Stevia extract containing selected steviol glycosides as flavor, salty and sweetness profile modifier

Abstract

Stevia extracts with selected major steviol glycosides (Reb A, stevioside, Reb D, Reb C) and minor steviol glycosides and glycosylated diterpene derivative plant molecules, derived from Stevia rebaudiana plant are found to improve the perception of flavor and taste perception, which includes the sweet, savory and salty perception in a wide range of food and beverage applications.

Description

FIELD OF THE INVENTION

The invention relates to the use of Stevia extracts as flavor modifiers that contain mixtures of steviol glycosides extracted from Stevia rebaudiana plant. This invention also relates to the application of the above-said Stevia extracts as sweetness profile modifier, not a sweetener, with other natural and artificial sweeteners. This invention also relates to the production and use of the above-mentioned Stevia extracts that can be used as flavor and sweetness profile modifier when used in food, beverage, and pharmaceutical products.

BACKGROUND

High intensity sweeteners possess sweetness level many times exceeding that of sucrose. They are essentially non-caloric and used widely in manufacturing of diet and reduced calorie food. Although natural caloric sweetener such as sucrose, fructose, and glucose provide the most desirable taste to consumers, they are caloric. High intensity sweeteners do not affect the blood glucose level and provide little or no nutritive value.

However, high intensity sweeteners that generally are used as substitutes for sucrose possess taste characteristics different than that of sugar, such as sweet taste with different temporal profile, maximal response, flavor profile, mouthfeel, and/or adaptation behavior than that of sugar. For example, the sweet taste of some high-potency sweeteners is slower in onset and longer in duration than that of sugar and thus changes the taste balance of a food composition. Because of these differences, usage of high-potency sweetener in replacing such a bulk sweetener as sugar in a food or beverage causes imbalance in temporal and/or flavor profile. If the taste profile of high-potency sweeteners could be modified to impart desired taste characteristics, it can provide low calorie beverages and food products with taste characteristics more desirable for consumers. To attain the sugar-like temporal and/or flavor profile, several ingredients have been suggested in different publications.

Non-limiting examples of synthetic sweeteners include sucralose, potassium acesulfame, aspartame, alitame, saccharin, neohesperidin dihydrochalcone synthetic derivatives, cyclamate, neotame, dulcin, suosan, N—[N-[3-(3-hydroxy-4-methoxyphenyl)propyl]-L-α-aspartyl]-L-phenylalanine 1-methyl ester, N—[N-[3-(3-hydroxy-4-methoxyphenyl)-3-methylbutyl]-L-α-aspartyl]-phenylalanine 1-methyl ester, N—[N-[3-(3-methoxy-4-hydroxyphenyl)propyl]-L-α-aspartyl]-phenylalanine 1-methyl ester, salts thereof, and the like.

Non-limiting examples of natural high intensity sweeteners include Stevioside, Rebaudioside A, Rebaudioside B, Rebaudioside C, Rebaudioside E, Rebaudioside F, Steviolbioside, Dulcoside A, Rubusoside, mogrosides, brazzein, neohesperidin dihydrochalcone (NHDC), glycyrrhizic acid and its salts, thaumatin, perillartine, pernandulcin, mukuroziosides, baiyunoside, phlomisoside-I, dimethyl-hexahydrofluorene-dicarboxylic acid, abrusosides, periandrin, carnosiflosides, cyclocarioside, pterocaryosides, polypodoside A, brazilin, hernandulcin, phillodulcin, glycyphyllin, phlorizin, trilobatin, dihydroflavonol, dihydroquercetin-3-acetate, neoastilibin, trans-cinnamaldehyde, monatin and its salts, selligueain A, hematoxylin, monellin, osladin, pterocaryoside A, pterocaryoside B, mabinlin, pentadin, miraculin, curculin, neoculin, chlorogenic acid, cynarin, siamenoside and others.

High intensity sweeteners can be derived from the modification of natural high intensity sweeteners, for example, by fermentation, enzymatic treatment, or derivatization.

A growing number of consumers perceive the ability to control their health by enhancing their current health and/or hedging against future diseases. This creates a demand for food products with enhanced characteristics and associated health benefits, specifically a food and consumer market trend towards “whole health solutions” lifestyle. The term “natural” is highly emotive in the world of sweeteners and has been identified as one of key trust, along with “whole grains”, “heart-healthy” and “low-sodium”. ‘Natural’ term is closely related to ‘healthier’.

Stevia rebaudiana is a perennial shrub of the Asteraceae (Compositae) family native to certain regions of South America. The leaves of the plant contain from 10 to 20% of diterpene glycosides, which are around 150 to 450 times sweeter than sugar. The leaves have been traditionally used for hundreds of years in Paraguay and Brazil to sweeten local beverages, foods and medicines.

At present there are more than 230 Stevia species with significant sweetening properties. The plant has been successfully grown under a wide range of conditions from its native subtropics to the cold northern latitudes.

Steviol glycosides have zero calories and can be used wherever sugar is used. They are ideal for diabetic and low calorie diets. In addition, the sweet steviol glycosides possess functional and sensory properties superior to those of many high potency sweeteners.

The extract of Stevia rebaudiana plant contains a mixture of different sweet diterpene glycosides, which have a single base—steviol and differ by the presence of carbohydrate residues at positions C13 and C19. These glycosides accumulate in Stevia leaves and compose approximately 10%-20% of the total dry weight. Typically, on a dry weight basis, the four major glycosides found in the leaves of Stevia are Dulcoside A (0.3%), Rebaudioside C (0.6%), Rebaudioside A (3.8%) and Stevioside (9.1%). Other glycosides identified in Stevia extract include Rebaudioside B, C, D, E, and F, Steviolbioside and Rubusoside (FIG. 1).

The chemical structures of the diterpene glycosides of Stevia rebaudiana are presented in FIG. 1. The physical and sensory properties are well studied only for Stevioside and Rebaudioside A. The sweetness potency of Stevioside is around 210 times higher than sucrose, Rebaudioside A around 300 times, and Rebaudioside C and Dulcoside A around 30 times. The Stevia extract containing Rebaudioside A and Stevioside as major components showed sweetness potency around 250 times. Rebaudioside A and Rebaudioside D are considered to have most favorable sensory attributes of all major Steviol Glycosides (TABLE 1).

TABLE 1
			Mol.	Solubility in	Relative
Name	Formula	TMelt, ° C.	Weight	water, %	sweetness	Quality of taste
Steviol	C20H30O3	212-213	318.45	ND	ND	Very bitter
Steviolmonoside	C26H40O8	ND	480.58	ND	ND	ND
Stevioside	C38H60O18	196-198	804.88	0.13	210	Bitter
Rebaudioside A	C44H70O23	242-244	967.01	0.80	200-400	Less Bitter
Rebaudioside B	C38H60O18	193-195	804.88	0.10	150	Bitter
Rebaudioside C	C44H70O22	215-217	951.01	0.21	30	Bitter
Rebaudioside D	C50H80O28	248-249	1129.15	1.00	220	Like sucrose
Rebaudioside E	C44H70O23	205-207	967.01	1.70	170	Like sucrose
Rebaudioside F	C43H68O22	ND	936.99	ND	ND	ND
Dulcoside A	C38H60O17	193-195	788.87	0.58	30	Very bitter
Steviolbioside	C32H50O13	188-192	642.73	0.03	90	Unpleasant
Rubusoside	C32H50O13	ND	642.73	ND	110	Very bitter

In addition to the commercially known steviol glycosides (Table 1), several new steviol glycosides (glycosylated diterpene) have been found in Stevia leaf extracts, as shown in Table 2a.

