Patent Publication Number: US-2022232860-A1

Title: Stevia flavor compositions

Description:
RELATED APPLICATIONS 
     This patent application incorporates by reference each of the following in their entirety: International Patent Application No. PCT/US2019/022581, filed on Mar. 15, 2019; International Patent Application No. PCT/US2019/022456, filed on Mar. 15, 2019; U.S. Provisional Application No. 62/806,646, filed on Feb. 15, 2019; U.S. Provisional Application No. 62/802,111, filed on Feb. 6, 2019; U.S. Provisional Application No. 62/771,937, filed on Nov. 27, 2018; U.S. Provisional Application No. 62/682,461, filed on Jun. 8, 2018; International Patent Application No. PCT/US2018/026920, filed on Apr. 10, 2018; U.S. Provisional Application No. 62/644,407, filed on Mar. 17, 2018; and U.S. Provisional Application No. 62/644,065, filed on Mar. 16, 2018. 
    
    
     SEQUENCE LISTING 
     The text file entitled “PC_77WO_Final_ST25.txt,” created on Apr. 10, 2018, having 15 kilobytes of data, and filed concurrently herewith, is hereby incorporated by reference in its entirety in this application. 
     SUMMARY OF THE INVENTION 
     The present invention is directed to novel  stevia  flavor compositions comprising a high level of rebaudioside AM (Reb AM). These compositions are flavors with modifying properties, or FMPs. An FMP is a flavor ingredient that modifies the flavor profile of a consumable product without imparting detectable sweetness to the product. Using an FMP, certain flavor notes in a consumable product can be enhanced, while other flavor notes may be suppressed. 
     The chemical structure of Reb AM is shown in  FIG. 1 . 
     The high level of Reb AM is obtained in the compositions by a bioconversion process to convert certain steviol glycoside molecules into Reb AM. The compositions have a total steviol glycoside content that is greater than or equal to about 80%, preferably greater than or equal to about 90% and more preferably greater than or equal to about 95%. The Reb AM content in the composition can range from greater than 10%, or greater than 50%, preferably greater than about 65%, and more preferably greater than about 70% In certain embodiments, the Reb AM content is about 68%, or about 72%, or about 75%. In other embodiments, the Reb AM content ranges from about 10-85%. Other levels of Reb AM content are encompassed by this invention. 
     The ratio of Reb AM to Reb M in the  stevia  flavor compositions may range, on a percent dry weight basis, from about 1:1 to about 10:1, preferably from about 2:1 to about 7:1, and more preferably from about 3:1 to about 5:1. In certain embodiments, the Reb AM to Reb M dry weight ratio is about 3.7:1, or about 4.8:1, or about 4.3:1. Other percent dry weight ratios of Reb AM to Reb M in the  stevia  flavor composition are contemplated by this invention. 
     The balance of the  stevia  flavor composition comprises other steviol glycosides, including known and unnamed steviol glycosides, found in the extract of  stevia  leaves. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1  shows the chemical structure of rebaudioside AM. 
         FIG. 2  shows the pathways of producing rebaudioside AM and various steviol glycosides from steviol. 
         FIG. 3  shows the biocatalytic production of rebaudioside AM from stevioside using the enzymes UGTS12 and UGT76G1 and concomitant recycling of UDP to UDP-glucose via sucrose synthase SuSy_At. 
         FIG. 4 , at the end of this document, shows the biocatalytic production of rebausioside AM from rebaudioside E using the enzyme UGT76G1 and concomitant recycling of UDP to UDP-glucose via sucrose synthase SuSy_At. 
         FIG. 5  is a graph showing the effect of Reb AM on the flavor modification of coconut water. 
         FIG. 6  is a graph showing the effect of Reb AM on the flavor modification of a chocolate protein shake. 
         FIG. 7  is a graph showing the effect of Reb AM on the flavor modification of vanilla cinnamon oatmilk. 
         FIG. 8  is a graph showing the effect of Reb AM on the flavor modification of strawberry jam. 
         FIG. 9  is a graph showing the effect of Reb AM on the flavor modification of a sugar reduced lemon poppy seed muffin. 
         FIG. 10  is a graph showing the effect of Reb AM on the flavor modification of seasoned peanuts. 
         FIG. 11  is a graph showing the effect of Reb AM on the flavor modification of a Moscow mule. 
         FIG. 12  is a graph showing the effect of Reb AM on the flavor modification of corn puff cereal. 
     
    
    
     DETAILED DESCRIPTION 
       FIG. 2  shows pathways of producing rebaudioside AM and various steviol glycosides from steviol. 
       FIG. 3  shows the biocatalytic production of rebaudioside AM from stevioside using the enzymes UGTS12 and UGT76G1 and concomitant recycling of UDP to UDP-glucose via sucrose synthase SuSy_At. 
       FIG. 4  shows the biocatalytic production of rebausioside AM from rebaudioside E using the enzyme UGT76G1 and concomitant recycling of UDP to UDP-glucose via sucrose synthase SuSy_At. 
     As used herein, the term “SuSy_AT”, unless specified otherwise, refers to sucrose synthase having amino-acid sequence “SEQ ID 1” of the attached Sequence Listing. 
     As used hereinafter, the term “UGTS12”, unless specified otherwise, refers to UDP-glucosyltransferase having amino-acid sequence “SEQ ID 2” of the attached Sequence Listing. 
     As used hereinafter, the term “UGT76G1”, unless specified otherwise, refers to UDP-glucosyltransferase having amino-acid sequence “SEQ ID 3” of the attached Sequence Listing. 
     One bioconversion process for making the  stevia  flavor composition, identified as “PCS-3028”, is described as follows: 
     
       
         
           
               
             
               
                   
               
               
                 Method of Manufacture 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Botanical Source 
                   Stevia rebaudiana  (Bertoni) Bertoni dried leaves 
               
               
                 Yield based on original source 
                 90% 
               
               
                 material, % 
                   
               
            
           
           
               
            
               
                 Detailed description of the steps involved in the isolation and purification of the natural complex 
               
               
                 Stage 1 
               
               
                 1 part of dried  stevia  leaves soaked in 10 parts of water (50° C., 1 hr) and the mixture is separated from 
               
               
                 biomass to obtain filtrate with dissolved glycosides. The glycosides are extracted by RO (reverse 
               
               
                 osmosis) water. No other solvents are used during this stage. The process is carried out in a continuous 
               
               
                 counter current extractor. 
               
               
                 Stage 2 
               
               
                 Filtrate with dissolved glycosides is treated with flocculant [Ca(OH) 2  2.5% wt] to precipitate 
               
               
                 polysaccharides, proteins and pigments. The treatment with flocculant is carried out at ambient 
               
               
                 temperature during 30 min. Then the mixture is filtered to obtain purified filtrate with dissolved 
               
               
                 glycosides. The filtration is carried out on a “frame-and-plate” type filter press similar to ones used in 
               
               
                 sugar production. 
               
               
                 Stage 3 
               
               
                 Purified filtrate is passed through columns packed with food grade ion exchange resins (Amberlite 
               
               
                 FPC 23 or equivalent in H +  form, and Amberlite FPA 51 or equivalent in OH −  form). Ion exchange 
               
               
                 resins are used to remove inorganic salts and organic ions. Cations are removed by Amberlite FPC 23 
               
               
                 (H + ), and anions by Amberlite FPA 51 (OH − ). The deionization step is carried out at ambient 
               
               
                 temperature. The solution is fed to columns at 0.5 BV/hr. This step provides deionized filtrate. 
               
               
                 Stage 4 
               
               
                 Deionized filtrate is passed through column packed with macroporous adsorbent resin (Diaion HP20 
               
               
                 or equivalent, ambient temperature, 0.5 BV/hr). Macroporous adsorbents are widely used in different 
               
               
                 food and pharmaceutical applications: separation of valuable compounds from plant extracts, 
               
               
                 extraction of antibiotic intermediates from fermentation broth, separation of peptides, or food 
               
               
                 additives, debittering of citrus juice etc. Steviol glycosides which have high affinity to macroporous 
               
               
                 resin adsorb to it whereas other impurities pass through the column freely. The steviol glycosides 
               
               
                 adsorbed on the resin are eluted by 70% (vol.) Ethanol to provide aqueous alcohol solution of steviol 
               
               
                 glycosides. Food grade Ethanol (complies with FCC VIII) is used in this stage. Ethanol is evaporated 
               
               
                 by a conventional distillation process and the resulting solution is deionized again with food grade ion 
               
               
                 exchange resins. The refined solution is concentrated and spray dried to yield crude  Stevia  extract. 
               
               
                 Stage 5 
               
               
                 Crude  Stevia  extract is dissolved in 5 volumes of Ethanol (&gt;90% v/v) and Rebaudioside A (reb A) 
               
               
                 crystallization is carried out at 20° C. during 24 hrs. The filtrate is separated by centrifugation and the 
               
               
                 Ethanol from obtained filtrate is evaporated by a conventional distillation process. The filtrate is then 
               
               
                 spray dried to yield Stevioside 50 powder (containing approx. 50% stevioside and 20% reb A). 
               
