Enzyme Assisted Extraction of Carotenoids from Tomatoes Comparison between classical method and new enzymatic method. Enzymes used: Carenzyme and Pectinex

1. Enzyme-Assisted
Extraction of
Carotenoids From
Tomatoes
By Manali Parab
M.Pharmacy First Year

2. Introduction to extraction of carotenoids
 Classical method: “Organic Solvent Procedure”
 Solvent used: petroleum, benzene, ethyl ether and
methanol
 Low yield- Activity of carotenoid is affected
 New method: Enzymatic Extraction
 The cell-wall degrading, hydrolytic enzymes such as
cellulases, hemicellulases and pectinases can penetrate
the complex tissue matrix of polysaccharides and
facilitate the removal of carotenoids
 Advantages: faster extraction, higher recovery, reduced
solvent usage and lower energy consumption

3. Carotenoids in tomatoes
 Lycopene is obtained from the tomato species, is a bright
red carotene and carotenoid pigment and phytochemical
found in tomatoes and other red fruits and vegetables,
such as red carrots, watermelons, and papayas.
 In plants, algae, and other photosynthetic organisms,
lycopene is an important intermediate in the biosynthesis
of many carotenoids, including beta-carotene, which is
responsible for yellow, orange, or red pigmentation,
photosynthesis, and photoprotection.

4. Structure of Lycopene

5. Enzymes Used:
 Enzyme-assisted extraction using comparatively
commercial and crude pectinases and cellulases obtained
from fermented fractions of Penicillium oxalicum and
Trichoderma reesei, respectively.
 Different ratios between the enzyme activity and
substrate (1, 5 and 10%) were used when incubated with
the tomato homogenate.

6. Extraction of Carotenoids from tomatoes
Tomatoes are washed and
homogenized, stirred
aliquots of 1 g each in
tightly-closed glass
tubes at -20°C.
Sample Preparation

7. EXTRACTION OF CARETENOIDS
To each aliquot of 1 g homogenized tomato sample, the
enzymes were added at pH 4.3 in each tube
Vortexed for two minutes & tubes were incubated at
40°C for 2 hours
Solvent Addition: hexane: acetone: ethanol 50:25:25
(v/v) and BHT (0.05% w/v in acetone) as antioxidant
Followed by two times extraction
Three concentrations pf enzymes were used: 1, 5 & 10% depending on
specific activity

8. The filtrate was partitioned in a separation
funnel, successively with water, diethyl ether
and saturated saline solution
The upper organic phase was evaporated at
35°C to dryness under vacuum, using a rotary
evaporator, and a nitrogen stream
The residue was redissolved in 0.5 ml of ethyl
acetate by mixing 2 min
Carotenoids were separated, identified and
quantified by HPLC.

9. Amount of carotenoids obtained by
Carenzyme and Pectinex
Carenzyme Amount of carotenoids (mg/kg FW)
obtained
1% 1.48
5% 2.25
10% 3.11
Pectinex Amount of carotenoids (mg/kg FW)
obtained
1% 1.54
5% 2.41
10% 2.17

10. Conclusion
 Optimal concentration of enzymes used for carotenoids release
were the crude cellulases at 10% followed by purified,
commercial cellulases at 10%.
 The crude pectinases showed higher efficiency, at 10%,
compared with the commercial ones, especially in the
extraction of phenolic compounds, improving the antioxidant
capacity of the hydrolysed extract.
 Beside the enhancement of release yield, the use of crude
enzymes prevents the use of high quantities of organic solvents
and assures the increase of antioxidant capacity of such
hydrolysed extracts.

11. REFERENCE
 Enzyme-Assisted Extraction of Carotenoids and Phenolic Derivatives from
Tomatoes, Bulletin UASVM Animal Science and Biotechnologies 71(1) / 2014,
20-26, By Diana NEAGU1), Loredana F. LEOPOLD1,3), Phillipe THONART2),
Jacqueline DESTAIN2), Carmen SOCACIU1,3), From 1)University of Agricultural
Sciences and Veterinary Medicine, Faculty of Food Science and Technology,3-5
Mănăştur Street, 400372 Cluj-Napoca, Romania;2)Gembloux Agro-Bio-Tech,
Université de Liège, Belgium: 3)CCD-BIODIATECH, Proplanta Centre, Cluj-
Napoca, Romania