TABLE 2a
Summary of formula and R-groups of identified steviol glycosides
(see FIG. 1 for backbone structure)
		Trivial	Mol.
#	Common name	formula	Wt.	R1	R2	Reference
1. Steviol + Glucose (SvGn)
1.1	Steviolmonoside	SvG1	481	H	Glcβ1-	Ohta et al.
						(2010)
1.2	Steviolmonoside	SvG1	481	Glcβ1-	H	Gardena et
	A					al. (2010)
1.3	Rubusoside	SvG2	643	Glcβ1-	Glcβ1-	Ohta et al.
						(2010)
1.4	Steviolbioside	SvG2	643	H	Glcβ(1-2)Glcβ1-	Kohda et
						al. (1976)
1.5	Stevioside	SvG3	805	Glcβ1-	Glcβ(1-2)Glcβ1-	Bridel &
						Lavielle
						(1931)
1.6	Stevioside A	SvG3	805	Glcβ(1-	Glcβ1-	Wu et al.
				2)Glcβ1-		(2012)
1.7	Rebaudioside B	SvG3	805	H	Glcβ(1-	Kohda et
					2)[Glcβ(1-	al. (1976)
					3)]Glcβ1-
1.8	Rebaudioside G	SvG3	805	Glcβ1-	Glcβ(1-3)Glcβ1-	Ohta et al.
						(2010)
1.9	Stevioside B	SvG3	805	Glcβ(1-	Glcβ1-	Chaturvedula
				3)Glcβ1-		& Zamora
						(2014)
1.10	Rebaudioside E	SvG4	967	Glcβ(1-	Glcβ(1-2)Glcβ1-	Sakamoto
				2)Glcβ1-		et al.
						(1977a)
1.11	Rebaudioside A	SvG4	967	Glcβ1-	Glcβ(1-	Kohda et
					2)[Glcβ(1-	al. (1976)
					3)]Glcβ1-
1.12	Rebaudioside A2	SvG4	967	Glcβ1-	Glcβ(1-	Chaturvedula
					6)Glcβ(1-	& Prakash
					2)Glcβ1-	(2011d)
1.13	Rebaudioside D	SvG5	1 129	Glcβ(1-	Glcβ(1-	Sakamoto
				2)Glcβ1-	2)[Glcβ(1-	et al.
					3)]Glcβ1-	(1977a)
1.14	Rebaudioside I	SvG5	1 129	Glcβ(1-	Glcβ(1-	Ohta et al.
				3)Glcβ1-	2)[Glcβ(1-	(2010)
					3)]Glcβ1-
1.15	Rebaudioside L	SvG5	1 129	Glcβ1-	Glcβ(1-	Ohta et al.
					6)Glcβ(1-	(2010)
					2)[Glcβ(1-
					3)]Glcβ1-
1.16	Rebaudioside Q2	SvG5	1 129	Glcα(1-	Glcβ(1-2)Glcβ1-	Chaturvedula
				2)Glcα(1-		& Prakash
				4)Glcβ1-		(2011c)
1.17	Rebaudioside Q	SvG5	1 129	Glcβ1-	Glcα(1-	—
					4)Glcβ(1-
					2)[Glcβ(1-
					3)]Glcβ1-
1.18	Rebaudioside I2	SvG5	1 129	Glcβ1-	Glcα(1-	Chaturvedula
					3)Glcβ(1-	et al.
					2)[Glcβ(1-	(2011c)
					3)]Glcβ1-
1.19	Rebaudioside Q3	SvG5	1 129	Glcβ1-	Glcα(1-	Chaturvedula
					4)Glcβ(1-	et al.
					3)[Glcβ(1-	(2011c)
					2)]Glcβ1-
1.20	Rebaudioside I3	SvG5	1 129	Glcβ(1-	Glcβ(1-2)Glcβ1-	Chaturvedula
				2)[Glcβ(1-		et al.
				6)]Glcβ1-		(2011c)
1.21	Rebaudioside M	SvG6	1 291	Glcβ(1-	Glcβ(1-	Ohta et al.
				2)[Glcβ (1-	2)[Glcβ(1-	(2010)
				3)]Glcβ1-	3)]Glcβ1-
2. Steviol + Rhamnose + Glucose (SvR1Gn)
2.1	Dulcoside A	SvR1G2	789	Glcβ1-	Rhaα(1-2)Glcβ1-	Kobayashi
						et al.
						(1977)
2.2	Dulcoside B	SvR1G2	789	H	Rhaα(1-	Ohta et al.
					2)[Glcβ(1-	(2010)
					3)]Glcβ1-
2.3	Rebaudioside C	SvR1G3	951	Glcβ1-	Rhaα(1-	Sakamoto
					2)[Glcβ(1-	et al.
					3)]Glcβ1-	(1977b)
2.4	Rebaudioside	SvR1G3	951	Rhaα(1-	Glcβ(1-2)Glcβ1-	Purkayastha
	C2a			2)Glcβ1-		(2016)
2.5	Rebaudioside S	SvR1G3	951	Rhaα(1-	Glcα (1-	Ibrahim et
				2)Glcβ1-	2)Glcβ1-	al (2016)
2.6	Rebaudioside H	SvR1G4	1 112	Glcβ1-	Glcβ(1-	Ohta et al.
					3)Rhaα(1-	(2010)
					2)[Glcβ(1-
					3)]Glcβ1-
2.7	Rebaudioside K	SvR1G4	1 112	Glcβ(1-	Rhaα(1-	Ohta et al.
				2)Glcβ1-	2)[Glcβ(1-	(2010)
					3)]Glcβ1-
2.8	Rebaudioside J	SvR1G4	1 112	Rhaα(1-	Glcβ(1-	Ohta et al.
				2)Glcβ1-	2)[Glcβ(1-	(2010)
					3)]Glcβ1-
2.9	Rebaudioside N	SvR1G5	1 274	Rhaα(1-	Glcβ(1-	Ohta et al.
				2)[Glcβ(1-	2)[Glcβ(1-	(2010)
				3)]Glcβ1-	3)]Glcβ1-
2.10	Rebaudioside O	SvR1G6	1 436	Glcβ(1-	Glcβ(1-	Ohta et al.
				3)Rhaα(1-	2)[Glcβ(1-	(2010)
				2)[Glcβ(1-	3)]Glcβ1-
				3)]Glcβ1-
2.11	Rebaudioside	SvR1G6	1 436	Glcβ(1-	Glcβ(1-	Purkayastha
	O2a			4*)Rhaα(1-	2)[Glcβ(1-	(2016)
				2)[Glcβ(1-	3)]Glcβ1-
				3)]Glcβ1-
2.12	Rebaudioside	SvR1G4	1 112	Glcβ(1-	Rhaα(1-	Purkayastha
	K2a			6)Glcβ1-	2)[Glcβ(1-	(2016)
					3)]Glcβ1-
3. Steviol + Xylose + Glucose (SvX1Gn)
3.1	Stevioside F	SvX1G2	775	Glcβ1-	Xylβ(1-2)Glcβ1-	Chaturvedula
						& Prakash
						(2011b)
3.2	Rebaudioside F	SvX1G3	937	Glcβ1-	Xylβ(1-	Starratt et
					2)[Glcβ(1-	al. (2002)
					3)]Glcβ1-
3.3	Rebaudioside F2	SvX1G3	937	Glcβ1-	Glcβ(1-	Chaturvedula
					2)[Xylβ(1-	& Prakash
					3)]Glcβ1-	(2011b)
3.4	Rebaudioside F3	SvX1G3	937	Xylβ(1-	Glcβ(1-2)Glcβ1-	Chaturvedula
				6)Glcβ1-		et al.
						(2011d)
3.5	Rebaudioside R	SvX1G3	937	Glcβ1-	Glcβ(1-	Ibrahim et
					2)[Glcβ(1-3)]	al (2016)
					Xylβ1-
3.6	Rebaudioside	SvX1G4	1 099	Xylβ(1-	Glcβ(1-	Purkayastha
	Ua			2)Glcβ1-	2)[Glcβ(1-	(2016)
					3)]Glcβ1-
3.7	Rebaudioside	SvX1G4	1 099	Xylβ(1-	Glcβ(1-2)Glcβ1-	Purkayastha
	U2a			2*)[Glcβ(1-		(2016)
				3)]Glcβ1-
3.8	Rebaudioside	SvX1G5	1 261	Glcβ(1-	Xylβ(1-	Purkayastha
	Va			2)[Glcβ(1-	2*)[Glcβ(1-	(2016)
				3)]Glcβ1-	3)]Glcβ1-
3.9	Rebaudioside	SvX1G5	1 261	Xylβ (1-	Glcβ(1-	Prakash &
	V2a			2)[Glcβ(1-	2)[Glcβ(1-	Chaturvedula
				3)]Glcβ1-	3)]Glcβ1-	(2013)
4. Steviol + Arabinose + Glucose (SvA1Gn)
4.1	Rebaudioside	SvA1G4	1 098	Glcβ(1-	Glcβ(1-2)Glcβ1-	Purkayastha
	Wa			2)[Araβ(1-		(2016)
				3*)]Glcβ1
4.2	Rebaudioside	SvA1G4	1 098	Araβ(1-	Glcβ(1-	Purkayastha
	W2a			2*)Glcβ1	2)[Glcβ(1-	(2016)
					3)]Glcβ1-
4.3	Rebaudioside	SvA1G4	1 098	Araβ(1-	Glcβ(1-	Purkayastha
	W3a			6)Glcβ1-	2)[Glcβ(1-	(2016)
					3)]Glcβ1-
4.4	Rebaudioside	SvA1G5	1 260	Glcβ(1-	Glcβ(1-	Purkayastha
	Ya			2)[Araβ(1-	2)[Glcβ(1-	(2016)
				3*)]Glcβ1	3)]Glcβ1-
5. Steviol + Fructose + Glucose (SvF1Gn)
5.1	Rebaudioside A3	SvF1G3	967	Glcβ1-	Glcβ(1-	Chaturvedula
					2)[Fruβ(1-	et al.
					3)]Glcβ1-	(2011b)
6. Steviol + galactose + Glucose (SvGa1Gn)
6.1	Rebaudioside	SvGa1	1 128	Galβ(1-	Glcβ(1-	Purkayastha
	Ta	G4		2*)Glcβ1	2)[Glcβ(1-	(2016){circumflex over ( )}
					3)]Glcβ1-
7. Steviol + de-oxy glucose + Glucose (SvdG1Gn)
7.1	Stevioside D	SvdG1G2	789	Glcβ1-	6-deoxyGlcβ(1-	Chaturvedula
					2)Glcβ1-	& Prakash
						(2011a)
7.2	Stevisoide E	SvdG1G3	951	Glcβ1-	6-deoxyGlcβ(1-	Chaturvedula
					2)[Glcβ(1-	& Prakash
					3)]Glcβ1-	(2011a)
7.3	Stevioside E2	SvdG1G3	951	6-	Glcβ(1-	Chaturvedula
				deoxyGlcβ1-	2)[Glcβ(1-	et al.
					3)]Glcβ1-	(2011e)

Besides diterpene glycosides, a number of flavonoids, labdane diterpene, triterpenes, sterols, and volatile oils have also been found in the extracts of Stevia rebaudiana, collectively referred to as plant molecules, as shown in Table 2b.

