REDUCING VARIABILITY

The typical variable and error rate in global glycemic testing has been shown to reach 80 percent, which is not acceptable for United States (U.S.) government claims on foods. To reduce this error-rate and variable down to less than 2 percent, Glycemic Research Laboratories (GRL) re-structured and re-designed glycemic testing protocols, which are now utilized in every clinical study.

Additionally, glycemic indices for foods can differ by fivefold, depending on level of adipose tissue body fat, metabolic Syndrome, BMI, insulin-resistance, diabetes, food form, and measurement/testing methods used.

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Analysis Directive
Glycemic Research Laboratories
Copyright © 2007
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Simultaneous ingestion of carbohydrate and protein reduces glycemic response in some foods, while protein ingestion increases insulin response. Ingesting carbohydrates with fat typically blunts blood glucose effect, but does not effect insulin.

Glycemic Research Laboratories (GRL) testing protocol has the highest rate of accuracy available (less than 2 % variability), with specific in-real-time analytical testing methods specifically developed by Glycemic Research Laboratories.

Specific protocols have been developed by Glycemic Research Laboratories for testing carbohydrate foods versus protein foods versus -0- calorie and low calorie foods.

Each GRL clinical protocol is designed to mitigate variables and stay within FDA and FTC legal guidelines for claims.

The variable reduction methodologies designed by Glycemic Research Laboratories are proprietary.

TARGETED PROTOCOLS

Targeted protocols are available to clients seeking clarification in glycemic and other metabolic responses. Targeted protocol subjects are selected on the basis of:

Analysis Directive
Glycemic Research Laboratories
Copyright © 2007
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CLINICAL INVESTIGATION PROTOCOLS

GLYCEMIC INDEXING
High glycemic foods and beverages that elevate blood glucose and insulin levels cause weight gain, increased diabetes risk, and tremendous metabolic stress on the human body, as the body compensates for excessive insulin levels by producing more adrenaline, cortisol, and other stress hormones.

Adrenaline, cortisol, and other stress hormones have two major effects:

MEALS
High glycemic meals promote elevated blood glucose and insulin levels, as well as direct adipose tissue fat storage. The glucose excursion that follows a low versus a high glycemic index meal directly affects postprandial glycemia. As an example, the change in plasma glucose one hour after eating 50 g of spaghetti is half of that seen 1 hour after eating 50 g of white bread (Reference: Glycemic Research Laboratories clinical trial for Mueller’s Pasta).

The glycemic response to a mixed meal can be identified by feeding subjects weighed portions of a mixed meal with varying percentages of carbohydrates, proteins, and fat.

Glycemic Research Laboratories conducts trials on mixed meals and frozen meals.

Analysis Directive
Glycemic Research Laboratories
Copyright © 2007
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GLYCEMIC RESPONSE: ALL CALORIES ARE NOT EQUAL

The Glycemic Response of foods & beverages refers to the effects elicited by oral ingestion of any edible agent (not just carbohydrate foods) on blood glucose concentration and insulin levels during the digestion process.

All foods and beverages can be designed and/or re-formulated to moderate and reduce blood glucose and insulin responses by utilizing Glycemic Research Laboratories Clinical Investigation Protocols.


DIABETICS
The Nurses’ Health Study, Harvard Medical School, found that “Women who ate the most foods with a high glycemic index had a 50% greater risk of diabetes than those who ate the least.”

The study went on report: “Not all foods affect blood glucose levels in the same way. Some foods have what is called a high glycemic index, which means that they can raise blood glucose levels rapidly.

Eating a lot of high glycemic index foods forces the body to produce insulin in large amounts to try to clear the high levels of glucose in the blood. Over time, this increase in insulin production can increase the risk of diabetes.”

Glycemic Research Laboratories Clinical Investigation Protocols provide a better understanding of the diabetic properties and risk associated with foods and beverages, as well as Nutraceuticals and Pharmaceuticals.

This allows for proper formulation and marketing of said products, and for design and re-formulating options by clients.

NURSES’ HEALTH STUDY ANNUAL NEWSLETTER
Volume 9, 2003
Nurses’ Health Study
Harvard Medical School

Analysis Directive
Glycemic Research Laboratories
Copyright © 2007
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SPORTS DRINK PROTOCOLS
In the development of sports-performance-related-products, professional athletes may be utilized. High glycemic sports drinks reduce sports performance (GRI Human Maximum Performance report), and are therefore contraindicated for professional athletes.

BEVERAGE PROTOCOLS
Zero-calorie beverages are no longer the answer to the growing obesity issue. Beverages that contain -0- calories and -0- carbohydrates are capable of increasing diabetes risk, and adding body weight, via the Cephalic Response.