               
                 Stage 6 
               
               
                 Stevioside 50 powder is dissolved in 5 volumes of Methanol (&gt;90% v/v) and stevioside crystallization 
               
               
                 is carried out at 20° C. during 24 hrs. The filtrate is separated by centrifugation and the Methanol from 
               
               
                 obtained filtrate is evaporated by a conventional distillation process. Crystals are then dried in a rotary 
               
               
                 drum vacuum dryer at 110° C. and 10 mbar, sifted through US 80 mesh stainless steel screens and 
               
               
                 passed through metal detectors to yield Stevioside 75 powder (containing approx. 75% stevioside and 
               
               
                 10% reb A). 
               
               
                 Stage 7 
               
               
                 Food enzymes UDP-glucosyltransferase (UGT; EC 2.4.1.x) and a sucrose-synthase (SuSy; EC 
               
               
                 2.4.1.13) are manufactured separately according to the following procedure, based on generally 
               
               
                 available and accepted methods used for production of microbial enzymes.  E. coli  production strain 
               
               
                 LE1B109 carrying the expression vector for the corresponding enzyme gene is inoculated in sterilized 
               
               
                 culture medium and fermented at a pH of between 6-8 and temperature of 25-37° C. until desired 
               
               
                 enzyme production yield is reached (usually &gt;15 hrs) and the fermentation is stopped. The enzyme 
               
               
                 is then isolated by first separating the biomass from the culture broth by standard solid/liquid 
               
               
                 separation techniques (centrifugation and/or filtration). The biomass is homogenized to disrupt the 
               
               
                 bacterial cells and treated with nuclease to degrade DNA/RNA nucleic acids released upon cell 
               
               
                 disruption. This is followed by solid/liquid separation to further remove cell debris and other insoluble 
               
               
                 matter. The cell-free supernatant is filtered to obtain the purified enzyme preparation. 
               
               
                 Stage 8 
               
               
                 Stevioside 75 powder (containing approx. 75% stevioside) and food-grade sucrose are dissolved in 
               
               
                 RO water and uridine 5′-diphosphate disodium salt (5′-UDP-Na 2 ; CAS No. 27821-45-0), UGT and 
               
               
                 SuSy enzymes are added to the solution. The reaction mixture is incubated at 40-50° C. during 10-48 
               
               
                 hrs of reaction. The resulting reaction mixture may be heated up to 80-100° C. for 10 minutes to 
               
               
                 inactivate enzymes. 
               
               
                 Stage 9 
               
               
                 The reaction mixture is filtered as described in Stage 2. The obtained clear solution is deionized by 
               
               
                 ion-exchange resins as described in Stage 3. The deionized filtrate is passed through column packed 
               
               
                 with macroporous adsorbent resin and the steviol glycosides adsorbed on the resin are eluted by 
               
               
                 aqueous ethanol, as described in Stage 4. Ethanol is evaporated by a conventional distillation process 
               
               
                 and the resulting solution is concentrated and spray dried to yield crude Rebaudioside AM powder. 
               
               
                 The obtained powder is sifted through US 80 mesh stainless steel screens and passed through metal 
               
               
                 detectors to yield PCS-3028  stevia  extract. Obtained product is a free flowing powder. 
               
            
           
           
               
               
            
               
                 Solvents Used 
                 Water, Ethanol, Methanol 
               
               
                 Reagents Used 
                 Ca(OH) 2 , sucrose, 5′-UDP-Na 2 , UGT, SuSy 
               
               
                   
               
            
           
         
       
     
     Other embodiments of the process are contemplated, including the use of different reagents such as different ion-exchange and adsorption resins, different apparatus including drying apparatus, different screen sizes, and different production strains for the enzymes. One skilled in the art will understand that variations of the reagents, solvents and apparatus will adjust the reaction parameters, and the invention is not limited to this embodiment. 
     The following Summary and Examples exemplify the use of the high Reb AM  stevia  flavor composition in various consumable applications, and the sweetness detection threshold determination of PCS-3028. 
     Summary of Sensory Tests Performed with PCS-3028 
     1. Threshold: 
     50 ppm of PCS-3028 solution in water provided sweetness perception significantly lower than that of 1.5% sugar solution. Therefore we selected 50 ppm of PCS-3028 as the recognition threshold concentration. 
     2. Beverage: 
     a. Raspberry Watermelon Coconut water (PCS-3028 usage level 50 ppm). The results indicated the test sample PCS-3028 had significantly higher watermelon flavor and overall liking compared to the control samples (at 95% confidence).
 
b. Test sample PCS-3028 had significantly lower sweet aftertaste intensity compared to the control samples (at 90% confidence).
 
     3. Dairy Products: 
     a. Chocolate Protein Shake with Reb A (PCS-3028 usage level 50 ppm). The panel found the test sample containing 50 ppm of PCS-3028 to be significantly lower bitterness, metallic note, whey protein and lower bitter aftertaste than the control (at 95% confidence).
 
b. The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher in cocoa flavor, dairy notes, vanilla flavor, and overall liking (at 95% confidence).
 
     4. Imitation Dairy: 
     a. Vanilla Cinnamon Oat milk with Reb A (PCS-3028 usage level 50 ppm). The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher cinnamon flavor, vanilla flavor, and overall liking than the control (at 95% confidence).
 
b. The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher in sweet aftertaste (at 90% confidence).
 
     5. Jams &amp; Jellies: 
     a. Strawberry Fruit Prep with Reb A (PCS-3028 usage level 50 ppm). The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher cooked strawberry flavor, and overall liking than the control (at 95% confidence and 90% confidence respectively).
 
b. The panel found the test sample containing 50 ppm of PCS-3028 to be significantly lower in bitter aftertaste (at 90% confidence).
 
     6. Baked Goods: 
     a. Threshold: 150 ppm of PCS-3028 in Lemon Poppy Seed Muffin provided sweetness perception significantly lower than that of 4.0% sugar solution. Therefore we selected 150 ppm of PCS-3028 as the recognition threshold concentration.
 
b. Lemon Poppy Seed Muffins (PCS-3028 usage level 150 ppm). The results indicated the test sample PCS-3028 had significantly higher lemon and brown flavor intensity compared to the control samples (at 95% confidence).
 
     7. Snack Foods/Seasoning/Nut Products: 
     a. Seasoned peanuts with Reb A (PCS-3028 usage level 50 ppm). Test sample had significantly less bitterness and lower aftertaste (sweet and bitter) than the Control sample (at 90% and 95% confidence respectively). 
     8. Alcoholic Beverages: 
     a. Moscow Mule (PCS-3028 usage level 50 ppm). The results indicated the test sample PCS-3028 had significantly higher ginger flavor and alcohol intensity compared to the control samples (at 95% confidence). 
     9. Breakfast Cereal: 
     a. Threshold: 150 ppm PCS-3028 in corn puff breakfast cereal provided sweetness perception significantly lower that 3.0% sugar solution. Therefore, we selected 150 ppm of PCS-3028 as the recognition threshold concentration.
 
b. Vanilla corn puffs (PCS-3028 usage level 150 ppm). Test sample had significantly higher vanilla, brown, corn flavor and overall liking than the Control sample (95% confidence). Additionally the test sample showed higher sweet aftertaste at 90% confidence.
 
     Example 1 
     Sweetness Perception 
     Threshold with PCS-3028 
     Application: Neutral Water 
     Test #1 
     The sweetness perception of 1.5% sugar solution and different solutions of PCS-3028 were tested with a consumer panel and found that 50 ppm of PCS-3028 solution in water provided sweetness perception significantly lower than that of 1.5% sugar solution. Therefore we selected 50 ppm of PCS-3028 as the recognition threshold concentration. 
                             METHODOLOGY                                        Nature of Participants:   Company employees       Number of Sessions   1       Number of Participants:   30       Test Design:   2- AFC, Balanced, randomized within pair.           Blind       Sensory Test Method:   Intensity ratings       Environmental Condition   Standard booth lighting       Attributes and Scales:   Which sample is sweeter?       Statistical Analysis:   Paired comparison Test       Sample Size   ~1.5 oz. in a clear capped plastic cup       Serving Temperature   Room temperature (~70° F.)       Serving/Panelists   Samples served simultaneously. Panelists       Instruction:   instructed to read ingredient statement,           evaluate each sample.                    
This is a re-evaluation of the recognition threshold concentration to follow methodology outlined in section 1.4.2 of the “Guidance for the Sensory Testing of Flavorings with Modifying Properties within the FEMA GRAS™ Program”.
 