TABLE 2b
Chemical Classes   Chemical Components
Monoterpenoids     Borneol
Diterpenoids       Austroinulin, 6-0-acetyl austroinulin, 6-acetyl austroinulin
                   7-0-acetyl austroinulin, Sterebin A, B, C, D, E, F, G, H, Jhanol
Triterpenoids      Amyrin beta acetate
Sesquiterpenes     α-bergamotene, Bisabolene, β-bourbonene, δ-cadinene, γ-cadinene
Essential oils     β-caryophyllene, Trans β-tarnesene, α-humulene, δ-cadiene
                   caryophyllene oxide, Nerolidol, Linalol, α-terpineol, Terpinen-4-ol
Sterol derivatives Stigmasterol , β-sitosterol, Campesterol
Flavonoids         Glucosy1-4′-O-apigenin, Glucosyl-7-O-luteolin, Rhamnosyl-3-O-
                   kaempferol, Quercetin, Glucosyl-3-O-quercetin, Arabinosyl-3-O-
                   quercetin, 5,7,3′-methoxyflavone, 3,6,4′-methoxyflavone,
                   Centaureidin, avicularin

All steviol glycosides provide sweetness and other taste attributes at a higher than certain threshold level of concentrations in water. Below the threshold level of concentration, the steviol glycoside components and their mixtures as found in a typical non-limiting Stevia extract as shown below has no recognizable sweetness taste. But such Stevia extract below the threshold level of significant sweetness recognition show remarkable characteristics of sweet and flavor profile modification in food and beverage applications.

This invention relates to use of the following Stevia extracts (Table 3) with the varying level of different steviol glycosides and other Stevia plant-derived glycosides, the combination of which contributes no significant sweetness but modifies flavor and sweetness profile at certain concentration in typical food and beverage applications.

TABLE 3
														Minor
														Steviol
														Glycosides
	Steviol Glycosides*, %		and related
Stevia				Reb		Dulco-	Rubu-		Steviolbio-				TSG*	plant
Extracts	Reb A	Stevioside	Reb D	F	Reb C	side A	soside	Reb B	side	Reb E	Reb N	Reb O	(%)	molecules
PCS-5001	10-20	4-12	1-4	1-5	10-25	1-5	1-4	0.5-5	0.5-5	1-4	0.5-4	0.5-4	45-65	35-50
PCS-1015	18-25	5-10	8-20	0-1	1-3	0-1	0-1	0.5-5	0-1	2-6	4-8	3-8	55-65	35-45
PSB-5005	15-30	3-12	1-10	0-5	5-15	0-5	0-5	0-8	0-5	0-5	0-6	0-5	45-70	30-55%
*TSG or Total Steviol Glycosides contain the Steviol Glycosides that are recognized by Codex Alimentarius (a commission of FAO and WHO) and major regulatory authorities

The present invention also relates to the Stevia extracts that contain major steviol glycosides (Table 3) and other minor steviol glycosides and glycosylated diterpene derivatives (water soluble molecules). The non-limiting examples of such minor molecules are Reb E, Reb G, Reb H, Reb I, Reb K, Reb L, Reb M, Reb N, Reb O (M. Ohta, S. Sasa, A. Inoue, et al. “Characterization of Novel Steviol Glycosides from Leaves of Stevia rebaudiana Morita.” J. Appl. Glycosci., 57, 199-209 (2010)).

The present invention is also directed to a method of making a specific Stevia extract composition, including: extracting steviol glycosides and other water soluble molecules from leaves of a Stevia rebaudiana plant, and separating the excess steviol glycosides than the amount and type of steviol glycosides required to contribute the taste and flavor modifying characteristics of the Stevia extract.

This invention combine the different natural sweeteners, especially steviol glycosides in certain proportion along with other water soluble molecules to provide enhanced sweetness and flavor profile in food and beverage application, which can be blended with other natural caloric sweeteners to impart more desirable sweetness profile. Non-limiting examples of caloric sweeteners include dextrose, fructose, sucrose, maltose, lactose, corn syrup, gluco-syrup derived from different carbohydrates, cane syrup, flavored sugar, honey, molasses,

This invention combine the different natural sweeteners, especially steviol glycosides in certain proportion along with other water soluble molecules to provide enhanced sweetness and flavor profile in food and beverage application, which can be blended with other natural non-caloric sweeteners to impart more desirable sweetness profile. Non-limiting examples of natural high intensity sweeteners include steviol glycosides, brazzein, monatin and its salt, neohesperidin dihydrochalcone (NHDC), glycyrrhizic acid and its salts, thaumatin, mogrosides and lu han guo extracts, perillartine, mabinlin, pentadin, miraculin, curculin, neoculin, chlorogenic acid, cynarin, siamenoside and others.

This invention combine the different natural sweeteners, especially steviol glycosides in certain proportion along with other water soluble molecules to provide enhanced sweetness and flavor profile in food and beverage application, which can be blended with other synthetic non-caloric sweeteners to impart more desirable sweetness profile. Non-limiting examples of synthetic sweeteners include sucralose, potassium acesulfame, aspartame, alitame, advantame, saccharin, neohesperidin dihydrochalcone synthetic derivatives, cyclamate, neotame, dulcin, suosan, N—[N-[3-(3-hydroxy-4-methoxyphenyl)propyl]-L-α-aspartyl]-phenylalanine 1-methyl ester, N—[N-[3-(3-hydroxy-4-methoxyphenyl)-3-methylbutyl]-α-aspartyl]-L-phenylalanine 1-methyl ester, N—[N-[3-(3-methoxy-4-hydroxyphenyl)propyl]-L-α-aspartyl]-L-phenylalanine 1-methyl ester, salts thereof, and the like.

BRIEF SUMMARY OF THE INVENTION

The present invention is directed to a taste and flavor modifying composition. The composition includes different steviol glycosides with other water soluble molecules derived from Stevia leaf, such as non-limiting examples of plant glycosides, flavonoids, labdane diterpene, triterpenes, which can modify the intensity of a taste and/or a flavor in a food or beverage product.

The present invention is also directed to a food or beverage product having an intense taste and flavor profile, wherein the food or beverage product includes a taste and flavor modifying composition comprising the Stevia extract of steviol glycosides and water soluble molecules derived from Stevia plant. A wide range of food and beverage products, such as, but not limited to, carbonated soft drinks, fruit juices, dairy foods, dairy beverages, baked goods, cereal products, snack foods, and table top sweeteners, may be made in accordance with the present invention. The taste and flavor profile of a food or beverage product including a taste and flavor modifying composition, wherein the taste and flavor modifying composition comprising the Stevia extract of steviol glycosides and water soluble molecules derived from Stevia plant, may be more intense than a comparative taste and flavor profile of a comparative food or beverage product which does not include the taste and flavor modifying composition. Moreover, the mouthfeel and overall taste perception of a food or beverage product including the taste and flavor modifying composition, wherein the taste and flavor enhancing composition includes the complex mixture of steviol glycosides and water soluble molecules, may be improved in relation to a mouthfeel and overall taste perception of a comparative food or beverage product which does not include the taste and flavor enhancing composition.

The present invention is further directed to a method of increasing the taste and flavor intensity of a food or beverage product, including the step of adding a taste and flavor enhancing composition to the food or beverage product, wherein the taste and flavor modifying composition comprising the Stevia extract of steviol glycosides and water soluble molecules derived from Stevia plant. The present invention is also directed to a method of improving the organoleptic properties of a food or beverage product including a high fructose syrup, including the step of adding the taste and flavor modifying composition to the food or beverage product. For example, adding the taste and flavor modifying composition may cause the high fructose syrup, such as high fructose corn syrup, to taste more like sugar. Also, if the high fructose syrup is high fructose corn syrup 42 (HFCS 42), adding the taste and flavor enhancing composition may cause the HFCS 42 to taste more like high fructose corn syrup 55 (HFCS 55).