Therefore, typical glycemic studies are no longer the sole answer to understanding the metabolic response of beverages.

Services are available to beverage clients seeking to identify the biochemical properties of a beverage, or to re-design current beverage products, and/or to develop new beverages. Targeted Clinical Investigation Protocols seek to identify the major factors involved in creating beverages.

Beverages focusing on the “Diet” market are encouraged to select protocols targeted to analyze:

SWEETENERS/SUGARS
Sugars and sweeteners, despite the caloric or carbohydrate content, are capable of high glycemic reactions on blood glucose and insulin levels. Sweeteners previously believed to have a glycemic response of zero have recently been proven to have definite glycemic properties.

In the case of sweeteners, the Test Food is prepared per instructions and confirmed by Brix refractometry.

STEVIA
Doses as low as 1 gram of Stevia elicit a glycemic index in clinical trials. As doses of Stevia increase, so does the glycemic index.

Sugar alcohols are not the preferred sweetener or bulk, as they can cause flatulence or a laxative effect in varying degrees in some individuals. This is due to their incomplete absorption (in the small intestine) properties.

Many food manufacturers claim that sugar alcohols do not affect blood sugar levels, but in reality, they do affect the postprandial blood glucose response in individuals both with and without diabetes.

PROTEIN TEST FOODS
Proteins eaten without carbohydrates can induce high glycemic responses and fat storage in humans. Consumption of high amounts of meats or protein (more than 30 grams ingested at one time) triggers adipose tissue fat storage and spillage into the urea cycle, causing liver and kidney problems, such as elevated liver enzymes, which can disqualify individuals from obtaining personal health insurance.

In many cases, ketogenic diets; high protein diets (Atkins, etc.), are responsible for skewed blood profiles.

Removing the patient from a high protein diet for 4-6 weeks typically returns serum profiles to normal.

Analysis Directive
Glycemic Research Laboratories
Copyright © 2007
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GI of ALCOHOL
Most alcoholic beverages contain low amounts of carbohydrate, ranging from 0 to 4 grams per 100 ml. Beer contains 3-4 grams of carbohydrate per 100 ml. Therefore, consuming large quantities of beer can over-elevate blood glucose levels. Consuming one glass of beer slightly elevates blood glucose levels.

The high caloric-values of alcohol respond to stimulation of fat-storage in humans. Favored-wines commonly contain high glycemic sugars, which can over-elevate blood glucose and insulin levels, independent of their alcohol content.

LEGAL SERVING SIZES
Legal use of the term “Low Glycemic” in the United States, as dictated by the Federal government, requires “appropriate serving size” amounts used in clinical tests.

Appropriate serving sizes are utilized during GRL clinical studies. In order to make the claim of “Low Glycemic” for any human-grade food product, the United States government requires Board Approved human In Vivo clinical trials.

In Vitro and non-clinical trial calculations, and/or software that claims to be able to determine glycemic index are not legally permitted for product labeling.

Glycemic Research Laboratories In Vivo Clinical trials focus on glycemic index, glycemic load, glycemic response, insulin response, Genetic Profiling, Metabolic Syndrome, fat-storing mechanisms and factors; Lipoprotein Lipase, Leptin, Neuropeptide Y, and Cephalic Response.

Test Food (s) are fed to pre-screened human subjects selected for specific protocols, such as diabetics, non-diabetics, obese, age, ethnic, children, and targeted other groups.

Protocols are designed by the GRL Medical Advisory Board (see About Us at www.GlycemicResearchLaboratories.com) based on the Protocol Design Session.

PROTOCOL DESIGN SESSION

The client participates in a Protocol Design Session prior to the testing phase, which includes:

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Glycemic Research Laboratories
Clinical Testing Methodologies
Copyright © 2007
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GENETIC VARIANCES in OBESITY
According to the American Diabetes Association (publication January 2007), “Half the U.S. population has the gene that puts them at greater risk of developing diabetes. The gene causes people to metabolize fat differently and may hurt their ability to remove sugar from the blood.” This genetic variant alters the way half the population in America processes food, driving foods into fat cells.

Many other genetic traits in humans have been identified which alter food and beverage metabolism. Foods and beverages can be formulated to address genetically-hard-wired metabolic variances as related to obesity, overweight, fat-cell activity, diabetes, and insulin-disorders.

METABOLIC SYNDROME SCREENING
Clients may elect to utilize subjects with Metabolic Syndrome as defined herein, or to eliminate all subjects diagnosed in-house with Metabolic Syndrome.

STANDARD CLINICAL DEFINITION of METABOLIC SYNDROME*

NEW PRODUCT DEVELOPMENT & FORMULATION ASSISTANCE

Clients submitting new products may opt for New Product Trial Feedback (NPTF) prior to finalizing a formula.