     
       
         
           
               
            
               
                   
               
               
                 DATA: n = 30 
               
            
           
           
               
               
            
               
                 Two-Tailed Analysis Table 
                   
               
               
                 Report for Result PCS-3028 
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 30 ppm 
                 Percent 
                 Binomial Distribution 
               
               
                   
                 1.5% 
                 PCS- 
                 Frequency 
                 Probability 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Sucrose 
                 3028 
                 Sample 1 
                 P-value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 PC 
                 29 
                 1 
                 96.7% 
                 0.0001 
                 *** 
               
               
                 % Frequency 
                 96.7% 
                 3.3% 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
            
               
                   
               
               
                 DATA: n = 30 
               
            
           
           
               
               
            
               
                 Two-Tailed Analysis Table 
                   
               
               
                 Report for Result PCS-3028 
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 50 ppm 
                 Percent 
                 Binomial Distribution 
               
               
                   
                 1.5% 
                 PCS- 
                 Frequency 
                 Probability 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Sucrose 
                 3028 
                 Sample 1 
                 P-value 
                 Sig 
               
               
                   
               
               
                 PC 
                 21 
                 9 
                 70.0 
                 0.04 
                 *** 
               
               
                 % Frequency 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
            
               
                   
               
               
                 DATA: n = 30 
               
            
           
           
               
               
            
               
                 Two-Tailed Analysis Table 
                   
               
               
                 Report for Result ISO3026A 
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 70 ppm of 
                 Percent 
                 Binomial Distribution 
               
               
                   
                 1.5% 
                 PCS- 
                 Frequency 
                 Probability 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Sucrose 
                 3028 
                 Sample 1 
                 P-value 
                 Sig 
               
               
                   
               
               
                 PC 
                 3 
                 27 
                 10.0% 
                 0.000 
                 *** 
               
               
                 % Frequency 
                 10% 
                 90% 
               
               
                   
               
            
           
         
       
     
     Example 2 
     Raspberry Watermelon Coconut Water with PCS-3028 
     Application: Non-Alcoholic Beverage 
     SUMMARY 
     Thirty company employees evaluated two samples of raspberry watermelon flavored coconut water for overall acceptance and attribute intensities (sweetness, Raspberry flavor, watermelon flavor, coconut water flavor, saltiness, bitterness, and sweet aftertaste, bitter aftertaste) in two sessions. In session one, the two samples included: 1) store-bought Raspberry Watermelon Coconut Water control sample and 2) store-bought Raspberry Watermelon Coconut Water test sample containing PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of a non-alcoholic beverage. The results indicated the test sample PCS-3028 had significantly higher mango peach flavor, coconut water flavor, and overall liking compared to the control samples (at 95% confidence). 
     Project Objective 
     The project objective is to assess if the addition of  stevia  extract solids has an effect on key flavor attributes in various beverage applications. 
     Test Objective 
     The test objective is to determine if the flavor profile and overall acceptance of a Control sample of flavored coconut water differs from a Test sample of the same beverage containing PCS-3028. 
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity and acceptance ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
                   
               
            
           
           
               
            
               
                 Overall Acceptance on a 10-pt hedonic scale where 10 = Extremely 
               
               
                 Like and 0 = Extremely Dislike 
               
               
                 Overall liking, sweetness, raspberry flavor, watermelon flavor, coconut 
               
               
                 water flavor, astringency, artificial chemical note, bitterness, and 
               
               
                 sweet aftertaste, bitter aftertaste. 10-pt continuous intensity scale 
               
               
                 where 0 = Imperceptible and 10 = Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Refrigerated temperature (~45° F.) 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     Samples 
       
     
       
         
           
               
            
               
                   
               
               
                 Beverage Type I, Non-alcoholic 
               
            
           
           
               
               
               
            
               
                   
                 Reference 
                 PCS-3028 
               
               
                   
               
            
           
           
               
               
               
            
               
                 *Coconut water raspberry watermelon juice 
                 100 
                 99.995 
               
               
                 PCS-3028 
                   
                 0.005 
               
               
                 Total (g) 
                 100 
                 100 
               
               
                   
               
               
                 *Vita Coco store brand 
               
            
           
         
       
     
     Results 
     Table 1 (below) summarizes the overall acceptance and mean attribute intensity results for each sample. 
     
       
         
           
               
             
               
                 TABLE 1 
               
             
            
               
                   
               
               
                 Mean Scores Raspberry Watermelon 
               
               
                 Coconut Water with 50 ppm PCS-3028 
               
               
                 Summary of Mean-Scores, P-Values, and Significance 
               
               
                 Test Result Code: Coconut Water (raspberry/ 
               
               
                 watermelon flavor) PCS-3028 at 50 ppm 
               
               
                 This test was performed on 30 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                 Coconut 
                 Coconut water 
                   
                   
               
               
                   
                 water 
                 with 50 ppm of 
                 P- 
                   
               
               
                 Attribute 
                 control 
                 PCS-3028 
                 Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 4.38 
                 4.44 
                 0.6990 
                 NS 
               
               
                 Bitter Intensity 
                 0.32 
                 0.24 
                 0.4267 
                 NS 
               
               
                 Astringency 
                 1.04 
                 1.10 
                 0.4942 
                 NS 
               
               
                 Coconut Flavor 
                 4.89 
                 5.11 
                 0.4372 
                 NS 
               
               
                 Watermelon Flavor 
                 b 
                 a 
                 0.0221 
                 *** 
               
               
                   
                 3.85 
                 4.41 
                   
                   
               
               
                 Raspberry Flavor 
                 0.68 
                 0.95 
                 0.2423 
                 NS 
               
               
                 Artificial/Chemical Note 
                 2.94 
                 2.55 
                 0.2583 
                 NS 
               
               
                 Sweet Aftertaste 
                 a 
                 b 
                 0.0905 
                 ** 
               
               
                   
                 1.60 
                 1.33 
                   
                   
               
               
                 Bitter Aftertaste 
                 0.36 
                 0.29 
                 0.5409 
                 NS 
               
               
                 Overall Liking 
                 b 
                 a 
                 0.0710 
                 ** 
               
               
                   
                 4.49 
                 5.04 
               
               
                   
               
               
                 *= 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             The results indicated the test sample PCS-3028 had significantly higher watermelon flavor and overall liking compared to the control samples (at 95% confidence). 
             Test sample PCS-3028 had significantly lower sweet aftertaste intensity compared to the control samples (at 90% confidence). 
           
         
       
    
     Example 2 Conclusion 
     Thirty company employees evaluated two samples of Raspberry Watermelon flavored coconut water for overall acceptance and attribute intensities (sweetness, watermelon flavor, raspberry flavor, coconut water flavor, astringency, artificial/chemical note, bitterness, and sweet aftertaste, bitter aftertaste) in two sessions. In session one, the two samples included: 1) store-bought Raspberry Watermelon Coconut Water control sample and 2) store-bought Raspberry Watermelon Coconut Water test sample containing PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of a non-alcoholic beverage. The results indicated the test sample PCS-3028 had significantly higher watermelon flavor and overall liking compared to the control samples (at 95% confidence).
 
Test sample PCS-3028 had significantly lower sweet aftertaste intensity compared to the control samples (at 90% confidence).
 
     Example 3 
     Chocolate Protein Shake with PCS-3028 
     Application: Milk/Dairy Product 
     SUMMARY 
     Thirty company employees evaluated two samples of chocolate flavored dairy protein shake for overall acceptance and attribute intensities (cocoa flavor, dairy note, whey protein, vanilla, metallic, sweetness, bitterness and aftertaste). The two samples included: 1) no sugar added “Control” sample containing 300 ppm PureCircle Reb A and 2) no sugar added “Test” sample containing 300 ppm PureCircle Reb A and 50 ppm PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of a milk product. The panel found the test sample containing 50 ppm of PCS-3028 to be significantly lower bitterness, metallic note, whey protein and lower bitter aftertaste than the control (at 95% confidence) and higher in cocoa flavor, dairy notes, vanilla flavor, and overall liking (at 95% confidence). Further, there was no significant impact on sweetness intensity. 
     Project Objective 
     The project objective is to assess if the addition of  stevia  extract solids has an effect on key flavor attributes in various beverage applications. 
     Test Objective 
     The test objective is to determine if the flavor profile and overall acceptance of a control sample of dairy beverage application differs from a Test sample of the same beverage containing PCS-3028. 
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity and acceptance ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
                   
               
            
           
           
               
            
               
                 Overall Acceptance on a 10-pt hedonic scale where 10 = Extremely 
               
               
                 Like and 0 = Extremely Dislike 
               
               
                 Overall liking, sweetness, bitterness, cocoa flavor, dairy notes, 
               
               
                 chocolate, whey protein notes, metallic note, vanilla note, and 
               
               
                 Aftertaste. 10-pt continuous intensity scale where 0 = 
               
               
                 Imperceptible and 10 = Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Refrigerated temperature (~45° F.) 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
            
               
                   
               
               
                 SAMPLES 
               
            
           
           
               
               
               
               
            
               
                   
                   
                 Sugar 
                 50 ppm 
               
               
                   
                 Ingredient list 
                 Reference 
                 PCS-3028 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                 Milk, 2% 
                 86.47 
                 86.465 
               
               
                   
                 Whey Protein 90 Instant - Non GMO 
                 6.8250 
                 6.8250 
               
               
                   
                 (Prod: 18618) 
               
               
                   
                 Non-Fat Dry Milk 
                 4.6269 
                 4.6269 
               
               
                   
                 Maltrin QD M585 
                 1.1066 
                 1.1066 
               
               
                   
                 Vitamin Blend - 
                 0.0063 
                 0.0063 
               
               
                   
                 Xanthan Gum (Cold dissolve) 
                 0.0359 
                 0.0359 
               
               
                   
                 Forbes 10/12 Cocoa powder 7113 
                 0.7194 
                 0.7194 
               
               
                   
                 Vanilla Flavor Powder 
                 0.1799 
                 0.1799 
               
               
                   
                 Reb A 
                 0.0300 
                 0.0300 
               
               
                   
                 PCS-3028 
                   
                 0.0050 
               
               
                   
                 TOTAL 
                 100 
                 100 
               
               
                   
                   
               
            
           
         
       
     
                                                 Sugar Contribution (grams)   Sugar   165 ppm           per 100 grams*   Reference   PCS-3028                                                        Milk, 2%   4.08   4.15           Non-Fat Dry Milk   2.41   2.41           Maltrin QD M585   0.08   0.08           TOTAL   8.07   6.64                       *Calculated with Genesis R&amp;D version 11.4            
Table 2 (below) summarizes the results.
 