The present invention is further directed to a method of increasing the taste and flavor intensity of a medical food and pharma product, including the step of adding a taste and flavor modifying composition to the food or beverage product, wherein the taste and flavor modifying composition comprising the Stevia extract of selected steviol glycosides and water soluble molecules derived from Stevia plant. The present invention is also directed to a method of improving the organoleptic properties of a medical food or pharma product containing functional food ingredients like vitamins, minerals and amino acids, including the step of adding the taste and flavor modifying composition to the food or beverage product. For example, adding the taste and flavor modifying composition may cause the off-taste due to vitamins, mineral, amino acids and other non-limiting functional ingredients, to improve taste and palatability.

The present invention is also directed to a method of making a taste and flavor enhancing composition, including: extracting steviol glycosides and other water soluble molecules from leaves of a Stevia rebaudiana plant, and separating the excess steviol glycosides than the amount and type of steviol glycosides required to contribute the taste and flavor modifying characteristics of the Stevia extract.

The foregoing has outlined rather broadly the features and technical advantages of the present invention in order that the detailed description of the invention that follows may be better understood. Additional features of the invention which form the subject of the claims of the invention will be described hereinafter. It should be appreciated by those skilled in the art that the specific embodiments disclosed may be readily utilized as a basis for modifying or designing other methods or structures for carrying out the same purposes of the present invention. It should also be realized by those skilled in the art that such equivalent constructions do not depart from the spirit and scope of the invention as set forth in the appended claims. The novel features which are believed to be characteristic of the invention, both as to its organization and method of operation, together with further objects and advantages will be better understood from the following description.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the chemical structure of the diterpene glycosides of Stevia rebaudiana.

FIG. 2 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to a cola flavored carbonated soft drink.

FIG. 3 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to an iced tea beverage.

FIG. 4 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to an iced tea beverage.

FIG. 5 is a bar graph showing the effect of Stevia extract on the flavor profile of roasted peanuts.

FIG. 6 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to tomato ketchup.

FIG. 7 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to chocolate milk.

FIG. 8 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to poppy seed muffins.

FIG. 9 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to tortilla chips.

FIG. 10 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to beef jerky.

FIG. 11 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to brown gravy.

FIG. 12 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to chocolate milk.

FIG. 13 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to vanilla custard.

FIG. 14 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to chocolate milk.

FIG. 15 is a bar graph showing the modification of flavor and sweetness profiles caused by the addition of Stevia extract to vanilla yogurt.

DETAILED DESCRIPTION

Embodiments of the present invention are described in the following examples.

EXAMPLES

Example 1A: Detection of Concentration Threshold for Sweetness Recognition

To detect the sweetness recognition level of PCS-5001, PCS 1015 and PSB 5005 (Stevia extract), the test method outlined by Harman, et al (Food Technology, November 2013) was used with ten trained panelists that have been previously qualified for their taste acuity and trained in the use of a sweetness intensity rating scale. The panelists evaluated a series of aqueous solutions of sucrose and the Stevia extract (PCS-5001, PCS-1015, or PSB 5005) at room temperature; the sucrose solutions of 1.5% concentration and the Stevia extract solutions with concentrations ranging between 100 and 120 ppm for PCS-5001, 70-80 ppm for PCS-1015, and 60-70 ppm for PSB 5005 were prepared with filtered water. The objective of the test was to determine the sweetness recognition level of the Stevia extract. The evaluations were done in triplicate using the same panelists so that a total of 30 values were generated for each average data point.

The samples were coded and presented in random order to panel members to taste and determine which sample was sweeter (ASTM E2164-08: Standard Method for Directional Difference Test). Panelists were asked to focus only on sweet attribute of those samples and to use warm water and salt solution in order to cleanse the palate between samples.

The results were tallied and significance was calculated by SIM 2000 (Sensory Computer System, NJ). Results are presented in Table 4. The overall sweetness of those samples was barely detectable. The 2-AFC shows that 100 ppm PCS-5001, 70 ppm of PCS-1015 and 60 ppm of PSB 5005 solutions were the least sweet samples and were significantly less sweet then the 1.5% sugar control. The sample with 120 ppm PCS-5001 and 80 ppm PCS-1015 were the sweetest samples showing significantly higher sweetness than the 1.5% sugar control (Table 4). The recognition threshold concentration of STEVIA EXTRACT (PCS-5001) in water was determined to be 100 ppm. The recognition threshold concentration of STEVIA EXTRACT (PCS-1015) in water was determined to be 70 ppm. The sweetness recognition threshold of STEVIA EXTRACT (PSB 5005) in water was determined to be 60 ppm.

TABLE 4
Sweetness perception of Stevia Extract in different concentration
against 1.5% sugar solution.
Comparison of sweetness
perception of STEVIA	Sugar solution	Stevia Extract
EXTRACT in water	(1.5%) sweeter?	solution sweeter?	P-Value	Significance
PCS-5001: 100 ppm, N=30	23	7	0.0052	***
PCS-5001: 110 ppm, N=30	20	10	0.0987	**
PCS-5001: 120 ppm, N=30	9	21	0.0457	***
PCS-1015:  70 ppm, N=30	26	4	0.0001	***
PCS-1015:  80 ppm, N=30	5	25	0.0003	***
PSB-5005:  60 ppm, N=30	24	6	0.0014	***
PSB-5005:  70 ppm, N=30	19	11	0.2005	NS

Example 1B: Sweetness Detection of Concentration Threshold for Sweetness Detection

The ten panel members evaluated a series of lemon-lime flavored carbonated soft drink (CSD) sweetened with sucrose and STEVIA EXTRACT at room temperature; the evaluations were done in triplicate using the same panelists so that at least 30 values were generated for each average data point. The lemon lime flavored carbonated soft drink control sample had 1.5% sucrose concentration and the test sample contained STEVIA EXTRACT (PCS-5001) with concentrations at 110 and 120 ppm or STEVIA EXTRACT (PCS-1015) with concentrations of 70 and 90 ppm. Other ingredients in the CSD samples were citric acid, lemon-lime flavor, sodium benzoate, potassium citrate and xanthan gum. The objective of the test was to determine the sweetness detection limit of STEVIA EXTRACT. Tests were conducted as outlined in Example 1A.

The samples with 120 ppm PCS-5001(STEVIA EXTRACT) and 90 ppm PCS-1015(STEVIA EXTRACT) showed no significant difference in sweetness than the 1.5% sugar control. The recognition threshold concentration of PCS-5001(STEVIA EXTRACT) in a lemon-lime flavored carbonated soft drink water was determined to be 110 ppm. The recognition threshold concentration of PCS-1015 (STEVIA EXTRACT) in a lemon-lime flavored carbonated soft drink water was determined to be 70 ppm. Results are shown in table 5.

TABLE 5
Sweetness perception of STEVIA EXTRACT in different concentrations against
1.5% sugar solution in a typical carbonated soft drink (CSD)
Sweetness perception of	CSD sample with	CSD sample with
SIEVIA EXTACT in CSD	Sugar sweeter?	Stevia Sweeter?	P-Value	Significance
PCS-5001: 110 ppm, N=30	23	7	0.0052	***
PCS-5001: 120 ppm, N=36	20	16	0.677	NS
PCS-1015:  70 ppm, N=30	21	9	0.0428	***
PCS-1015:  90 ppm, N=30	12	18	0.3616	NS

Example 2: Effect of Stevia Extract on Flavor Modification in a Typical Carbonated Soft Drink Application

A cola flavored carbonated soft drink was developed to evaluate the effect of PCS-5001 and PCS-1015 (Stevia extract) on the sweetness and flavor profile of the beverage that was sweetened with sugar and Stevia sweetener to achieve 30% sugar reduction (Table 6). The samples with and without PCS-5001 or PCS-1015 were evaluated by thirty consumer panel members, who assigned relative values to each sample for overall Liking, sweetness, vanilla flavor, brown note, and aftertaste on a 10-pt continuous intensity scale as outlined in Table 7.

