Food design in clinical trials: the impact of food formulation on - PowerPoint PPT Presentation

Food design in clinical trials: the impact of food formulation on absorption and metabolism of bioactive compounds Jennifer Ahn-Jarvis Jennifer.Ahn-Jarvis@quadram.ac.uk 2 July 2019

OSU Agriculture Campus- Wooster OSU Food Science Department Quadram Experimental Kitchen Clinical Research Facility Test Kitchen OSU Medical Center

Importance of Oral Disease • Global Burden of Disease 2015 study reports that an estimated 3.5 billion people are affected with oral disease 1 . • Untreated caries is the most prevalent oral condition in the world. • Oral disease is linked to many diseases 2,3 1. Kassebaum, N.J. et al J Dent Res 2017;96:380-387 2. Williams, R.C. et al Curr Med Res Opin 2008;24:1635-43 https://doctorbrianwest.com/services/m 3. Kaye, E.A. J Am Dent Assoc 2007;138:616-9. outh-body-connection/

Oral Health and Diet • Diets rich in fruits and vegetables have shown to promote oral health by improving overall nutrition and microbiome profile 1,2 . • Among the many fruits, black raspberries ( Rubus occidentalis ) have shown in rodent and cell studies to improve oral health and prevent oral cancer 3,4 . • Clinical trials have shown mixed effects 1. Winn, D.M. Am J Clin Nutr 1995 Feb;61:437S-445S 2. Hezel, M.P. and Weitzberg, E. Oral Dis . 2015;21:7-16 3. Casto, B.C. et al Anticancer Res 2002; 22:4005-15 4. Mallery, S.R. et al Cancer Prev Res 2011; 8:1209-21

Oral Residence Time and Oral Metabolites Enhanced Absorption vs. Enhanced Degradation 1 Mouth Esophagus Conjugated-cyanidin Aglycone-cyanidin Mono or disaccharide Cyanidin degradation products BRB : Freeze-dried black raspberries 1. Ahn-Jarvis et al Cancer Res 2014;74:19

Gut derived metabolites Ellagitannins Ellagic acid Dimethyl ellagic acid Urolithin D Proposed pathway of ellagic acid metabolism by gut microbiome Urolithin C Isourolithin A Urolithin B 1. Tomás ‐ Barberán et al Mol Nutr Food Res . 2017;61:1-35

Standardization of Black Raspberry Powder (BRB) Ellagitannin (Sanguiin H-6) mAU 520 nm Cyanindin-3-xylorutinoside 70 260 nm Cyanindin-3-glucoside 355 nm 60 Quercetin hexonic acid 50 Ellagic acid- pentose 40 Ellagic Acid 30 20 Rutin 10 0 6 7 8 9 10 11 12 min Large Scale Production Black raspberries were HPLC chromatogram of BRB mixture after blending and peak procured from Dale Stokes in Wilmington, Ohio (15 kg) identities verified using external standards and tandem mass and Berri Health, Oregon (6 kg). spectroscopy 1 1. Gu et al J Agric Food Chem . 2014;62:3997-4006

Dissolution Apparatus to Approximate Oral Residence Time a 100% fast Percentage of Total Phenolic Released b intermediate 80% slow 60% c* 40% Hard Candy Saliva Hard Candy PBS Starch- Saliva USP I dissolution apparatus was used to determine oral 20% Pectin- Saliva residence time of various amorphous forms using PBS Pectin PBS and synthetic saliva (100 RPM, pH 6.5 at 37°C) 6,7 . Starch PBS 0% 0 50 100 150 200 250 300 Time (minutes) Three amorphous forms show 3 distinct release behaviors 1 *Complete dissolution occurred at 600 minutes. Different letters denote differences in Hard Candy Pectin Starch rate of total phenolic release using ANOVA (p≤0.05) with Tukey’s posthoc test (visco-elastic) (elastic) (glassy) 1. Gu et al J Food Sci . 2015; 80:E610-8

Sensory Evaluation for Clinical Adherence and Optimization of Formulation Hard Candy Like 9 Pectin Extremely Starch 8 9 Point Hedonic Scale a a 7 ab a a b a a a ab 6 b a b a b b b 5 c 4 Sensory Evaluation confections conducted under sensory booth 3 conditions (n=65). Letters denote mean separation where significant 2 differences (p≤0.05) were found Dislike 1 using ANOVA with Tukey’s Extremely Overall Color Aroma Fruit Flavor Sweetness Texture posthoc test. Liking Attributes