     
       
         
           
               
             
               
                 TABLE 2 
               
             
            
               
                   
               
               
                 Effect of PCS PCS-3028 on flavor 
               
               
                 modification of Chocolate Protein shake 
               
               
                 Summary of Mean-Scores, P-Values, and Significance 
               
               
                 Test Result Code: PROTEIN6 
               
               
                 Test Description: Chocolate Vanilla 
               
               
                 Protein Dairy Shake: 500 ppm Reb AM 
               
               
                 This test was performed on 30 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 Test - NSA 
                   
                   
               
               
                   
                 Control - NSA 
                 Protein Shake 
               
               
                   
                 Protein Shake 
                 w Reb A &amp; 50 
               
               
                 Attribute 
                 w/Reb A 
                 ppm PCS-3028 
                 P-Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 6.04 
                 5.98 
                 0.7329 
                 NS 
               
               
                 Bitterness 
                 a 
                 b 
                 0.0138 
                 *** 
               
               
                   
                 1.98 
                 1.46 
               
               
                 Metallic Note 
                 a 
                 b 
                 0.0311 
                 *** 
               
               
                   
                 1.93 
                 1.48 
               
               
                 Cocoa Flavor 
                 b 
                 a 
                 0.0409 
                 *** 
               
               
                   
                 4.06 
                 4.55 
               
               
                 Dairy Note 
                 b 
                 a 
                 0.0515 
                 ** 
               
               
                   
                 4.10 
                 4.59 
               
               
                 Whey Protein Note 
                 a 
                 b 
                 0.0460 
                 *** 
               
               
                   
                 4.79 
                 4.32 
               
               
                 Vanilla Note 
                 b 
                 a 
                 0.0174 
                 *** 
               
               
                   
                 2.10 
                 2.52 
               
               
                 Sweet Aftertaste 
                 1.82 
                 1.65 
                 0.2130 
                 NS 
               
               
                 Bitter Aftertaste 
                 a 
                 b 
                 0.0495 
                 *** 
               
               
                   
                 1.03 
                 0.77 
               
               
                 Overall Liking 
                 b 
                 a 
                 0.0001 
                 *** 
               
               
                   
                 4.80 
                 5.59 
               
               
                   
               
               
                 *= 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             The panel found the test sample containing 50 ppm of PCS-3028 to be significantly lower bitterness, metallic note, whey protein and lower bitter aftertaste than the control (at 95% confidence). 
             The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher in cocoa flavor, dairy notes, vanilla flavor, and overall liking (at 95% confidence). 
           
         
       
    
     Example 3 Conclusion 
     Thirty company employees evaluated two samples of chocolate flavored dairy protein shake for overall acceptance and attribute intensities (cocoa flavor, dairy note, whey protein, vanilla, metallic, sweetness, bitterness and aftertaste). The two samples included: 1) no sugar added “Control” sample containing 300 ppm PureCircle Reb A and 2) no sugar added “Test” sample containing 300 ppm PureCircle Reb A and 50 ppm PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of a milk product. The panel found the test sample containing 50 ppm of PCS-3028 to be significantly lower bitterness, metallic note, whey protein and lower bitter aftertaste than the control (at 95% confidence) and higher in cocoa flavor, dairy notes, vanilla flavor, and overall liking (at 95% confidence). Further, there was no significant impact on sweetness intensity. 
     Example 4 
     Cinnamon Vanilla Oatmilk with PCS-3028 
     Application: Non-Dairy/Imitation Dairy Product 
     SUMMARY 
     Thirty company employees evaluated two samples of cinnamon vanilla oatmilk for overall acceptance and attribute intensities (cinnamon, vanilla, oat, sweetness, bitterness and aftertaste). The two samples included: 1) no sugar added “Control” sample containing 150 ppm PureCircle Reb A and 2) no sugar added “Test” sample containing 150 ppm PureCircle Reb A and 50 ppm PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of imitation dairy. The panel found the test sample containing 50 ppm of PCS-3028 to have significantly higher cinnamon, vanilla flavor and overall liking than the control (at 95% confidence). Further, there was no significant impact on sweetness intensity. 
     Project Objective 
     The project objective is to assess if the addition of  stevia  extract solids has an effect on key flavor attributes in various beverage applications. 
     Test Objective 
     The test objective is to determine if the flavor profile and overall acceptance of a control sample of imitation dairy beverage application differs from a Test sample of the same beverage containing PCS-3028. 
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity and acceptance ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
               
            
           
           
               
            
               
                 Overall Acceptance on a 10-pt hedonic scale where 
               
               
                 10 = Extremely Like and 0 = Extremely Dislike 
               
               
                 Overall Liking, sweetness, bitterness, vanilla, cinnamon, oat, and 
               
               
                 aftertaste. 10-pt continuous intensity scale where 0 = Imperceptible 
               
               
                 and 10 = Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Refrigerated temperature (~45° F.) 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
            
               
                   
               
               
                 SAMPLES 
               
            
           
           
               
               
               
            
               
                   
                 NSA Reb A 
                 Reb A + PCS3028 
               
               
                   
                   
               
            
           
           
               
               
               
            
               
                 Oatmilk Unsweetened plain 
                 99.735 
                 99.48 
               
               
                 Vanilla flavor 
                 0.15 
                 0.3 
               
               
                 Cinnamon flavor 
                 0.1 
                 0.2 
               
               
                 Reb A 97 
                 0.015 
                 0.015 
               
               
                 PCS-3028 
                   
                 0.005 
               
               
                 Total g 
                 100 
                 100 
               
               
                   
               
            
           
         
       
     
     Results 
     Table 3 (below) summarizes the results. 
     
       
         
           
               
             
               
                 TABLE 3 
               
             
            
               
                   
               
               
                 Effect of PCS-3028 on flavor modification of Vanilla Cinnamon Oatmilk 
               
               
                 Summary of Mean-Scores, P-Values, and Significance 
               
               
                 Test Description: Quaker Oatmilk with Vanilla and Cinnamon 
               
               
                 Flavor and Reb A 97 at 150 ppm with PCS3028 at 50 ppm 
               
               
                 This test was performed on 30 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 Vanilla Cinnamon 
                   
                   
               
               
                   
                   
                 Oatmilk with reb 
               
               
                   
                 Vanilla Cinnamon 
                 A 97 at 150 ppm 
               
               
                   
                 Oatmilk with Reb 
                 and PCS-3028 
                 P- 
               
               
                 Attribute 
                 A 97 at 150 ppm 
                 at 50 ppm 
                 Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 3.99 
                 4.12 
                 0.2662 
                 NS 
               
               
                 Bitter Intensity 
                 0.11 
                 0.14 
                 0.6024 
                 NS 
               
               
                 Cinnamon Flavor 
                 b 
                 a 
                 0.0001 
                 *** 
               
               
                   
                 2.19 
                 3.32 
               
               
                 Vanilla Flavor 
                 b 
                 a 
                 0.0038 
                 *** 
               
               
                   
                 3.07 
                 3.68 
               
               
                 Oat Flavor 
                 5.96 
                 5.75 
                 0.2865 
                 NS 
               
               
                 Sweet Aftertaste 
                 b 
                 a 
                 0.0940 
                 ** 
               
               
                   
                 0.95 
                 1.17 
               
               
                 Bitter Aftertaste 
                 0.21 
                 0.15 
                 0.5725 
                 NS 
               
               
                 Overall Liking 
                 b 
                 a 
                 0.0282 
                 *** 
               
               
                   
                 5.72 
                 6.34 
               
               
                   
               
               
                 *= 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher cinnamon flavor, vanilla flavor, and overall liking than the control (at 95% confidence). 
             The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher in sweet aftertaste (at 90% confidence). 
           
         
       
    
     Example 4 Conclusion 
     Thirty company employees evaluated two samples of cinnamon vanilla oatmilk for overall acceptance and attribute intensities (cinnamon, vanilla, oat, sweetness, bitterness and aftertaste). The two samples included: 1) no sugar added “Control” sample containing 150 ppm PureCircle Reb A and 2) no sugar added “Test” sample containing 150 ppm PureCircle Reb A and 50 ppm PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of imitation dairy. The panel found the test sample containing 50 ppm of PCS-3028 to have significantly higher cinnamon, vanilla flavor and overall liking than the control (at 95% confidence). Further, there was no significant impact on sweetness intensity. 
     Example 5 
     Strawberry Jam/Fruit Prep with PCS-3028 
     Application: Jams and Jellies 
     SUMMARY 
     Thirty company employees evaluated two samples of strawberry jam/fruit prep for overall acceptance and attribute intensities (strawberry, sour, sweetness, bitterness and aftertaste). The two samples included: 1) no sugar added “Control” sample containing 400 ppm PureCircle Reb A and 2) no sugar added “Test” sample containing 400 ppm PureCircle Reb A and 50 ppm PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of strawberry jam/fruit prep. The panel found the test sample containing 50 ppm of PCS-3028 to have significantly higher cooked strawberry flavor and overall liking than the control (at 95% confidence and 90% confidence respectively). Additionally, the product had significantly lower bitter aftertaste than the control (at 90% confidence). Further, there was no significant impact on sweetness intensity. 
     Project Objective 
     The project objective is to assess if the addition of  stevia  extract solids has an effect on key flavor attributes in various beverage applications. 
     Test Objective 
     The test objective is to determine if the flavor profile and overall acceptance of a control sample of jam and jelly application differs from a Test sample of the same beverage containing PCS-3028. 
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity and acceptance ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
               
            
           
           
               
            
               
                 Overall Acceptance on a 10-pt hedonic scale where 10 = Extremely 
               
               
                 Like and 0 = Extremely Dislike 
               
               
                 Overall Liking, sweetness, bitterness, strawberry, sour and aftertaste. 
               