TABLE 6
Cola flavored Soft drink for sensory evaluation
	Control:	Test: 30% Sugar	Test: 30% Sugar
COLA BEVERAGE	30%	Reduction with	Reduction with
FORMULA	Sugar Reduction	PCS-5001	PCS-1015
Water	91.68	91.67	91.67
Sugar	7.89	7.89	7.89
Cola Flavor-Flavor Systems	0.375	0.375	0.375
Phosphoric Acid 85%	0.0333	0.0333	0.0333
Caffeine	0.0100	0.0100	0.0100
Steviol glycoside	0.0100	0.0100	0.0100
PCS-5001		0.0110
PCS-1015			0.0080
Total	100	100	100

TABLE 7
Sensory evaluation of Cola flavored carbonated soft drink
Nature of Participants: Company employees
Number of Sessions      1
Number of               30
Participants:
Test Design:            Balanced, randomized within pair. Blind
Sensory Test            Intensity and acceptance ratings
Method:
Environmental           Standard booth lighting
Condition
Attributes and Scales:
Overall Acceptance on a 10-pt hedonic scale where 10 = Extremely Like and 0 =
Extremely Dislike
Overall Liking, Sweetness, Vanilla flavor, Brown note, and Sweet Aftertaste.
10-pt continuous intensity scale where 0 = Imperceptible and 10 = Extremely
Pronounced
Statistical Analysis:   ANOVA (by Block) with Post Hoc Duncan'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 instructed to
Instruction:            read ingredient statement, evaluate each sample.

FIG. 2 shows the modification of flavor and sweetness profiles caused by the addition of Stevia extract (PCS-5001). The results indicated the sample containing Stevia extract PCS-5001 and the sample containing PCS-1015 had significantly higher cola flavor, vanilla flavor, brown spice notes and overall liking compared to the control samples (at 95% confidence). The sample containing PCS-5001 had directionally lower bitterness, and bitter aftertaste intensity compared to the control samples (at 90% and 95% confidence respectively). The sample containing PCS-1015 had directionally lower bitterness, and sweet aftertaste intensity compared to the control samples (at 80% confidence). In addition, the sample with Stevia extract (PCS-1015) had significantly lower bitter aftertaste compared to the control sample (at 95% confidence).

Example 3: Peach Flavored Tea Beverage for Sensory Evaluation

A peach flavored black tea drink was developed to evaluate the effect of STEVIA EXTRACT on the sweetness and flavor profile of the beverage that was sweetened with sugar and Stevia sweetener to achieve 30% sugar reduction (Table 8). The samples with and without STEVIA EXTRACT were evaluated as outlined in EXAMPLE 2 by thirty consumer panel members, who assigned relative values to sweetness, bitterness, peach flavor, tea flavor, acid intensity, astringency, and aftertaste on 10-pt continuous intensity scale where 0=Imperceptible and 10=extremely pronounced.

TABLE 8
Peach Flavored Tea Beverage samples for sensory evaluation
		Reduce Sugar	Reduce Sugar
	Reduced Sugar	Tea with PCS-	Tea with PCS-
	Tea	5001	1015
Water	95.71	95.70	95.71
Sucrose	3.850	3.850	3.850
Black Tea Powder	0.275	0.275	0.275
Citric Acid	0.0880	0.0880	0.0880
Peach Flavor	0.0330	0.0330	0.0330
Sodium Citrate	0.0150	0.0150	0.0150
Potassium Sorbate	0.0150	0.0150	0.0150
Steviol Glycoside	0.0140	0.0140	0.0140
Stevia Extract PCS-5001		0.0120
Stevia Extract PCS-1015			0.0080
Xanthan Gum-TIC	0.0013	0.0013	0.0013

FIG. 3 shows the modification of flavor and sweetness profiles contributed by the addition of STEVIA EXTRACT (PCS-5001) in peach flavored ice tea beverage. The results indicated that the test sample containing PCS-5001 had significantly higher peach flavor, and overall liking (at 95%, confidence). The sample containing PCS-5001 had significantly lower astringency than the control sample (at 95% confidence). The results shown in FIG. 4 indicated that the test sample containing PCS-1015 had significantly higher peach flavor, black tea flavor, and overall liking (at 95%, confidence). The sample PCS-1015 also had significantly lower astringency, sweet intensity, bitter intensity, and bitter aftertaste than the Control sample (at 95% confidence). In addition, the PCS-1015 sample had lower sweet aftertaste intensity than the Control sample at 90% confidence).

Example 4: Effect of Stevia Extract on Flavor Modification of Savory Applications

A seasoning blend was developed to determine the flavor modification effect of Stevia extract in a seasoning blend on reduced sugar roasted peanut samples. 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 two samples (Table 9) included: 1) 50% sugar reduced control sample containing Stevia glycosides, and 2) 50% reduced sugar test sample containing steviol glycoside and Stevia extract, PCS-5001 or PCS-1015.

The objective of the test was to determine if the addition of Stevia extract affects the flavor profile of a savory snack food. The results indicated that the addition of PCS-5001 at 110 ppm and PCS-1015 at 70 ppm provided flavor modification (FIG. 5). The test samples containing 110 ppm PCS-5001 had significantly higher salt intensity, smoke flavor, and bitter intensity compared to the control (95% confidence). The test sample also had lower sweet intensity than the control (95% confidence). In addition, the test sample containing Stevia extract had directionally higher spice and chili notes (90% confidence). The test sample containing PCS-1015 had significantly higher salt intensity than the control sample (at 95% confidence). The test sample showed an increase in heat/spice intensity, and chili flavor compared to the control.

TABLE 9
Effect of STEVIA EXTRACT on snack and seasoning applications
	Steviol	Steviol Glycoside +	Steviol Glycoside +
	Glycoside	Stevia Extract	Stevia Extract
Unsalted Peanuts	86.8	86.8	86.8
Vegetable oil	2.93	2.93	2.93
Sugar	5.88	5.88	5.88
Salt	2.93	2.93	2.93
Chilli powder	0.174	0.174	0.174
Cumin powder	0.286	0.286	0.286
Garlic powder	0.156	0.156	0.156
Cayenne pepper	0.156	0.156	0.156
Smoke liquid	0.729	0.729	0.729
Steviol Glycoside	0.0243	0.0243	0.0243
PCS-5001		0.0110
PCS-1015			0.0070
Total wt. (g)	100	100	100

TABLE 10
Sensory evaluation of snack and seasoning applications
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 9-pt hedonic scale where 9 = Like Extremely, 5 = Neither Like
Nor Dislike, and 1 = Dislike Extremely
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
Open Ended General Comments
Statistical Analysis:   ANOVA (by Block) with Post Hoc Duncan'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 evaluate each
Instruction:            sample once.

Example 5: Flavor Modification of Sauce and Vegetable Preparation

A tomato ketchup preparation was developed to determine the flavor modification effect of Stevia extract (PCS-1015). A panel of thirty company employees evaluated the overall acceptance and attribute intensities (tomato, onion, vinegar, sweet, saltiness, bitterness and aftertaste) of each sample. The sensory evaluation methodology outlined in Example 4 was adopted for the sauce samples as presented in Table 11.

TABLE 11
Effect of PCS-1015 (stevia extract) on tomato ketchup
		Steviol Glycoside
	Steviol Glycoside	w/Stevia Extract
Tomato Juice (Sieved)	52.4863	52.4793
Tomato Puree	24.6236	24.6236
White Distilled Vinegar	11.3454	11.3454
Water	1.5845	1.5845
Sucrose	2.6511	2.6511
Tomato Paste	5.8311	5.8311
Onion Powder	0.8649	0.8649
Salt	0.5811	0.5811
Steviol glycoside	0.032	0.032
Stevia Extract (PCS 1015)		0.007
Total	100	100

FIG. 6 shows the modification of flavor and sweetness profiles caused by the addition of Stevia extract (PCS-1015). The results indicate the test samples containing Stevia extract, PCS-1015, had a significant increase in herbal notes, and savory (onion/garlic) notes at a 95% confidence interval. The test sample containing PCS-1015 had directionally lower bitterness, bitter aftertaste and overall liking at a 90% confidence interval compared to the control sample.

Example 6: Effect of PCS-1015 (Stevia Extract) on Flavor Modification of Dairy Applications

A chocolate flavored dairy beverage was developed to determine the flavor modification effect of Stevia extract (PCS-1015) in dairy beverage. The panel evaluated samples of chocolate milk for overall acceptance and attribute intensities (chocolate flavor, dairy notes, sweetness, bitterness and aftertaste). The two samples (Table 12) included: 1) 50% sugar reduced control sample containing Stevia glycosides, and 2) 50% reduced sugar test sample containing Stevia glycoside and 80 ppm of Stevia extract, PCS-1015.

TABLE 12
Effect of PCS-1015 (stevia extract) on flavored dairy beverage
	50% Total Sugar	50% Total Sugar
	Reduction	Reduction with stevia
	with steviol	extract and stevia
Dairy Formula	glycoside	glycoside
2% Reduced fat Milk	96.5803	96.5753
Sugar	2.40	2.40
Cocoa Powder	0.80	0.80
Palsgaard 150 ChoMilk	0.20	0.20
Steviol Glycosides	0.0197	0.0197
PCS-1015		0.080
Total	100	100

TABLE 13
Sensory evaluation of Dairy beverage
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           Standard booth lighting
Condition
Attributes and Scales:
Overall Acceptance on a 10-pt hedonic scale where 10 = Extremely Like and 0 =
Extremely Dislike
Overall Liking, sweetness, bitterness, dairy notes, chocolate, and Aftertaste. 10-
pt continuous intensity scale where 0 = Imperceptible and 10 = Extremely
Pronounced
Statistical Analysis:   ANOVA (by Block) with Post Hoc Duncan'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 instructed to read
Instruction:            ingredient statement, evaluate each sample.