This option entails pre-testing of formulas to develop the most glycemically acceptable form of the Test Food, assistance with ingredients selection, pre-screening and testing of formula ingredients and options, and preferred-outcome selection of formula raw materials.

Glycemic Research Laboratories, Medical Advisory Board, represent expertise in glycemic product development, having received the first glycemic patent ever awarded worldwide. For the past 23 years, GRL staff has been at the forefront of glycemic research and development.

TRADE SECRETS
Glycemic Research Laboratories is bound to protect, and hold private, trade and formula secrets involved in product testing and product development. GRL does not publish any clinical trial results, without express written permission from clients, as this would compromise proprietary product development.

GRL proprietary low glycemic, non-Cephalic development protocols are held in strict confidence by the GRL development staff, and are not made public in any circumstances whatsoever.

In the case of proprietary product development, and patent applications, Glycemic Research Laboratories will not accept competing-development projects (on a case-by-case basis).

Glycemic Research Laboratories conducts testing and product development for the largest food companies in the world, and as such, does not compromise proprietary trade secrets.

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Glycemic Research Laboratories
Study Options
Copyright © 2007
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CERTIFICATION TRIALS
If clients intend to apply for the Glycemic Research Institute (GRI) Certification Marks; Low Glycemic and/or Low Glycemic for Diabetics, GRL will apply appropriate protocols during the trial period, as specified by GRI’s guidelines.

To utilize GRI’s Diabetic Certification Marks, it is mandatory to use diabetic subjects, as diabetics respond very differently than non-diabetics to foods, drinks, and Nutraceuticals ingested.

The Glycemic Research Institute is a non-profit organization that allows Pro Bono use of its Federally registered Certification Marks, based on submitted and accepted Human In Vivo Clinical trials. The Glycemic Research Institute does not accept In Vitro or non-approved clinical trials as acceptable proof of glycemic response. The certification Marks may be viewed at www.Glycemic.com.

TRADE JOURNAL PUBLICATION
Protocols can be specifically designed to meet the requirements of peer reviewed journals. This must be implemented prior to the onset of the GRL clinical trial.

The glycemic index is a numerical classification based on Human In Vivo clinical trials designed to quantify the relative blood glucose response to foods, drinks, Nutraceuticals, Pharmaceuticals, and any edible agent.

Glycemic Research Laboratories (GRL) Human In Vivo Clinical trials have been developed over a 20-year period, focusing on reduction of testing variables. GRL trials are conducted under direction of the Glycemic Research Laboratories (GRL) Medical Advisory Board, M.D.’s, Professor’s of Medicine, and PhD statisticians.

Testing methods are approved by the Institutional Review Boards for the State of Florida, and the International Clinical Study Review Board. Specific analytical testing methods are the property of GRL.

METHODS
All blood work and analytical calculations are conducted in-house in Real-Time. Utilizing standardized Glycemic Research Laboratories Board-Approved clinical protocols, accommodations are made for low-end or high-end carbohydrate Test Foods.

Ten pre-screened human subjects are typically used for each product tested. Clients may elect to use larger pools of subjects.

White bread is used as the standard. Each subject is fed a minimum of three bread standards for comparison to the products tested. Calculations are made using the area under the curve (AUC) as compared to bread standards (converted to the glucose scale). AUC is calculated by GRL statisticians using standard Glycemic Research Laboratories protocols.

Fasting blood glucose measurements are made, and at 15-minute intervals throughout the trial, for 2-4 hours, or until blood glucose levels stabilize.

Capillary blood is preferred: the results for capillary blood glucose (BG) are less variable than that of venous plasma glucose. Additionally, elevations in BG are greater in capillary blood than venous plasma, and the differences in Test Foods and bread standards are easier to detect statistically using capillary blood glucose.

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Glycemic Research Laboratories
Research Design & Methods
Copyright © 2007
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PROTOCOLS
When VBP is called for in clinical trials, the GRL protocol calls for an overnight fast of 12 h, a blood sampling i.v. cannula was inserted into the antecubital vein. Blood samples are taken at -5, -10 and -15 minutes (analysed as a pool) before the Test Food, and every 15 minutes for the first hour, and every 30 minutes thereafter, to a 5-hour postprandial period.

Taste, mouth-feel, gastrointestinal issues; nausea, flatulence, bloating, are recorded.

Results presented in the final Test Food report are based on the glucose scale. Glycemic index and glycemic load values are converted to the glucose = 100 scale by multiplication with the factor 0.7.