               
                 10-pt continuous intensity scale where 0 = Imperceptible and 10 = 
               
               
                 Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Refrigerated temperature (~45° F.) 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     SAMPLES 
     50% Sugar Reduced Strawberry Fruit Prep 
       
                                     Ingredients   50% SR Reb A   50% SR Reb A + PCS3028                                            Water   41.7571   41.7571       Sodium Citrate   0.0453   0.0453       Potassium Sorbate   0.0453   0.0453       Sucrose   9.0634   0.0634       Starch   3.5   3.5       Reb A   0.04   0.04       Pureed Frozen Strawberries   45.317   45.3172       Red Food Coloring   0.2719   0.2719       PCS-3028       0.005       Total:   100   100                    
Table 4 (below) summarizes the results.
 
     
       
         
           
               
             
               
                 TABLE 4 
               
             
            
               
                   
               
               
                 Effect of PCS-3028 on flavor modification of Strawberry Fruit Prep/Jam 
               
               
                 Summary of Mean-Scores, P-Values, and Significance 
               
               
                 Test Description: Fruit Prep Strawberries 
               
               
                 Reb A 97 at 400 ppm with PCS-3028 as 50 ppm 
               
               
                 This test was performed on 30 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 Fruit Prep 
                   
                   
               
               
                   
                   
                 Strawberries 
               
               
                   
                 Fruit Prep 
                 with Reb A 97 
               
               
                   
                 Strawberries 
                 and PCS3028 
               
               
                 Attribute 
                 with Reb A 97 
                 at 50 ppm 
                 P-Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 7.83 
                 7.85 
                 0.9318 
                 NS 
               
               
                 Bitter intensity 
                 a 
                 b 
                 0.1771 
                 * 
               
               
                   
                 0.27 
                 0.13 
               
               
                 Fresh Strawberry 
                 5.08 
                 5.30 
                 0.4223 
                 NS 
               
               
                 Flavor 
               
               
                 Cooked Strawberry 
                 b 
                 a 
                 0.0166 
                 *** 
               
               
                 Flavor 
                 2.18 
                 2.89 
               
               
                 Sour Intensity 
                 2.03 
                 2.06 
                 0.7956 
                 NS 
               
               
                 Sweet Aftertaste 
                 1.70 
                 1.60 
                 0.5091 
                 NS 
               
               
                 Bitter Aftertaste 
                 a 
                 b 
                 0.0786 
                 ** 
               
               
                   
                 0.19 
                 0.05 
               
               
                 Overall Liking 
                 b 
                 a 
                 0.0870 
                 ** 
               
               
                   
                 5.96 
                 6.37 
               
               
                   
               
               
                 * = 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             The panel found the test sample containing 50 ppm of PCS-3028 to be significantly higher cooked strawberry flavor, and overall liking than the control (at 95% confidence and 90% confidence respectively). 
             The panel found the test sample containing 50 ppm of PCS-3028 to be significantly lower in bitter aftertaste (at 90% confidence). 
           
         
       
    
     Example 5 Conclusion 
     Thirty company employees evaluated two samples of strawberry jam/fruit prep for overall acceptance and attribute intensities (strawberry, sour, sweetness, bitterness and aftertaste). The two samples included: 1) no sugar added “Control” sample containing 400 ppm PureCircle Reb A and 2) no sugar added “Test” sample containing 400 ppm PureCircle Reb A and 50 ppm PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of strawberry jam/fruit prep. The panel found the test sample containing 50 ppm of PCS-3028 to have significantly higher cooked strawberry flavor and overall liking than the control (at 95% confidence and 90% confidence respectively). Additionally, the product had significantly lower bitter aftertaste than the control (at 90% confidence). Further, there was no significant impact on sweetness intensity. 
     Example 6a 
     Sweetness Perception Threshold with PCS-3028 
     Application: Baked Goods 
     SUMMARY 
     The sweetness perception of 4.0% sugar solution and different solutions of PCS3028 were tested with a consumer panel and found that 150 ppm of PCS3028 in Lemon Poppy Seed Muffin provided sweetness perception significantly less than that of 4.0% sugar solution in a Lemon Poppy Seed Muffin. Additional threshold test data found for Lemon Poppy Seed Muffin provided in Appendix. 
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 ≥30 
               
               
                 Test Design: 
                 2- AFC, Balanced, randomized within pair. 
               
               
                   
                 Blind 
               
               
                 Sensory Test Method: 
                 Intensity ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
                 Which sample is sweeter? 
               
               
                 Statistical Analysis: 
                 Paired comparison Test 
               
               
                 Sample Size 
                 ~2.0 oz. in a clear plastic cup 
               
               
                 Serving Temperature 
                 Room temperature (~70° F.) 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
            
               
                   
               
               
                 DATA: n = 30 
               
            
           
           
               
               
            
               
                 Two-Tailed Analysis 
                   
               
               
                 Table Report for Result 
                   
               
            
           
           
               
               
               
            
               
                   
                 70 ppm 
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                 4% sugar 
                 PCS3028 
                 Percent 
                 Binomial Distribution 
               
               
                   
                 in muffins 
                 in muffins 
                 Frequency 
                 Probability 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Sample 1 
                 Sample 2 
                 Sample 1 
                 P-value 
                 Sig 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 PC 
                 23 
                 7 
                 76.7% 
                 0.0052 
                 *** 
               
               
                 Frequency 
                 76.7% 
                 23.3% 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
            
               
                   
               
               
                 DATA: n = 30 
               
            
           
           
               
               
            
               
                 Two-Tailed Analysis 
                   
               
               
                 Table Report for Result 
                   
               
            
           
           
               
               
               
            
               
                   
                 150 ppm 
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                 4% sugar 
                 PCS3028 
                 Percent 
                 Binomial Distribution 
               
               
                   
                 in muffins 
                 in muffins 
                 Frequency 
                 Probability 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Sample 1 
                 Sample 2 
                 Sample 1 
                 P-value 
                 Sig 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 PC 
                 21 
                 9 
                 70% 
                 0.0428 
                 *** 
               
               
                 Frequency 
                 70% 
                 30% 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
            
               
                   
               
               
                 DATA: n = 30 
               
            
           
           
               
               
            
               
                 Two-Tailed Analysis 
                   
               
               
                 Table Report for Result 
                   
               
            
           
           
               
               
               
            
               
                   
                 250 ppm 
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                 4% sugar 
                 PCS3028 
                 Percent 
                 Binomial Distribution 
               
               
                   
                 in muffins 
                 in muffins 
                 Frequency 
                 Probability 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Sample 1 
                 Sample 2 
                 Sample 1 
                 P-value 
                 Sig 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 PC 
                 10 
                 20 
                 33.3% 
                 0.0987 
                 ** 
               
               
                 Frequency 
                 33.3% 
                 66.7% 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                   
               
               
                 Ingredient list 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
            
               
                   
                   
                 SUGAR 
                 PCS-3028 
               
               
                 Dry Ingredients 
                 BASE 
                 Reference 
                 150 ppm 
               
               
                   
               
               
                 All Purpose Flour 
                 17.6600 
                 17.6600 
                 17.7099 
               
               
                 Whole Wheat Flour 
                 5.8818 
                 5.8818 
                 5.8818 
               
               
                 Poppy Seeds 
                 1.0658 
                 1.0658 
                 1.0658 
               
               
                 Maltodextrin - 10DE 
                 8.3307 
                 4.3307 
                 8.2807 
               
               
                 Fibersol2 
                 4.1618 
                 4.1618 
                 4.1618 
               
               
                 Modified Starch - Inscosity 656 
                 4.1618 
                 4.1618 
                 4.1618 
               
               
                 (Grain Products) 
               
               
                 Lemon Flavor 
                 0.8868 
                 0.8868 
                 0.8868 
               
               
                 Salt (Sodium Chloride) 
                 0.7486 
                 0.7486 
                 0.7486 
               
               
                 Baking Powder 
                 1.0658 
                 1.0658 
                 1.0658 
               
               
                 Baking Soda 
                 0.3208 
                 0.3208 
                 0.3208 
               
               
                 Sucrose 
                   
                 4.0000 
               
               
                 PCS-3028 
                   
                   
                 0.0150 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                   
                 Sugar 
                 PCS-3028 
               
               
                   
                 Wet Ingredients 
                 BASE 
                 Reference 
                 150 ppm 
               
               
                   
                   
               
               
                   
                 Milk, 2% 
                 27.2700 
                 27.2700 
                 27.2700 
               
               
                   
                 Soybean Oil 
                 11.7635 
                 11.7635 
                 11.7635 
               
               
                   
                 Whole Eggs 
                 8.5553 
                 8.5553 
                 8.5553 
               
               
                   
                 Water 
                 5.3470 
                 5.3470 
                 5.3470 
               
               
                   