FIG. 7 shows the modification of flavor and sweetness profiles caused by the addition of Stevia extract (PCS-1015). The results indicate the 50% sugar reduced sample containing steviol glycoside sweetener and Stevia extract, PCS-1015, had significantly higher chocolate flavor.

Example 7: Effect of Stevia Extract (PCS-5001) on Flavor Modification of Baked Goods Applications

A lemon poppy seed flavored muffin formulation was developed to determine the flavor modification effect of Stevia extract (PCS-5001) in baked good applications. To test the contribution of PCS-5001 in baked goods, lemon flavored poppy seed muffins were baked with a 45% sugar reduced formulation with steviol glycoside as control, and sugar reduced formulation with steviol glycoside and Stevia extract (PCS-5001) as a test sample as shown in Table 14. A thirty member consumer panel evaluated two samples of lemon poppy seed muffins for several attributes (lemon, vanilla flavors, brown notes, sweet & bitter aftertaste).

TABLE 14
Effect of PCS-5001 (stevia extract) on baked goods
	Steviol	Steviol
	Glycoside	glycoside w/
	(400 ppm)	120 ppm
Ingredients	Control	stevia extract
DRY Ingredients
Sucrose	12.3722	12.3682
All Purpose Flour	17.6434	17.6434
Whole Wheat Flour	5.8763	5.8763
Poppy Seeds	1.0648	1.0648
Maltodextrin—10DE	2.1368	2.1368
Fibersol2 (ADM/Matsutani)	1.0648	1.0648
Modified Starch—Inscosity 656	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
Steviol Glycoside	0.0400	0.0400
Stevia extract (PCS-5001)		0.0120
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
	100	100

FIG. 8 shows the modification of flavor and sweetness profiles caused by the addition of Stevia extract (PCS-5001). The panel found that the addition of Stevia extract provided an increase in brown note than control sample without Stevia extract (at 90% confidence).

Example 8: Effect of Stevia Extract (PCS-5001) on Flavor Modification of Reduced Sodium Applications

A 30% salt reduced tortilla chip formulation was developed to determine the flavor modification effect of Stevia extract (PCS-5001) in a salt reduced applications. To test the contribution of PCS-5001 in a salt reduced application, cheddar cheese flavor tortilla chips were coated with a control salt formulation, and a 30% salt reduced formulation with Stevia extract (PCS-5001) as a test sample as shown in Table 15. A sixteen member consumer panel evaluated two samples of cheddar cheese flavored tortilla chips for different attributes (sweet intensity, saltiness, cheese flavor, dairy notes, corn flavor, bitterness, and sweet & bitter aftertaste).

FIG. 9 shows the modification of flavor and salt perception caused by the addition of Stevia extract (PCS-5001). The panel found the addition of Stevia extract in a 30% salt reduced formulation provided an increase in salt perception, parity to the full sodium control. In addition, Stevia extract provided an increase in sweet intensity and dairy note higher than control sample without Stevia extract (at 95% confidence).

TABLE 15
Tortilla Chips with Cheddar Cheese 30% less sodium
	Control	30% Less Salt
Corn chips	78	78.33
Cheese seasoning	10	10.04
Vegetable Oil	11	11.05
Added Salt	1	0.57
PCS-5001		0.01
Total w (g)	100	100.00

Example 9: Effect of Stevia Extract (PCS-5001) on Flavor Modification of Dried Meat Applications

A beef jerky formulation was developed to determine the flavor modification effect of Stevia extract (PCS-5001) in a dried meat applications. To test the contribution of PCS-5001 in a dried meat application, flank steak was marinated with a reduced sugar control formulation, and a 30% sugar reduced formulation with steviol glycosides and Stevia extract (PCS-5001) as a test sample as shown in Table 16. A twenty member consumer panel evaluated two samples of beef jerky for different attributes (sweet intensity, saltiness, black pepper, teriyaki flavor, fat-like intensity, beef flavor and sweet aftertaste).

FIG. 10 shows the modification of flavor and salt perception caused by the addition of Stevia extract (PCS-5001). The panel found the addition of Stevia extract in a 30% sugar reduced formulation provided an increase in salt perception.

TABLE 16
30% sugar reduced Beef Jerky
	Control (%)	Stevia Extract
Flank Steak	75.44	75.44
Balsamic vinegar	10.15	10.15
Salt	2.46	2.46
Pepper	0.83	0.83
Sugar	6.88	6.88
Liquid smoke	0.86	0.86
Water
Garlic powder	0.44	0.44
Onion powder	0.44	0.44
Steviol Glycoside	0.018	0.018
PCS-5001 (stevia extract)		0.0100
Worcestershire sauce	2.46	2.46
	100	100

Example 10: Effect of Stevia Extract (PCS-5001) on Flavor Modification of Reduced Sodium Applications in Brown Gravy

A 30% sodium reduced brown gravy formulation was developed to determine the flavor modification effect of Stevia extract (PCS-5001) in a salt reduced applications. To test the contribution of PCS-5001 in a salt reduced application, a 30% sodium reduced brown gravy formulation, and a 30% salt reduced formulation with Stevia extract (PCS-5001) as a test sample. A thirty member consumer panel evaluated two samples of brown gravy for different attributes (sweet intensity, saltiness, black pepper, beef flavor, and onion/savory notes, bitterness, and sweet & bitter aftertaste).

FIG. 11 shows the modification of flavor and salt perception caused by the addition of Stevia extract (PCS-5001). The panel found the addition of Stevia extract in a 30% salt reduced formulation provided an increase in salt perception compared to 30% sodium reduced control. In addition, Stevia extract provided an increase in savory and black pepper note higher than control sample without Stevia extract (at 95% confidence). There was also a decrease in bitter aftertaste.

Example 11: Effect of Stevia Extract on Flavor Modification of Dairy Product

To evaluate the contribution of PCS-1015 (MLD-1), a Stevia extract, to a dairy product, two 50% reduced sugar chocolate milk samples were prepared and tested by a consumer panel of 30 company employees. The consumer panel evaluated those two samples of chocolate milk for overall acceptance and attribute intensities (chocolate flavor, dairy notes, sweetness, bitterness and aftertaste) in two sessions. In session one, the two samples included: 1) a 50% sugar reduced control sample containing PureCircle Alpha (steviol glycoside sweetener) and 2) 50% sugar reduced test sample containing PureCircle Alpha and 70 ppm PCS-1015 (MLD-1). In session two, the two samples included: 1) a 50% sugar reduced control sample containing PureCircle Alpha (steviol glycoside sweetener) and 2) 50% sugar reduced test sample containing PureCircle Alpha and 80 ppm PCS-1015 (MLD-1). Tables 17 shows the formula of the control and test samples of 50% reduced sugar.

TABLE 17
50% sugar reduced Chocolate Milk with PCS-1015
		50% Total	50% Total
	50% Total	Sugar	Sugar
	Sugar	Reduction	Reduction
	Reduction	with	with
	with	PC	PC
	PureCircle	Alpha &	Alpha &
Dairy Formula	Alpha	PCS-1015	PCS-1015
2% Reduced	96.5803	96.5743	96.5753
fat Milk
Sugar	2.40	2.40	2.40
Cocoa Powder	0.80	0.80	0.80
10/12
Palsgaard 150	0.20	0.20	0.20
ChoMilk
PureCircle	0.0197	0.0197	0.0197
Alpha
PCS-1015		0.0070	0.0080
(MLD-1)
Total	100	100	100

Table 18 shows the sensory results with the two test samples. Both test samples showed the impact of the Stevia extract (PCS 1015) on the Chocolate flavor notes and dairy note. At 80 ppm use level, the chocolate milk sample showed better sweetness profile and overall liking than the control sample. FIG. 12 shows the comparison of the taste profile between the control and the test sample with 80 ppm Stevia extract PCS 1015.