Optimized Confection Formulations Hard Pectin Starch Composition (g) Candy Confection Confection BRB powder 1.25 1.25 1.25 Sugar 3.15 2.18 0.63 Corn syrup 1.85 0.75 2.28 Water - 1.85 1.70 Starch/pectin - 0.09 (pectin) 0.39 (starch) 50% (w/w) citric acid - 0.13 - Total 6.25 6.25 6.25 BRB: Standardized freeze-dried black raspberry powder Starch: Tate and Lyle Confectioners G cornstarch Pectin: Danisco Grindsted CF 130B

Study Design n = 10 Berry Confection 2: Hard Candy R Dose 1 (3 pieces / day) 67 Adults A Berry Confection 2: Hard Candy n = 10 (33 men and 34 N Dose 2 (6 pieces / day) women) D Berry Confection 2: Pectin n = 10 Dose 1 (3 pieces / day) O 18 to 65 years old n = 10 Berry Confection 2: Pectin BMI 18 to 35 kg/m 2 M Dose 2 (6 pieces / day) Never or past smokers I Berry Confection 3: Starch n = 10 for >10 years Z Dose 1 (3 pieces / day) E n = 10 Berry Confection 3: Starch Dose 2 (6 pieces / day) Washout (2 weeks) Low phenolic diet Intervention (2 weeks) Day 14 Day -14 Day 0 -buccal scrape -buccal scrape -consent and enrollment -urine collection -urine collection -instruction of washout (restricted low -blood collection -blood collection phenolic diet) - multivitamin regimen -Durability Test

Demographic and Adherence Measures of Confection Study 33 ± 11 years old Age (mean ± SD) 26.6 ± 9.2 kg/m 2 BMI (mean ± SD) Compliance (mean ±SD) 93.2 ± 15.0% Hard Candy 93.9 ± 12.8% Pectin 96.4 ± 12.3% Starch 13% (9/67) History of dental surgery 66% (44/67) Twice daily teeth brushing 69% (46/67) Daily flossing Regular alcohol consumers (average 3.25 servings/ week) 69% (46/67) 89% (60/67) Never smokers 11% (7/67) Past smokers (0.5 pack/ 5.5 years)

Oral Residence Time (Assessment of Confection Durability) Each subject was asked to record start and stop times for each confection • Three confections were administered in a randomized crossover design during a single visit. • Participants were instructed to vigorously tumble confection but prohibited from chewing *Oral residence time in minutes (A) and quantity of saliva produced confection. with each confection (B). Letters denote differences using ANOVA (p≤0.05) with Tukey’s posthoc test

Salivary Metabolites Maximum total cyanidin in 9 7.E+05 confection (8.5 ± 0.6 mg) hard 8 pectin Total Anthocyanins in saliva collected 6.E+05 starch 7 Relative abundance (mAU) 5.E+05 6 b 4.E+05 5 b 4 3.E+05 3 a 2.E+05 2 1.E+05 1 0 0.E+00 Cyanidin-3-glucoside pca pga Different letters denote significant (p≤0.05) differences usingANOVAwith Tukey’s posthoc test

24 hour Total Urolithins • 3 confections at two doses (4 and 8g/day) consumed for 14 days (n=62). • Total urolithin reflects urolithin A to D and isourolithin A. Different letters denote mean separation where significant differences (p≤0.05) were found in total urolithin in 24 hour urine using ANOVA with Tukey’s posthoc test

Conclusions 1. Oral residence time : hard candy was the most durable having longest oral residence time resulting in the greatest volume of saliva 2. Salivary metabolites : cyanidin-3-glucoside significantly less during hard candy consumption but ellagitannin and ellagic acid increased. 3. Huge inter-individual differences observed in the rate of confection consumption, quantity of saliva produce, and polyphenolic profile 4. 24 hour urine: No differences in total urolithins excreted with 4g dose but significant differences were observed between pectin and hard candies with 8g dose of BRB suggesting that confection matrix may impact ellagic acid metabolism

Acknowledgements College of Public Health – Division of Environmental and Health Sciences Christopher Weghorst, Ph.D. Thomas Knobloch, Ph.D. Steve Oghumu, Ph.D. College of Agriculture and Food Sciences Department of Food Science and Technology Ken Riedl, Ph.D. Steven J. Schwartz, Ph.D. Matthew Teegarden, M.S. Yael Vodovotz, Ph.D. Center for Advancement of Functional Foods and Entrepreneurship Sensory and Clinical Studies Volunteers College of Medicine CTOC Postdoctoral Fellowship National Institute of Dental and Division of Internal Medicine Craniofacial Research (T32 DE014320). Steven K. Clinton, M.D., Ph.D. Comprehensive Cancer Center and the National Cancer Institute Department of Biomedical Statistics (P30 CA016058). Denis Pearl, Ph.D. Center for Clinical and Translational Science and the National College of Pharmacy Center For Advancing Translational Sciences (UL1TR001070). James Fuchs, Ph.D. United States Department of Agriculture: USDA 38903 ‐ 03560