SUBJECTS undergo a two-visit protocol, the first to determine glucose tolerance status and the second to measure SI. Subjects fast for 12 h before each of the two visits, and abstain from alcohol for 24 h. Smoking is prohibited on the day of the study.

Anthropometric measures are taken for each subject. Height and weight are measured in duplicate and recorded to the nearest 0.5 cm and 0.1 kg, respectively. BMI is calculated as weight (in kilograms) divided by the square of height (in meters). Waist circumference is measured at the natural indentation or at a level midway between the iliac crest and the lower edge of the rib cage if no natural indentation was visible. Waist is recorded to the nearest 0.5 cm, and the mean of two measures within 1 cm of each other is used.

A 2-h, 75-g oral glucose tolerance test is performed during the first visit, and World Health Organization (WHO) criteria is used to assign glucose tolerance status. Subjects taking oral hypoglycemic medications are classified as type 2 diabetics.

Glycemic Research Laboratories
Research Design & Methods
Copyright © 2007
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Acute insulin response (AIR) and SI are assessed using a 12-sample, insulin-enhanced, frequently sampled intravenous glucose tolerance test (FSIGT) with minimal model analysis. Modifications of the protocol are used when appropriate for targeted Trials. AIR and fasting insulin are log transformed: logarithmic transformations, the disposition index, typically calculated as the product of AIR and SI, is preferentially created as the sum of log (AIR + 20) and log (SI + 1).

AIR is calculated based on insulin levels through the 8-min blood samples before insulin infusion. Fasting plasma insulin was determined by radioimmunoassay.

SI is calculated by mathematical modeling methods; the time course of plasma glucose was fit using nonlinear least squares methods with the plasma insulin values as a known input to the system.

Mean glycemic index values are assigned to white bread standard purchased at available grocery stores.

In our subject pre-screening, typical glycemic index (GI) and glycemic load (GL) are 58 and 128 g/day, respectively. A higher SI value expresses increased insulin sensitivity, while higher fasting insulin implies increased insulin resistance. Higher AIR indicates greater insulin secretion in response to glucose, and higher disposition index implies increasing pancreatic functionality. Positive linear relationships are observed between food/liquid intake and levels of fasting insulin, BMI, and waist circumference.

Adjustments are made for non-carbohydrate Test Foods using the Residual Method.

Dietary fiber intake and measures of SI, insulin secretion and adiposity are made, including multivariate adjustment and scoring, as dietary fiber in a Test Food is associated with SI, fasting insulin, BMI, and waist circumference. In our trials, it is observed that 1 8-10 gram fiber content is associated with lower level of fasting insulin with statistically higher level of SI. Significant linear relationship between glycemic load and outcome levels is observed, that are positive for fasting insulin, BMI, and waist circumference and inverse for SI.

Outliers are recorded.

Subject responses to Test Food activation of adipose-tissue fat-storage mechanisms, IE LPL, are tracked and recorded per GRL protocols.

Glycemic Research Laboratories
Research Design & Methods
Copyright © 2007
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Glycemic Index (GI) alone is unable to predict the glycemic response/impact when different amounts of carbohydrates are eaten. Glycemic Load must be utilized in conjunction with GI to differentiate the acute impact on blood glucose and insulin responses induced by Test Foods.


GLYCEMIC LOAD (GL)
Glycemic Load is based on a specific quantity and carbohydrate content of the test food. GL is calculated by multiplying the weighted mean of the dietary glycemic index by the percentage of total energy from the test food.

When the test food contains quantifiable carbohydrates, the Glycemic Load equals GI (%) x grams of carbohydrate per serving. One unit of GL approximates the glycemic effect of 1 gram of glucose. Typical diets contain from 60-180 GL units per day.


A HIGH GLYCEMIC LOAD diet is defined as: 60% carbohydrate, 20% protein, 20% fat (glycemic load 116 g/1000 kcal).

A LOW GLYCMIC LOAD diet is defined as: 40% carbohydrate, 30% protein, 30% fat, (glycemic load 45 g/1000 kcal).

GLUCOSE SCALE
Results presented in the final Test Food report are based on the glucose scale. Glycemic index and glycemic load values are converted to the glucose = 100 scale by multiplication with the factor 0.7.

SAMPLE STUDY
The following Human In-Vivo Clinical Trial was conducted by Glycemic Research Laboratories (GRL) in 2007, and is utilized as an example (Report ID: GTD-0307) of a typical GRL clinical trial. No copies of this report may be made, transferred, or used in any format whatsoever, and remains the sole property of Glycemic Research Laboratories.

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Mathematic Modeling Methods
Glycemic Research Laboratories.
Copyright © 2007

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Mathematic Modeling Methods
Glycemic Research Laboratories.
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