2.

Chemical and physical Properties

In the following chapter the chemical properties of two main sweeteners Aspartame and Stevia will be
described. The main accent will be put on the stability of the compounds, because it is the most important
aspect in food industry. Foods and beverages are commonly produced at high temperatures and low pH.

2.1 Aspartame
Aspartame as a molecule is a dipeptide formed of two amino-acids: L-aspartic acid and
L-phenylalanine. Phenyl-alanine represents an essential amino-acid. These components exist in many
foods and are processed (metabolized) exactly in the same way, as they would come from meat, fish,
cheese, vegetables, milk, or fruit juice. [9] Aspartame’s taste profile is the closest one to sugar’s among
all other artificially made sweeteners and it is approximately 200 times sweeter than sugar.
The structural formula of aspartame is:

Figure 1: Functional groups of Aspartame [10,11]

Property name Property value

IUPAC Name (3S)-3-amino-4-[[(2S)-1-methoxy-1-oxo-3-
phenylpropan-2-yl]amino]-4-oxobutanoic acid
Solubility in water 44.1 ug/Ml
Molecular weight 294.307 g/mol
Taste Sweet
Melting point 245-247 °C
Odor Odorless
Dissociation Constants pKa1 = 2.96 (carboxyl); pKa2 = 7.30 (amine) (est)
Table 1: Chemical Properties of Aspartame[13]
Stability in different buffers of Aspartame
The stability of it varies depending on temperature, time, pH, and water activity [12]. Aspartame is less
stable in the liquid systems and its stability is primarily of a function of time, temperature, and pH. The
aspartame molecule hydrolyzes slowly at a low pH in order to produce a tasteless molecule methanol and
aspartyl-phenylalanine. Another route at pH=5 and above is that aspartame can cyclize to form its diketo-
piperazine with the elimination of methanol. These conversion products can be subsequently hydrolyzed
to its individual amino acids—aspartic acid and phenylalanine. (see figure 2)

Figure 1:Hydrolis of Aspartame[http://capellisquared.blogspot.com/2008/05/aspartame-kills-with-sweet-seduction.html]

A study was made by Yakici and Arici regarding the storage stability of Aspartame in Orange Flavored
soft drinks. The samples with different pH values (2.75;3.25;4,57) were kept in 20℃, 30℃ and 40℃ over
a 5 months’ period. The aspartame that remained was detected by using HPLC (High-pressure liquid
chromatography). They determined that aspartame is less stable at pH=2.75 and t=40℃. The graphs of
Aspartame’s degradation in different systems will be provided in the figures below. [15]

Figure 3: Storage stability at different temperatures, pH=2.75 [15]

 Figure 4: Storage stability at different temperatures, pH=3.25 [15]

Figure 5: Storage stability at different temperatures, pH=4.57 [15]

In dry products the stability of Aspartame is good. When used in its dry application the stability is quite
similar to that of the pure form of the compound. The degradation of aspartame in acidic solutions and at
high temperatures is accelerated by the ascorbic acid.

After providing and analyzing data from many sources, it can be concluded that aspartame is the most
stable at a pH=4.3. At elevated temperatures such as that found in short time pasteurization (80℃),
aspartame’s stability is decreasing, notably in the pH=6-7. It was also suggested that the degradation of
this molecule at ultrahigh temperature, can be ignored because the time involved is short for many changes
to take place. However, this view may be not correct, depending on the activation energy (Ea) for
decomposition processes.
2.2 Stevia
Stevia is a genus of about 200 species of herbs and shrubs in the Asteraceae family (sun-flower family)[1].
It can grow up to 1 m tall. The plant is a perennial herb with a root system that is extensive and brittle
stems producing small, elliptic leaves. [16] The leaf is formed of eight terpene glycosides identified as
Stevioside, Rebaudioside A, B, C, D, and E, dulcoside A and C. The sweetness of it is due the presence
of three glucose groups. In spite of the fact that the Stevioside is the major component among the diterpene
glycosides, its presence in high levels creates a post digestive bitter taste, this being a main reason for
which Stevia is rejected by consumers. [61]

The extracts from Stevia Rebaudiana have different chemical structure. Extracts from Stevia Rebaudiana
consists of 5-10 percent stevioside,2-4 percent rebaudioside A. 1-2 percent is for rebaudioside C and other
glycosides like rebaudiosides B, D, E; dulcoside A and steviolbioside. Some studies suggest that
rebaudioside B and steviolbioside are not constituents of Stevia, but are provided as a result of partial
hydrolysis.[63]