TABLE 18
Summary of the overall acceptance and mean attribute
intensity results for each reduced sugar chocolate
milk samples tested by 30 panel members.
Summary of Mean-Scores, P-Values, and Significance
Test Result Code—chocolate milk with 70 ppm MLD-1
	197 ppm	70 ppm
	of	of
	Alpha	MLD-1
	Only	w/ PC
Attribute	(Control)	Alpha	P-Value	Sig
Sweet Intensity	8.85	8.89	0.8555	NS
Chocolate Flavor	6.82 b	7.70 a	0.0482	***
Dairy Note	3.61 b	4.19 a	0.1934	*
Bitterness	0.84	0.83	0.9500	NS
Bitter Aftertaste	0.74	0.70	0.6096	NS
Sweet Aftertaste	3.02	3.15	0.7232	NS
Overall Liking	7.12	7.42	0.5114	NS
Summary of Mean-Scores, P-Values, and Significance
Test Result Code—chocolate milk with 80 ppm MLD1
	197 ppm	80 ppm
	of	of
	Alpha	MLD-1
	only	w/ PC
Attribute	(Control)	Alpha	P-Value	Sig
Sweet Intensity	8.90 b	9.05 a	0.1557	*
Chocolate Flavor	6.89 b	7.53 a	0.0048	***
Dairy Note	4.12 b	4.44 a	0.1470	*
Bitterness	0.49	0.35	0.2473	NS
Bitter Aftertaste	0.71 a	0.55 b	0.1824	*
Sweet Aftertaste	2.66	2.82	0.5177	NS
Overall Liking	6.49 b	6.89 a	0.1908	*
* = 80% CI, ** = 90% CI, *** = 95% CI

Example 12: Effect of Stevia Extract on Desserts (Vanilla Custard)

To test the contribution of the Stevia extract, PCS-1015 in gelatin and puddings, two 30% calorie reduced vanilla custard samples were tested: 1) sweetened with PureCircle Alpha, a PureCircle Stevia sweetener, 2) sweetened with PureCircle Alpha and PCS-1015 (MLD-1). Table 19 shows the formulation of the control and test samples. A panel of 30 trained panelists with extensive experience in profiling sensory attributes tasted both samples.

To prepare the sample, blend the PureCircle Alpha and the test ingredient (PCS-1015) with the dry ingredients. Add the dry ingredients to the milk using good agitation. Heat on low until all ingredients are dissolved. Heat up to 95° C. for 10 minutes to cook up the starches. Add flavors, stir it, cool, stir it before place it in the refrigerator. Serve at chilled in 1 oz cups.

TABLE 19
Reduced sugar dessert (Vanilla Custard) with PCS-1015
	Control with	Test with PureCircle
	PureCircle Alpha	Alpha w/ stevia extract
Milk ( 1% fat)	94.27	94.27
Sucrose	4.00	4.00
Starch Perma Flo	1.25	1.25
Tate & Lyle
TIC Carrageenan	0.09	0.09
Salt	0.06	0.06
ROHA Beta Carotene	0.05	0.05
French Vanilla Flavor	0.15	0.15
UV 420-066-7
Steviol Glycoside	0.0166	0.0166
Stevia Extract	—	0.0080
Total	100	100

The trained panel found that the test sample had stronger sweet intensity, vanilla, dairy flavor notes and overall liking at 80% confidence. The sample containing Stevia extract also had significantly higher egg note at 95% confidence. FIG. 13 shows the pictorial rendition of the sensory difference between the control and test dessert samples

TABLE 20
Summary of the overall acceptance and mean attribute
intensity results for reduced sugar dessert
(Vanilla Custard) with PCS-1015
	166 ppm
	of	70 ppm
	Alpha	of
	Only	MLD-1
Attribute	(Control)	with Alpha	P-Value	Sig
Sweet Intensity	7.01 a	7.13 b	0.1095	*
Vanilla Flavor	3.22 a	3.5 b	0.1299	*
Egg Note	1.22 a	1.56 b	0.0497	***
Dairy / Creaminess	3.04 a	3.22 b	0.1164	*
Bitterness	0.43	0.5	0.3001	NS
Bitter Aftertaste	0.36	0.38	0.7692	NS
Sweet Aftertaste	2.23	2.24	0.8794	NS
Overall Liking	6.49 a	6.87 b	0.1149	*

Example 13: Effect of Stevia Extract on Flavor Modification of Chocolate-Flavored Beverage with Cocoa Powder Reduction

A chocolate flavored dairy beverage was developed to determine the flavor modification effect of Stevia extract flavor with modifying properties (FMP) in a dairy beverage. The two samples included: 1) control sample with full amounts of sugar and cocoa powder, and 2) test sample with 15% reduced sugar and 20% reduced cocoa, containing 60 ppm of Stevia extract FMP, as shown in Table 21.

TABLE 21
Reduced Sugar and Cocoa Chocolate-Flavored Beverage
(Chocolate Milk) with Stevia Extract
		Test with
	Control with	Reduced
	Full Amount	Sugar and
	of Sugar	Cocoa with
	and Cocoa	Stevia Extract
Milk, 2% milkfat	85.200	86.554
Sugar	8.00	6.80
Hot Water, 190° F.	6	6
Natural 10/12	0.80	0.64
Cocoa Powder
Stevia Extract	—	0.006
FMP (PSB-5005)
Total	100	100

A 15 member trained panel evaluated samples of chocolate milk for overall acceptance and attribute intensities (sweet intensity, bitterness, cocoa flavor, dairy note, sweet aftertaste and bitter aftertaste). The parameters for the sensory evaluation are shown in Table 22.

Table 23 shows the sensory results for the control and test products. The test product with 20% reduced cocoa powder and Stevia extract FMP shows no significant difference in cocoa flavor from the control. The test sample with 60 ppm Stevia extract FMP was higher in sweet intensity (90% confidence) and sweet aftertaste (directional). FIG. 14 illustrates this comparison.

TABLE 22
Sensory evaluation of chocolate flavored beverage
Nature of Participants: Trained panel
Number of Sessions      1
Number of Participants: 15
Test Design:            Balanced, randomized within set.
                        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, sweet intensity, bitterness, cocoa flavor, dairy note, sweet
aftertaste and bitter aftertaste. 10-pt continuous intensity scale where
0 = Imperceptible and 10 = Extremely Pronounced
Statistical Analysis:   ANOVA (by Block) with
                        Post Hoc Duncan's Test
Sample Size             ~1.5 oz. in a clear capped plastic cup
Serving Temperature     Refrigerated temperature (~45° F.)
Serving/Panelists       Samples served simultaneously.
Instruction:            Panelists instructed to evaluate
                        each sample.

TABLE 23
Sensory Results
		Test with
	Control	Reduced
	with Full	Sugar and
	Amount of	Cocoa with
	Sugar and	Stevia
Attribute	Cocoa	Extract FMP
Sweet intensity	7.05	7.26
Bitterness	0.65	0.55
Cocoa flavor	4.4	4.28
Dairy note	4.91	5.13
Sweet aftertaste	1.24	1.43
Bitter aftertaste	0.21	0.16
Overall liking	7.43	7.12

As seen in Table 23, the test product with reduced sugar and cocoa, and containing Stevia extract FMP, in this case PSB-5005, had statistically similar overall liking and mean cocoa flavor intensity results as compared to a full-cocoa formulation. The test product containing Stevia extract FMP had lower bitterness attribute and bitter aftertaste ratings compared to the control product made without Stevia extract. The dairy note was rated higher in the test product compared to the control product. From these results it can be seen that a reduction in cocoa and sugar content in a dairy beverage can be suitably accomplished using a Stevia extract FMP, such as PSB-5005, and unexpectedly with a decrease in bitterness which is typically associated with Stevia ingredients.

Example 14: Effect of Stevia Extract FMP on Flavor Modification of Vanilla-Flavored Dairy Product

A 50% sugar-reduced vanilla yogurt was developed to determine the flavor modification effect of Stevia extract flavor with modifying properties (FMP) in a reduced-sugar vanilla-flavored dairy product. The two samples as shown in Table 24 included: 1) control sample with 180 ppm steviol glycoside sweetener, and 2) test sample with 180 ppm steviol glycoside sweetener and 100 ppm of Stevia extract FMP.

TABLE 24
Sugar Reduced Vanilla-Flavored Dairy Product (Vanilla Yogurt)
		Test with
	Control	Reduced
	180 ppm	Sugar and
	steviol	Cocoa with
	glycoside	Stevia
	sweetener	Extract FMP
Plain nonfat yogurt	96.132	96.122
Sugar	3.750	3.750
Vanilla Flavor	0.100	0.100
Steviol Glycoside	0.018	0.018
Stevia Extract	—	0.010
(PCS 5001)
Total	100.000	100.000

A 30 member panel evaluated samples of vanilla yogurt for overall acceptance and attribute intensities (sweet intensity, bitterness, vanilla flavor, dairy, astringency, sweet aftertaste and bitter aftertaste). Table 25 lists the sensory evaluation parameters.

TABLE 25
Sensory evaluation of vanilla flavored dairy product
Nature of Participants: Trained sensory panel
Number of Sessions      1
Number of Participants: 30
Test Design:            Balanced, randomized within set.
                        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, sweet intensity, bitterness, vanilla flavor, dairy, astringency,
sweet aftertaste and bitter aftertaste. 10-pt continuous intensity scale where
0 = Imperceptible and 10 = Extremely Pronounced
Statistical Analysis:   ANOVA (by Block) with
                        Post Hoc Duncan's Test
Sample Size             ~1.5 oz. in a clear capped plastic cup
Serving Temperature     Refrigerated temperature (~45° F.)
Serving/Panelists       Samples served simultaneously.
Instruction:            Panelists instructed to evaluate
                        each sample.

