Stevia Rebaudiana

Stevia is the generic term used for food ingredient derived from the herb stevia
rebaudiana. Because of high sweetener levels found in its leaves, stevia is commonly
called as sweet leaf or sugar leaf. Stevia has important industrial uses in food and
beverages, as well as used as medicine such as for low uric acid treatment, anesthetic
and anti-imflammatory. The extraction of stevia sweetener exert beneficial effect on
human health, including antioxidant, carcinogenic and anti-human rotavirus.

Before, there were several toxicological studies have been carried out to verify
the possible mutagenic and genotoxic effect of srevia extracts on bacterial cells and
mammalian species. The extract obtained from stevia leaves contain a complex mixture
of compounds, among them is glycosides such as stevioside and rebaudioside A. The
residual taste associated with stevia extracts is partially due to the glycoside and some
other compound such as terpenes (Nor, 2012)

Stevioside

Stevioside is a diterpene glycoside present in S. rebaudiana. A simple enzymatic

method is describes for the determination of stevioside from S.rebaudiana based on the
hydrolysis of stevioside with crude hesperidinase. The reaction is followed by
monitoring the production of glucose with a glucose oxidase-peroxidase-2 system.
Stevioside appear as a white, crystalline and odourless powder. Unlike many low-
calorie sweeteners, stevioside is stable at high (100°C) temperatures and over a range
of pH values (pH 3-9). It is contain no calories, and does not darken upon cooking. (Nor,
2012)
Solid-liquid extraction

According to solid-liquid extraction by (Afandi, A et al., 2013), the stevia leaves

are extracted with hot water or alcohols. In some cases, the leaves are pre-treated with
non polar solvents such as chloroform or hexane to remove the essential oils, lipids,
chlorophyll and other non polar substances. The extract is clarified by precipitation with

salt or alkaline solutions (Midmore and Rank, 2006). The extract is concentrated and
redissolved in methanol for crystallization of the glycosides. The crystals are formed
almost by pure stevioside.

Methanol gave improved (5.2%) extraction yield compared with water (4.7%)
when used in PHE for the isolation of stevioside from S. rebaudiana within the
temperature range of 110–160 °C. However, water represents a greener alternative to
methanol, therefore it can be a preferable solvent even with slightly lower yields (Puri.
M et al., 2012). Enzyme assisted extraction can be used to improve yields of stevioside
and also for improved extraction of a variety of bioactives.

Enzyme used in Enzymatic Assisted Extraction

Basically, there are four groups of plant enzymes, and each one of them is
responsible for breaking down a certain type of nutrient. Protease is responsible for
breaking down a protein, amylase can breaks down the sugar, while lipase works on the
fats, and cellulose helps to break down the carbohydrates. Typically, all whole foods
contain the necessary enzymes for the body to properly digest that particular food. The
enzymes that can degrade plant cell wall materials include cellulase, hemicellulases,
pectinase, chitinase and many ancillary enzymes.

Cellulases are part of a large group of glycosyl hydrolases that have been
categorized into several families on the basis of their amino acid homology.
Hemicellulases are able to degrade hemicelluloses, a class of polysaccharides that can
form hydrogen bonds with cellulose fibrils and form a network in plant cell walls.
Cellulase is an enzymatic protein that hydrolyzes the cellulose polymer to smaller
oligosaccharide, cellobiose and glucose. Cellulase randomly splits cellulose chains into
glucose. (Nor, 2012)
Enzyme assisted extraction

Principle and mechanism

Enzymes can be selected for specific functionalities as well as for

optimumprocess conditions, such as temperature and concentration. Although enzymes
normally function atan optimal temperature, they can still be used over a range of
temperatures, providing flexibility for both cost and product quality. Substrate particle
size reduction prior to enzymatic treatment provides better accessibility of the enzyme
to the cell to increase extraction yields significantly.

In enzyme-assisted aqueous extraction, the enzymes can rupture the

polysaccharide–protein colloid in the cell wallcreating an emulsion that interferes with
extraction. Therefore, non-aqueous systemsare preferable for some materials because
they minimize the formation of polysaccharide–protein colloid emulsions (Puri. M et
al., 2012).

Enzymatic pre-treatment has been considered as a novel and an effective way

to release bounded compounds and increase overall yield. The addition of specific
enzymes like cellulase, a-amylase, and pectinase during extraction enhances recovery
by breaking the cell wall and hydrolyzing the structural polysaccharides and lipid
bodies.
Procedure
Justification

The application of enzymes for complete extraction of bioactives without the

use of solvents is an attractive proposition. Enzyme pretreatment of raw material
normally results in a reduction in extraction time, minimizes usage of solvents and
provides increased yield and quality of product. Prior knowledge of the cell wall
composition of the raw materials helps in the selection of an enzyme or enzymes useful
for pretreatment. Decreased solvent use during extraction is particularly important for
both regulatory and environmental reasons, providing a ‘greener’ option than
traditional non-enzymatic extraction (Nor, 2012).

Solid-liquid extraction of stevioside from Stevia rebaudiana has incomplete

removal of interferences, low recovery of analytes, and high variability in results.
Quality of the stevioside produced from Stevia rebaudiana via enzyme assisted
extraction is higher than solid-liquid extraction. Efficiency of enzymes and the
specificity of their chemical activity in enzyme assisted extraction method allows
shorter extraction times. Therefore, enzyme assisted extraction is more preferred for the
extraction of stevioside from Stevia rebaudiana compared to solid liquid extraction.

Enzyme-assisted extraction of bioactive compounds from plants has potential

commercial and technical limitations: the cost of enzymes is relatively expensive for
processing large volumes of raw material (Puri. M et al., 2012). However, if the above
limitations can be overcome, then enzyme-based extraction could provide an
opportunity to not only increase extraction yields, but also to enhance product quality
by enabling the use of milder processing conditions such as lower extraction
temperatures.

Conclusion

In short, stevioside is a glycoside derived from the stevia plant, which can be
used as a sweetener. By comparing the effectiveness of solid-liquid extraction and
enzyme-assisted extraction methods, it can be concluded that enzyme assisted methods
shows more advantages than solid-liquid extraction method.
Reference

Puri, M., Sharma, D., & Barrow, C. J. (2012, January). Enzyme-assisted extraction of
bioactives from plants. Trends in Biotechnology.

Afandi, A., Sarijan, S., Ranajit, A., & Shaha, K. (2013). OPTIMIZATION OF
REBAUDIOSIDE A EXTRACTION FROM STEVIA REBAUDIANA
(BERTONI) AND QUANTIFICATION BY HIGH PERFOMANCE LIQUID
CHROMATOGRAPHY ANALYSIS. Journal of Tropical Resources and
Sustainable Science Universiti Malaysia Kelantan Publisher Journal of
Tropical Resources and Sustainable Science, 1(11), 2289–3946.

Nor, H. B. S. (2012, July). EXTRACTION OF STEVIOSIDE FROM STEVIA

REBAUDIANA LEAVES USING. Retrieved from UNIVERSITY MALAYSIA
PAHANG INSTITUTIONAL REPOSITORY:
http://umpir.ump.edu.my/7161/1/CD7144.pdf