Figure 6: All the Steviol Glycosides [64]

Property name Property value
IUPAC Name 13-[(2-O-β-D-glucopyranosyl-3-O-β-D-
glucopyranosyl-β-D-glucopyranosyl)oxy]-ent-
kaur-16-en-19-oic acid β-D-glucopyranosyl
ester
Solubility in water 0.365 g/ml
Molecular weight 967.021 g/mol
Taste Sweet
Water holding capacity 4.7 ml/g
Odor Odorless
pH of solution 5.95
Energy value 2.7 kcal/g
Table 2: Chemical properties of Stevioside [19]

Stability in different buffers solutions of Steviol Glycosides

The stability of the two most present steviol glycosides rebaudioside A and stevioside and the possible
formation of the aglycones (the compound remaining after the glycosyl group on a glycoside is replaced
by a hydrogen atom) will be provided below. [65] Little data is available on topic of practical use in foods
and beverages, and we notice an absence of detailed information about steviosides stability, reactions with
other food-ingredients and also about storage conditions. Good stability of stevioside has been provided
in water solutions and in the soy sauce at 30 Celsius degrees for 30 days, but at the same time some losses
were discovered in vinegar at 80 Celsius Degrees. [66]
In the long-term tests of storage of these two most found glycosides in citric and phosphoric acidified
drinks no changes were noticed at 4 Celsius degrees and at the room temperature for 3 months, while some
modifications occurred at 37 Celsius degrees with 17.1 percent loss after 4 months of being stored in the
citric acidified beverage and a loss of 36 percent in the phosphoric acidified beverage. As degradation
products glucose, steviolbioside and some else non-analyzed substances were found. [67] Rebaudioside
A did not show significant changes during 4 months at 4 Celsius degrees.,1 month at 37 degrees or 3
months at 22.5 Celsius degrees in either phosphoric and citric acidified beverage. But after 4 months at
37 degrees a 13 percent loss was detected in the citric acid system and ¼ loss in the phosphoric acid
beverage. [68] Using data provided by Chang and Cook (1983) a kinetic plot can be constructed. The
graph below shows the reducing amount of Rebaudioside A under certain conditions: pH=2.6; t=100°C
(pseudo-first order react.).
 Figure 7: Degradation of rebaudioside A at pH 2.6; t=100 ℃ modeled pseudo-first
order [63,69] Figure 2:

In 2008, Clos et. Alumni questioned the stability under light of Rebaudioside A presented before by Chang
and Cook in 1983 and redid their experience. Samples were stored under identic condition and therefore
it was concluded that chemically that Rebaudioside A is not affected by light. Anyway, their experiment
actually shows 5 times bigger loss when stored under light than in full darkness. [70]

Furthermore, the stability of Stevioside under different conditions will be analyzed. In solutions it is
remarkable stable over a big range of pH and temperature. After thermic treatment at 60 degrees up to 137
degrees’ significant degradation was not observed in acidic solutions. This was equally true for the citric
and phosphoric acidified systems. Thin-layer chromatography (TLC) was used to identify if new chemical
structures appeared in the solutions. TLC technique is often used to separate non-volatile mixtures, being
done on a sheet of plastic, glass or aluminum foil which is covered with a thin layer of adsorbent material
like silica-gel or cellulose.

Figure 8: TLC of Stevia Extracts [71]

The stability of stevioside at different temperatures was analyzed, and a graph was constructed using the
data:

Figure 9: Stability and degradation rate of Stevioside as solid substance at elevated

temperatures [68]

In comparison with aspartame stevia is far more stable. Stevia is generally stable to heating so the
sweetening power is not lost during processing [20]. For instance, High temperature short time (HTST)
and ultrahigh temperature processing (UHT) are used for many applications such as iced tea, milk and
juices. Process testing done evaluated a sweetener comprised of many Steviol glycosides at two different
conditions commonly viewed in beverage processing: pH of 3.2 at 88 °C for 0.5 minutes (HTST) and pH
of 6.5 at 138 °C for 10 seconds (UHT). Almost no loss is observed in the quantity of glycosides. [21]

Figure 10: HTST and UHT treatments of Steviol Glycosides[22]