.3 Yes 2 Yes 2.1 Yes 2.2 Yes 2.3 Yes 2.4 Yes 3 Yes 3.1 Yes 3.2 Yes 3.3 Yes 3.4 N/A 3.5 N/A 42

44 sectional studies.) If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., gold standard )? 3.6 N/A 44. Was method of handling withdrawals described? Were follow up methods described and the same for all groups? Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) Were all enrolled subjects/patients (in the original sample) accounted for? Were reasons for withdrawals similar across groups If diagnostic test, was decision to perform reference test not dependent on results of test under study? 45. Was blinding used to prevent introduction of bias? In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? In case control study, was case definition explicit and case ascertainment not influenced by exposure status? In diagnostic study, were test results blinded to patient history and other test results? 46. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were intervening factors described? In RCT or other intervention trial, were protocols described for all regimens studied? In observational study, were interventions, study settings, and clinicians/provider described? Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Was the amount of exposure and, if relevant, subject/patient compliance measured? Were co-interventions (e.g., ancillary treatments, other therapies) described? Were extra or unplanned treatments described? Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? In diagnostic study, were details of test administration and replication sufficient? 47. Were outcomes clearly defined and the measurements valid and reliable? Were primary and secondary endpoints described and relevant to the question? Were nutrition measures appropriate to question and outcomes of concern? Was the period of follow-up long enough for important outcome(s) to occur? Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Was the measurement of effect at an appropriate level of precision? 4 Yes 4.1 Yes 4.2 Yes 4.3 Yes 4.4 Yes 4.5 N/A 5 Yes 5.1 Yes 5.2 Unclear 5.3 N/A 5.4 N/A 5.5 N/A 6 Yes 6.1 Yes 6.2 N/A 6.3 Yes 6.4 Yes 6.5 Yes 6.6 N/A 6.7 Unclear 6.8 N/A 7 Yes 7.1 Yes 7.2 Yes 7.3 Yes 7.4 Yes 7.5 Yes 43

45 47.6. Were other factors accounted for (measured) that could affect outcomes? Were the measurements conducted consistently across groups? 7.6 Yes 7.7 Yes 48. Was the statistical analysis appropriate for the study design and type of outcome indicators? Were statistical analyses adequately described the results reported appropriately? Were correct statistical tests used and assumptions of test not violated? Were statistics reported with levels of significance and/or confidence intervals? Was intent to treat analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? Was clinical significance as well as statistical significance reported? If negative findings, was a power calculation reported to address type 2 error? 49. Are conclusions supported by results with biases and limitations taken into consideration? Is there a discussion of findings? Are biases and study limitations identified and discussed? 50. Is bias due to study s funding or sponsorship unlikely? Were sources of funding and investigators affiliations described? Was there no apparent conflict of interest? 8 Yes 8.1 Yes 8.2 Unclear 8.3 Yes 8.4 Yes 8.5 Yes 8.6 Yes 8.7 N/A 9 Yes 9.1 Yes 9.2 Yes 10 Yes 10.1 No 10.2 Yes MINUS/NEGATIVE (-) If most (six or more) of the answers to the above validity questions are No, the report should be designated with a minus (-) symbol on the Evidence Worksheet. NEUTRAL ( ) If the answers to validity criteria questions 2, 3, 6, and 7 do not indicate that the study is exceptionally strong, the report should be designated with a neutral ( ) symbol on the Evidence Worksheet. PLUS/POSITIVE (+) If most of the answers to the above validity questions are Yes (including criteria 2, 3, 6, 7 and at least one additional Yes ), the report should be designated with a plus symbol (+) on the Evidence Worksheet. 44

46 Academy of Nutrition and Dietetics Evidence Analysis Library Worksheet Template and Quality Criteria Checklist: Primary Research Citation Study Design Class Acheson KJ, Blondel-Lubrano A, Oguey-Araymon S, et. al. Protein choices targeting thermogenesis and metabolism. Am J Clin Nutr. 2011;93: Randomized Crossover Control Trial A Quality Rating + (Positive) - (Negative) (Neutral) Research Purpose Inclusion Criteria Exclusion Criteria The objective of the study is to analyze the effects of three proteins on energy metabolism, satiety, and glucose control. Sedentary, lean, and healthy individuals with normal clinical signs and lab values. Not explicitly outlined Recruitment: Participants were recruited from among the Nestle Research Center's Personnel. Design: 23 participants visited the clinic on six separate occasions. On occasion one the subject came to the clinic after an overnight fast and the participant's RMR was measured. The recorded values were used to determine the 24 hour energy requirements of each subject. On occasions 2-6, the subject appeared at the clinic the day before the test and consumed a controlled breakfast, lunch, and dinner and spent the night in the clinic. The following morning subjects were woken at 6 am. Overnight urine was collected and participants were transferred Description of Study Protocol to a whole body indirect calorimetry chamber where continuous respiratory exchange measurements were made continuously for 4 hours. Patients consumed one of 4 isocaloric test meals (three of the test meals consisted of 50% protein (whey, casein, or soy), 40% carbohydrate, and 10% fat and the fourth meal consisted of 95.5% carbohydrate. These were compared with a glucose meal that provided the same glucose load as the protein meals) and exited the chamber for a second urine collection. Respiratory exchange measurements occurred for an additional 2 1/2 hours and participants exited the chamber for a third urine sample. At each visit the subject consumed a different test meal selected at random. Blinding used (if applicable): Double 45