                 Yogurt, Plain Nonfat 
                 1.6041 
                 1.6041 
                 1.6041 
               
               
                   
                 Lemon Juice, 100% 
                 0.6416 
                 0.6416 
                 0.6416 
               
               
                   
                 Vanilla Extract 
                 0.5347 
                 0.5347 
                 0.5347 
               
               
                   
                   
                 100 
                 100 
                 100 
               
               
                   
                   
               
            
           
         
       
     
     Example 6b 
     Lemon Poppy Seed Muffin with PCS-3028 
     Application: Baked Goods 
     SUMMARY 
     To test the contribution of PCS-3028 in baked goods, lemon flavored poppy seed muffins were baked with a 50% sugar reduced formulation with Reb A as control, and sugar reduced formulation with Reb A and  stevia  extract (PCS-3028) as a test sample. A thirty consumer panel members evaluated two samples of lemon poppy seed muffins for different attributes (lemon flavor, vanilla flavor, brown notes, and sweet). The panel found that the test samples with PCS-3028 significantly higher lemon flavor, and brown flavor intensity than the control sample (at 95% confidence). 
     Project Objective 
     The project objective is to assess if the addition of PCS-3028 has an effect on key flavor attributes in baked applications. 
     Test Objective 
     The test objective is to determine if the flavor profile of a control sample of sugar reduced lemon poppy seed muffin differs from Test samples of the same muffin containing PCS-3028. 
     Sample Preparation 
     
         
         
           
             1. Grease muffin pans or line with muffin baking cups. Preheat oven to 350° F. (175° C.). 
             2. In a large mixing bowl, blend dry ingredients. Make a well in the center of the bowl. 
             3. Add wet ingredients and mix until smooth. 
             4. Pour batter evenly into prepared pans. Bake for 25-30 minutes, or until a toothpick inserted 
             into the center of the muffins comes out clean. 
             5. Cool in muffins in baking pan for 5 minutes then remove to a wire cooling rack. 
             6. Serve muffins at room temperature. 
           
         
       
    
     
       
         
           
               
               
               
             
               
                   
                   
               
               
                   
                 Reb A 
                 Reb A w/ 
               
               
                   
                 Control 
                 PCS-3028 
               
               
                   
                   
               
             
            
               
                   
               
            
           
           
               
               
               
            
               
                 Dry Ingredients 
                   
                   
               
               
                 Sucrose 
                 12.3722 
                 12.3722 
               
               
                 All Purpose Flour 
                 17.6434 
                 17.6434 
               
               
                 Whole Wheat Flour 
                 5.8763 
                 5.8763 
               
               
                 Poppy Seeds 
                 1.0648 
                 1.0648 
               
               
                 Maltodextrin - 10DE (Tate &amp; Lyle) 
                 2.1368 
                 2.1368 
               
               
                 Fibersol2 (ADM/Matsutani) 
                 1.0648 
                 1.0648 
               
               
                 Modified Starch - Inscosity 656 (Grain Products) 
                 1.0648 
                 1.0648 
               
               
                 Lemon Flavor - Firmenich 
                 0.8860 
                 0.8860 
               
               
                 Salt (Sodium Chloride) 
                 0.7479 
                 0.7479 
               
               
                 Baking Powder 
                 1.0648 
                 1.0648 
               
               
                 Baking Soda 
                 0.3205 
                 0.3205 
               
               
                 Reb A 
                 0.0200 
                 0.0200 
               
               
                 PCS-3028 
                   
                 0.0150 
               
               
                 Wet Ingredients 
               
               
                 Milk, 2% 
                 27.2444 
                 27.2444 
               
               
                 Soybean Oil 
                 11.7525 
                 11.7525 
               
               
                 Whole Eggs 
                 8.5473 
                 8.5473 
               
               
                 Water 
                 5.3420 
                 5.3420 
               
               
                 Yogurt, Plain Nonfat 
                 1.6026 
                 1.6026 
               
               
                 Lemon Juice, 100% 
                 0.6410 
                 0.6410 
               
               
                 Vanilla Extract 
                 0.5342 
                 0.5342 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
               
            
           
           
               
            
               
                 Lemon Flavor, Vanila Flavor, Brown Note, and Sweetness, aftertaste 
               
               
                 (sweet and bitter) 10-pt continuous intensity scale where 0 = 
               
               
                 Imperceptible and 10 = Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Room temperature (~70° F.) 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     Results 
     Table 5 (below) summarizes the overall acceptance and mean attribute intensity results for each sample. 
     
       
         
           
               
             
               
                 TABLE 5 
               
             
            
               
                   
               
               
                 Mean Scores and degree of confidence (P value) 
               
               
                 Summary of Mean-Scores, P-Values, and Significance 
               
               
                 Test Description: 50% Sugar Reduced Lemon Poppy Seed 
               
               
                 Muffins with 200 ppm of Reb A vs. 150 ppm of PCS-3028 
               
               
                 This test was performed on 30 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 50% SR Muffins 
                   
                   
               
               
                   
                 50% SR Muffins 
                 with 200 ppm 
               
               
                   
                 with 200 ppm 
                 of Reb A and 150 
                 P- 
               
               
                 Attribute 
                 of Reb A 
                 ppm PCS-3028 
                 Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 b 
                 a 
                 0.1808 
                 * 
               
               
                   
                 7.56 
                 7.80 
               
               
                 Bitter Intensity 
                 0.85 
                 0.76 
                 0.2829 
                 NS 
               
               
                 Lemon Flavor 
                 b 
                 a 
                 0.0202 
                 *** 
               
               
                   
                 3.70 
                 4.21 
               
               
                 Brown Note 
                 b 
                 a 
                 0.0147 
                 *** 
               
               
                   
                 1.99 
                 2.45 
               
               
                 Vanilla Flavor 
                 1.22 
                 1.41 
                 0.3233 
                 NS 
               
               
                 Sweet Aftertaste 
                 2.38 
                 2.29 
                 0.5064 
                 NS 
               
               
                 Bitter Aftertaste 
                 0.45 
                 0.49 
                 0.6803 
                 NS 
               
               
                 Overall Liking 
                 b 
                 a 
                 0.1107 
                 * 
               
               
                   
                 6.39 
                 6.56 
               
               
                   
               
               
                 * = 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             The Test samples with PCS-3028 had significantly higher lemon flavor, and brown flavor intensity than the Control sample (95% confidence). 
           
         
       
    
     Example 6 Conclusion 
     To test the contribution of PCS-3028 in baked goods, lemon flavored poppy seed muffins were baked with a sugar reduced formulation with Reb A as control, and sugar reduced formulation with Reb A and  stevia  extract (PCS-3028) as a test sample. A thirty consumer panel members evaluated two samples of lemon poppy seed muffins for different attributes (lemon flavor, vanilla flavor, brown notes, and sweet). The panel found that the Test samples with PCS-3028 had significantly higher lemon flavor, and brown flavor intensity than the control sample (at 95% confidence). 
     Example 7 
     Barbeque Flavored Peanut with PCS-3028 
     Application: Snack Food/Seasoning/Nut Product 
     SUMMARY 
     Roasted Peanuts were used for testing the modification of flavor profile by the PCS-3028. The two samples included: 1) a 50% sugar reduced control sample containing Reb A, 2) a 50% reduced sugar test sample containing Reb A and PCS-3028.
 
Thirty consumer panel members evaluated two samples of the peanuts for overall acceptance and attribute intensities (overall flavor, saltiness, sweetness, smoke flavor, spice/heat intensity, peanut flavor, chili powder flavor, bitterness and lingering sweet aftertaste intensity). The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of a savory snack food. The results indicated:
         Test sample had significantly less bitterness and lower aftertaste (sweet and bitter) than the Control sample (at 90% and 95% confidence respectively).       

     Project Objective 
     The project objective is to assess if the addition of  stevia  extract solids has an effect on key flavor attributes in various applications. 
     Test Objective 
     The test objective is to determine if the flavor profile and overall acceptance of a Control sample of barbeque flavored peanuts (containing no  stevia  extract solids) differs from a Test sample of the same snack food (containing  stevia  extract solids). 
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity and acceptance ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
               
            
           
           
               
            
               
                 Overall Acceptance on a 10-pt hedonic scale where 10 + Extremely 
               
               
                 Like and 0 = Extremely Dislike 
               
               
                 Overall Flavor, Saltiness, Sweetness, Smoke Intensity, Heat/spice 
               
               
                 intensity, peanut flavor, chili powder and Aftertaste Intensity 
               
               
                 (sweet and bitter) on a 10-pt continuous intensity scale where 
               
               
                 0 = Imperceptible and 10 = Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Room temperature 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     SAMPLE PREPARATION 
     
         
         
           
             Mix peanuts with liquid ingredients, until peanuts are completely coated. 
             Blend dry ingredients with Reb A and the test ingredient (PCS 5001). 
             Add to the peanuts and mix well until they are fully coated. 
             Serve at room temperature in 1 oz cups. 
           