Table 26 shows the sensory results for the control and test products. At 95% confidence, the test sample containing Stevia extract FMP was significantly higher for sweet intensity and vanilla flavor and significantly and unexpectedly lower in bitterness, astringency and sweet aftertaste. At 90% confidence, the test sample was higher in dairy and had higher overall liking. FIG. 15 illustrates this comparison.

TABLE 26
Summary of the overall acceptance and mean attribute
intensity results for vanilla-flavored dairy product
(vanilla yogurt) with stevia extract
		Test with
		180 ppm
		steviol
	Control	glycoside
	180 ppm	sweetener and
	steviol	100 ppm
	glycoside	stevia extract
Attribute	sweetener	FMP	p-value	Sig
Sweet intensity	6.88	7.05	0.0281	***
Bitterness	2.53	1.74	0.0374	***
Vanilla flavor	4.95	5.87	0.0452	***
Dairy	4.78	5.65	0.0507	**
Astringency	2.36	1.63	0.0407	***
Sweet aftertaste	1.73	0.95	0.0144	***
Bitter aftertaste	1.55	0.94	0.0682	**
Overall liking	5.26	6.37	0.0629	**

Example 15: Usage Levels of Stevia Extract FMP

Useful or maximum usage levels of Stevia extract FMPs were evaluated. To be useful as a flavor with modifying properties, the level of use of the Stevia extract must be below a certain sweetness detection threshold in a particular food or beverage product. To determine this threshold, a sensory evaluation is conducted with a full sugar product as the control, and a test product containing different levels of the Stevia extract FMP. Sensory panel members are then asked to identify which product is sweeter.

Using the Flavor and Extract Manufacturers' Association (FEMA) guidance document called “Guidance for the Sensory Testing of Flavorings with Modifying Properties within the FEMA GRAS™ Program, 2013”, the recognition threshold was determined using a 2-alternatve forced choice (2-AFC) methodology, as described in Table 27.

TABLE 27
Sensory evaluation to determine usage levels
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     Refrigerated temperature (~42° F.)
                        or room temperature, depending
                        on sample requirements
Serving/Panelists       Samples served simultaneously.
Instruction:            Panelists instructed to read ingredient
                        statement, evaluate each sample.

Usage levels for Stevia extract FMP are determined by those levels at which the Stevia extract FMP provides a sweetness perception that is significantly less than the full sugar control. For products other than baked goods and breakfast cereals, the sugar level in the control product was 1.5%. In baked goods, the sugar level in the control product was 4%, and in breakfast cereals, the sugar level in the control product was 3%. The test products contained no added sugar and contain various levels of Stevia extract FMP.

Table 28 shows usage levels of Stevia extract FMP in various food and beverage applications as determined using the FEMA sensory testing guidance.

TABLE 28
Usage levels of stevia extract FMP
Category                  Usage Level (ppm)
Baked Goods               500
Beverages, Non-Alcoholic  110
Beverages, Alcoholic      130
Breakfast Cereals         600
Chewing Gum               100
Condiments and Relishes   100
Confections and Frostings 100
Fats and Oils             180
Frozen Dairy              100
Fruit Ices                100
Gelatins and Puddings     100
Gravies                   100
Hard Candy                100
Imitation Dairy           165
Instant Coffee and Tea    200
Jams and Jellies          100
Milk Products             165
Nut Products              230
Processed Fruits          100
Processed Vegetables      100
Seasonings and Flavors    230
Snack Foods               230
Soft Candy                100
Soups                     100
Sugar Substitutes         100
Sweet Sauces              100

It was unexpectedly discovered that Stevia extract FMPs can be used at various levels to favorably impact the taste and flavor profile of a food or beverage product while having little or no detectable sweetness perception in that product. These usage levels serve as examples of use, and other usage levels of the Stevia extract FMP in various consumable products are contemplated by this invention.

Although various embodiments of the present invention have been disclosed here for purposes of illustration, it should be understood that a variety of changes, modifications and substitutions may be incorporated without departing from either the spirit or the scope of the invention.

Claims

1. A Stevia extract flavor with modifying properties (FMP) comprising: a. rebaudioside A at a level of up to 30% by weight;b. stevioside at a level of up to about 12% by weight; andc. minor steviol glycosides and plant molecules at a level between 30-55% by weight,wherein the Stevia extract FMP modifies the flavor and sweetness profile of a consumable product without imparting significant sweetness to the consumable product.

2. The Stevia extract FMP of claim 1, further comprising between 1-25% rebaudioside C by weight and between 1-20% rebaudioside D by weight.

3. The Stevia extract FMP of claim 1, further comprising rebaudioside E, wherein the rebaudioside E is present in an amount up to about 6% of the Stevia extract FMP by weight, rebaudioside N, wherein the rebaudioside N is present in an amount up to about 8% by weight, and rebaudioside O, wherein the rebaudioside O is present in an amount up to about 8% by weight.

4. The Stevia extract FMP of claim 1, wherein the minor steviol glycosides comprise one or more compounds selected from the group consisting of: rebaudioside G; rebaudioside H; rebaudioside I, rebaudioside J; rebaudioside K; and rebaudioside L.

5. The Stevia extract FMP of claim 1, wherein the plant glycosides further comprise a detectable amount of one or more compounds selected from the group consisting of: glucosyl-4′-O-apigenin; glucosyl-7-O-luteolin; rhamnosyl-3-O-kaempferol; glucosyl-3-O-quercitin; and arabinosyl-3-O-quercetin.

6. The Stevia extract FMP of claim 1, further comprising rebaudioside F, wherein the rebaudioside F is present in an amount up to 5% by weight.

7. The Stevia extract FMP of claim 1, further comprising rebaudioside B, wherein the rebaudioside B is present in an amount up to 8% by weight.

8. The Stevia extract FMP of claim 1, further comprising dulcoside A, wherein the dulcoside A is present in an amount up to 5% by weight.

9. The Stevia extract FMP of claim 1, further comprising rububoside, wherein the rububoside is present in an amount up to 5% by weight.

10. The Stevia extract FMP of claim 1, further comprising steviolbioside, wherein the steviolbioside is present in an amount up to 5% by weight.

11. A consumable product comprising a Stevia extract flavor with modifying properties (FMP) wherein the Stevia extract FMP is present at an amount below a Stevia extract FMP sweetness detection threshold level of about 120 ppm, wherein the Stevia extract FMP comprises rebaudioside A at a level of up to 30% by weight, stevioside at a level of up to about 12% by weight, and minor steviol glycosides and plant molecules at a level of up to 55% by weight.

12. The consumable product of claim 11, wherein the Stevia extract FMP further comprises between 1-25% rebaudioside C by weight and up to about 20% rebaudioside D by weight.

13. The consumable product of claim 11, wherein the Stevia extract FMP further comprises rebaudioside E, wherein the rebaudioside E is present in an amount up to about 6% of the Stevia extract FMP by weight, rebaudioside N, wherein the rebaudioside N is present in an amount up to about 8% by weight, and rebaudioside O, wherein the rebaudioside O is present in an amount up to about 8% by weight.

14. The consumable product of claim 11, wherein the Stevia extract FMP further comprises rebaudioside F, wherein the rebaudioside F is present in an amount up to 5% by weight.

15. The consumable product of claim 11, wherein the Stevia extract FMP further comprises rebaudioside B, wherein the rebaudioside B is present in an amount up to 8% by weight.

16. The consumable product of claim 11, wherein the Stevia extract FMP further comprises dulcoside A, wherein the dulcoside A is present in an amount up to 5% by weight.

17. The consumable product of claim 11, wherein the Stevia extract FMP further comprises rububoside, wherein the rububoside is present in an amount up to 5% by weight.

18. The consumable product of claim 11, wherein the Stevia extract FMP further comprises steviolbioside, wherein the steviolbioside is present in an amount up to 5% by weight.

19. A Stevia extract FMP comprising rebaudioside A, stevioside, and minor steviol glycosides and plant molecules, wherein; a. the ratio of rebaudioside A to stevioside is at least 5:6 and at most 10:1;b. the ratio of rebaudioside A to minor steviol glycosides and plant molecules is at least 1:5 and at most 1:1; andc. the ratio of stevioside to minor steviol glycosides and plant molecules is at least 3:55 and at most 2:5.

20. The consumable product of claim 11, wherein the minor steviol glycosides and plant molecules are present at a level between 30-50% by weight.

21. The consumable product of claim 12, wherein the rebaudioside D is present in an amount of between 1-20% by weight.