         
       
    
     
       
         
           
               
            
               
                   
               
               
                 SAMPLES 
               
            
           
           
               
               
               
            
               
                   
                 Reb A 
                 Reb A + PCS3028 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                 Unsalted Peanuts 
                 86.812 
                 86.7277 
               
               
                   
                 Vegetable oil 
                 2.934 
                 2.934 
               
               
                   
                 Sugar 
                 5.876 
                 5.876 
               
               
                   
                 Salt 
                 2.934 
                 2.934 
               
               
                   
                 Chilli powder 
                 0.173 
                 0.173 
               
               
                   
                 Cumin powder 
                 0.286 
                 0.286 
               
               
                   
                 Garlic powder 
                 0.156 
                 0.156 
               
               
                   
                 Cayenne pepper 
                 0.156 
                 0.156 
               
               
                   
                 Smoke liquid 
                 0.729 
                 0.729 
               
               
                   
                 Reb A 
                 0.0243 
                 0.0243 
               
               
                   
                 PCS-3028 
                   
                 0.0040 
               
               
                   
                 Total wt. (g) 
                 100 
                 100 
               
               
                   
                   
               
            
           
         
       
     
     Results 
     Table 6 (below) summarizes the overall acceptance and mean attribute intensity results for each sample. 
     
       
         
           
               
             
               
                 TABLE 6 
               
             
            
               
                   
               
               
                 Mean Scores 
               
               
                 Summary of Mean-Scores, P-Vaules, and Significance 
               
               
                 Test Description: 50% SR BBQ Peanuts w/PCS-3028 at 40 ppm 
               
               
                 This test was performed on 31 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 50% SR w/PCS- 
                   
                   
               
               
                   
                 50% SR control 
                 3028 at 40 ppm 
               
               
                 Attribute 
                 BBQ peanuts 
                 BBQ peanuts 
                 P-Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 3.95 
                 4.09 
                 0.6444 
                 NS 
               
               
                 Saltiness 
                 5.23 
                 5.10 
                 0.6366 
                 NS 
               
               
                 Smoked Flavor 
                 2.30 
                 2.48 
                 0.4667 
                 NS 
               
               
                 Heat/Spice 
                 1.57 
                 1.41 
                 0.4026 
                 NS 
               
               
                 Peanut Flavor 
                 7.61 
                 7.74 
                 0.2933 
                 NS 
               
               
                 Chilli Powder 
                 3.12 
                 3.19 
                 0.6781 
                 NS 
               
               
                 Bitterness 
                 a 
                 b 
                 0.0719 
                 ** 
               
               
                   
                 1.07 
                 0.85 
               
               
                 Bitter Aftertaste 
                 a 
                 b 
                 0.0291 
                 *** 
               
               
                   
                 0.86 
                 0.60 
               
               
                 Sweet Aftertaste 
                 a 
                 b 
                 0.0395 
                 *** 
               
               
                   
                 1.85 
                 1.45 
               
               
                 Overall Liking 
                 b 
                 a 
                 0.1986 
                 * 
               
               
                   
                 5.66 
                 4.96 
               
               
                   
               
               
                 * = 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             Test sample had significantly less bitterness and lower aftertaste (sweet and bitter) than the Control sample (at 90% and 95% confidence respectively). 
           
         
       
    
     Example 7 Conclusion 
     Thirty consumer panel members evaluated two samples of the peanuts for overall acceptance and attribute intensities (overall flavor, saltiness, sweetness, smoke flavor, spice/heat intensity, peanut flavor, chili powder flavor, bitterness and lingering sweet aftertaste intensity). The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of a savory snack food. The results indicated Test sample had significantly less bitterness and lower aftertaste (sweet and bitter) than the Control sample (at 90% and 95% confidence respectively). 
     Example 8 
     Alcoholic Beverage—Moscow Mule with PCS-3028 
     Application: Alcoholic Beverage 
     SUMMARY 
     Thirty company employees evaluated two samples of alcoholic beverage for overall acceptance and attribute intensities. The two samples included: 1) Moscow mule—Control and 2) Moscow Mule test sample containing PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of an alcoholic beverage. The results indicated the Test sample PCS-3028 had significantly higher ginger and alcohol note compared to the control samples (at 95% confidence). 
     Project Objective 
     The project objective is to assess if the addition of  stevia  extract solids has an effect on key flavor attributes in alcoholic beverage applications. 
     Test Objective 
     The test objective is to determine if the flavor profile and overall acceptance of a Control sample of alcoholic beverage differs from a Test sample of the same beverage containing PCS-3028. 
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity and acceptance ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
               
            
           
           
               
            
               
                 Overall Acceptance on a 10-pt hedonic scale where 10 + Extremely 
               
               
                 Like and 0 = Extremely Dislike 
               
               
                 Overall Liking, Sweetness, Bitterness, Ginger, Llime, Alcohol, and 
               
               
                 Aftertaste (Sweet, bitter, and Alcohol). 10-pt continuous intensity 
               
               
                 scale where 0 = Imperceptible and 10 = Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Refrigerated temperature (~45° F.) 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     Samples 
       
     
       
         
           
               
            
               
                   
               
               
                 Ingredient list 
               
            
           
           
               
               
               
            
               
                   
                 Moscow 
                 50 ppm in 
               
               
                   
                 Mule 
                 Moscow Mule 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                 Mule 20, Moscow Mule 
                 100 
                 99.995 
               
               
                   
                 PCS-3028 
                   
                 0.0050 
               
               
                   
                 TOTAL 
                 100 
                 100 
               
               
                   
                   
               
               
                   
                 *Mule 20, Moscow Mule, 8% alcohol/volume 
               
            
           
         
       
     
     Results 
     Table 7 (below) summarizes the overall acceptance and mean attribute intensity results for each sample. 
     
       
         
           
               
             
               
                 TABLE 7 
               
             
            
               
                   
               
               
                 Mean Scores 
               
               
                 Summary of Mean-Scores, P-Values, and Significance 
               
               
                 Test Description: Moscow Mule Pre-Mixed 8% 
               
               
                 The test was performed on 34 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 Moscow Mule 
                   
                   
               
               
                   
                 Moscow Mule 
                 Pre-Mixed 8% 
               
               
                   
                 Pre-Mixed 8% 
                 Alcohol w/ 
               
               
                   
                 Alcohol 
                 PCS3028 @ 
                 P- 
               
               
                 Attribute 
                 Control 
                 50 ppm 
                 Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 b 
                 a 
                 0.1052 
                 * 
               
               
                   
                 5.78 
                 6.13 
               
               
                 Bitted Intensity 
                 4.08 
                 4.00 
                 0.6238 
                 NS 
               
               
                 Ginger Flavor 
                 b 
                 a 
                 0.0173 
                 *** 
               
               
                   
                 2.97 
                 3.61 
               
               
                 Lime Flavor 
                 1.21 
                 1.15 
                 0.6304 
                 NS 
               
               
                 Alcohol Note 
                 b 
                 a 
                 0.0438 
                 *** 
               
               
                   
                 6.41 
                 7.06 
               
               
                 Sweet Aftertaste 
                 2.46 
                 2.34 
                 0.3686 
                 NS 
               
               
                 Bitter Aftertaste 
                 1.86 
                 1.86 
                 1.0000 
                 NS 
               
               
                 Alcohol Aftertaste 
                 3.00 
                 2.85 
                 0.5491 
                 NS 
               
               
                 Overall Liking 
                 5.40 
                 5.68 
                 0.2230 
                 NS 
               
               
                   
               
               
                 * = 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             The results indicated the Test sample PCS-3028 had significantly higher ginger flavor, and alcohol note compared to the control samples (at 95% confidence). 
           
         
       
    
     Example 8 Conclusion 
     Thirty company employees evaluated two samples of alcoholic beverage for overall acceptance and attribute intensities. The two samples included: 1) Moscow mule—Control and 2) Moscow Mule test sample containing PCS-3028. The objective of the test was to determine if the addition of PCS-3028 affects the flavor profile of an alcoholic beverage. The results indicated the Test sample PCS-3028 had significantly higher ginger and alcohol note compared to the control samples (at 95% confidence). 
     Example 9 
     Application: Breakfast Cereal 
     To estimate the sweetness threshold perception of PCS-3028 in breakfast cereal, puffed corn cereal was made and sweetened with 3.0% sugar (control) or with PCS-3028 (test sample). As recommended by the FEMA Sensory guidance document,—Guidance for the Sensory Testing of Flavorings with Modifying Properties, within the FEMA GRAS™ Program, 2013—the recognition threshold concentration was determined using 2-alternateve forced choice (2-AFC) methodology.
 
A consumer panel (N=30) tested control (3.0% sugar) and test samples containing 150 ppm PCS-3028 in a puffed corn cereal and found that control sample was significantly sweeter than test samples containing 150 ppm PCS-3028.
 
Therefore, it is proposed that the average maximum level be 150 ppm of PCS-3028 for the breakfast cereal product category.
 
     Example 9a 
     Sweetness Perception Threshold of PCS-3028 in Breakfast Cereal 
     Application: Breakfast Cereal 
     SUMMARY 
     The sweetness perception of 3.0% sugar and 150 ppms of PCS-3028 added a low sugar cereal was tested with a consumer panel and found that 150 ppm of PCS-3028 added to a low sugar cereal provided sweetness perception that was significantly less sweet than 3.0% sugar added to the low sugar cereal. 
                             METHODOLOGY                                        Nature of Participants:   Company employees       Number of Sessions   1       Number of Participants:   ≥30       Test Design:   2- AFC, Balanced, randomized within pair.           Blind       Sensory Test Method:   Intensity ratings       Environmental Condition   Standard booth lighting       Attributes and Scales:   Which sample is sweeter?       Statistical Analysis:   Paired comparison Test       Sample Size   ~1.5 oz. in a clear capped plastic cup       Serving Temperature   Room Temperature       Serving/Panelists   Samples served simultaneously. Panelists       Instruction:   instructed to read ingredient statement,           evaluate each sample.                    
This is an evaluation of the recognition threshold concentration to follow methodology outlined in section 1.4.2 of the “Guidance for the Sensory Testing of Flavorings with Modifying Properties within the FEMA GRAS™ Program”.
 
     
       
         
           
               
            
               
                   
               
               
                 DATA: n = 30 
               
               
                 Two-Tailed Analysis Table Report for Result 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Puffed Corn 
                 Puffed Corn 
                 Percent 
                   
                   
               
               
                   
                 Cereal + 8% 
                 Cereal + 150 
                 Frequency 
               
               
                   
                 Sugar 
                 ppm PCS-3028 
                 Sample 1 
                 P-value 
                 Sig 
               
               
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 PC 
                 28 
                 2 
                 93.3% 
                 0.00 
                 *** 
               
               
                 % Frequency 
                 93.3 
                 6.7 
               
            
           
           
               
            
               
                 Analysis: 
               
               
                   
               
            
           
         
       
     
                            DATA: n = 30       Two-Tailed Analysis Table Report for Result                                         Puffed Corn   Puffed Corn   Percent                   Cereal + 8%   Cereal + 250   Frequency           Sugar   ppm PCS-3028   Sample 1   P-value   Sig                                                 PC   13   17   43.3   0.5847           % Frequency   43.3   56.7                 Analysis:                    
Sample Preparation: The puffed corn cereal (with no added sugar or flavor) purchased from local stores and sprayed with aqueous sucrose solutions using a portable atomizer to create a fine mist while tumbling the dry cereal grains. All the samples were dried to remove the added water from the samples. About 3 g of cereal was portioned into appropriately labeled blind-coded plastic 2 oz soufflé cup with lids. Samples were served dry to all panelists.
 
     Example 9b 
     Breakfast Cereal with PCS-3028 
     Application: Breakfast Cereal 
     SUMMARY 
     To evaluate the contribution of PCS-3028 in breakfast cereal, two vanilla flavored corn puff cereal samples were prepared and tested by a consumer panel of thirty company employees. The employee panel evaluated the samples of breakfast cereal for overall acceptance and attribute intensities (corn, vanilla, brown, sweetness, bitterness, and aftertaste). The two samples included: 1) Control of vanilla flavored corn puffs and 2) Test sample of vanilla flavored corn puffs with 150 ppm PCS-3028. The panel found the test sample containing 150 ppm of PCS-3028 to be significantly higher in corn puffs, vanilla, brown, overall liking than the control (at 95% confidence). Additionally, results showed significantly higher sweet aftertaste (at 90% confidence). 
     Project Objective 
     The project objective is to assess if the addition of  stevia  extract solids has an effect on key flavor attributes in various food applications. 
     Test Objective 
     The test objective is to determine if the flavor profile of a control sample of breakfast cereal differs from a Test sample of the same cereal containing PCS-3028. 
     Samples 
     Breakfast Cereal 
       
     
       
         
           
               
            
               
                   
               
               
                 Corn Puff Caramel Flavored 
               
            
           
           
               
               
               
            
               
                   
                 Reference 
                 PCS-3028 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                 *Corn Puff Dry Cereal 
                 93.9 
                 93.885 
               
               
                   
                 Sugar 
                 6 
                 6 
               
               
                   
                 Oil 
                 0.1 
                 0.1 
               
               
                   
                 PCS-3028 
                   
                 0.015 
               
               
                   
                 Total (g) 
                 100 
                 100 
               
               
                   
                   
               
               
                   
                 *Arrowhead Mills brand 
               
               
                   
                 Bake at 200 F., 8 minutes 
               
            
           
         
       
     
     
       
         
           
               
               
             
               
                   
                   
               
               
                   
                 Sugar slurry 
               
               
                   
                   
               
             
            
               
                   
               
            
           
           
               
               
               
            
               
                   
                 water 
                 72.8 
               
               
                   
                 sugar 
                 120 
               
               
                   
                 oil 
                 0.2 
               
               
                   
                 Vanilla Flavor 
                 7 
               
               
                   
                 Total g 
                 200 
               
               
                   
                   
               
               
                   
                 *Use 10 g of slurry to make 100 gr of final product. 
               
            
           
         
       
     
     
       
         
           
               
             
               
                   
               
               
                 METHODOLOGY 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                 Nature of Participants: 
                 Company employees 
               
               
                 Number of Sessions 
                 1 
               
               
                 Number of Participants: 
                 30 
               
               
                 Test Design: 
                 Balanced, randomized within pair. Blind 
               
               
                 Sensory Test Method: 
                 Intensity and acceptance ratings 
               
               
                 Environmental Condition 
                 Standard booth lighting 
               
               
                 Attributes and Scales: 
               
            
           
           
               
            
               
                 Overall Acceptance on a 10-pt hedonic scale where 10 + Extremely 
               
               
                 Like and 0 = Extremely Dislike 
               
               
                 Overall Liking, Corn, brown note, vanilla flavor, sweetness, bitterness, 
               
               
                 and aftertaste. 10-pt continuous intensity scale where 0 = Imperceptible 
               
               
                 and 10 = Extremely Pronounced 
               
            
           
           
               
               
            
               
                 Statistical Analysis: 
                 ANOVA (by Block) with Post Hoc Duncan&#39;s 
               
               
                   
                 Test 
               
               
                 Sample Size 
                 ~1.5 oz. in a clear capped plastic cup 
               
               
                 Serving Temperature 
                 Room temperature 
               
               
                 Serving/Panelists 
                 Samples served simultaneously. Panelists 
               
               
                 Instruction: 
                 instructed to read ingredient statement, 
               
               
                   
                 evaluate each sample. 
               
               
                   
               
            
           
         
       
     
     Results 
     Table 8 (below) summarizes the overall acceptance and mean attribute intensity results for each sample. 
     
       
         
           
               
             
               
                 TABLE 8 
               
             
            
               
                   
               
               
                 Mean Scores 
               
               
                 Summary of Mean-Scores, P-Values, and Significance 
               
               
                 Test Description: Vanilla Corn puffs cereal with 
               
               
                 6% Sucrose vs. 6% Sucrose and PCS-3028 at 150 ppm 
               
               
                 This test was performed on 30 panelists. 
               
            
           
           
               
               
               
               
               
            
               
                   
                 6% Sucrose 
                 6% Sucrose with 150 
                   
                   
               
               
                 Attribute 
                 Only 
                 ppm of PCS-3028 
                 P-Value 
                 Sig 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Sweet Intensity 
                 2.16 
                 2.14 
                 0.9321 
                 NS 
               
               
                 Bitted Intensity 
                 0.19 
                 0.16 
                 0.5349 
                 NS 
               
               
                 Vanilla Flavor 
                 b 
                 a 
                 0.0168 
                 *** 
               
               
                   
                 2.32 
                 3.31 
               
               
                 Brown Note 
                 b 
                 a 
                 0.0131 
                 *** 
               
               
                   
                 3.57 
                 4.50 
               
               
                 Corn Flavor 
                 b 
                 a 
                 0.0236 
                 *** 
               
               
                   
                 6.04 
                 7.00 
               
               
                 Sweet Aftertaste 
                 b 
                 a 
                 0.0662 
                 ** 
               
               
                   
                 0.92 
                 1.22 
               
               
                 Bitter Aftertaste 
                 0.47 
                 0.38 
                 0.2726 
                 NS 
               
               
                 Overall Liking 
                 b 
                 a 
                 0.0045 
                 *** 
               
               
                   
                 4.81 
                 5.56 
               
               
                   
               
               
                 *= 80% CI, 
               
               
                 ** = 90% CI, 
               
               
                 *** = 95% CI 
               
            
           
         
       
         
         
           
             The panel found the test sample containing 150 ppm of PCS-3028 to be significantly higher in corn, vanilla, brown, and overall liking than the control (at 95% confidence). 
             Additionally, results showed significantly higher sweet aftertaste (at 90% confidence). 
           
         
       
    
     Example 9 Conclusion 
     To evaluate the contribution of PCS-3028 in breakfast cereal, two vanilla flavored corn puff cereal samples were prepared and tested by a consumer panel of thirty company employees. The employee panel evaluated the samples of breakfast cereal for overall acceptance and attribute intensities (corn, vanilla, brown, sweetness, bitterness, and aftertaste). The two samples included: 1) Control of vanilla flavored corn puffs and 2) Test sample of vanilla flavored corn puffs with 150 ppm PCS-3028. The panel found the test sample containing 150 ppm of PCS-3028 to be significantly higher in corn puffs, vanilla, brown, overall liking than the control (at 95% confidence). Additionally, results showed significantly higher sweet aftertaste (at 90% confidence). 
     Although the invention and its advantages have been described in detail, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims. Moreover, the scope of the application is not intended to be limited to the particular embodiments of the invention described in the specification. As one of ordinary skill in the art will readily appreciate from the disclosure of the invention, the compositions, processes, methods, and steps, presently existing or later to be developed that perform substantially the same function or achieve substantially the same result as the corresponding embodiments described herein may be utilized according to the invention.