(LUẬN văn THẠC sĩ) optimization of some factors influencing lycopene extraction from tomato processing waste using response surface methodology

VIETNAM NATIONAL UNIVERSITY OF AGRICULTURE

NGUYEN THI KIM THANH

OPTIMIZATION OF SOME FACTORS INFLUENCING
LYCOPENE EXTRACTION FROM
TOMATO PROCESSING WASTE USING
RESPONSE SURFACE METHODOLOGY

Major:

Food Technology

Code:

24 18 05 57

Suppevisors:

1. Assoc. Prof. Tran Thi Dinh
2. Prof. Marie-Louise Scippo

AGRICULTURAL UNIVERSITY PRESS - 2017

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DECLARATION
I hereby declare that the data and results of research in my thesis are honest. There
is no material that has been accepted for the award of any other degrees or diploma in
any educational institution and, to the best of my knowledge and belief, it contains no

material previously published or written by another person, except where due reference
is made in the text of the thesis
I hereby declare that, all the help to carry out of my thesis was thanked and the
cited information in this thesis has been written clearly the source.

Hanoi, May 10th, 2017
Master candidate

Nguyen Thi Kim Thanh

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ACKNOWLEDGEMENTS
This thesis was realized at Department of Food Processing Technology and
Central laboratories of Food technology-Vietnam national university of Agriculture
under the supervisor of Assoc. Prof. Tran Thi Dinh and Prof. Marie-Louise Scippo. To
complete this thesis, besides the effort of myself, I have received encouragement and
great help of many individuals and groups.
Foremost, I would like express my deep gratitude to my supervisor Assoc. Prof.
Tran Thi Dinh and Prof. Marie-Louise Scippo for their valuable advices and continuous
guidance, encouragement and time sharing during my study. I would like to express my
sincere thanks to Msc. Nguyen Thi Hoang Lan and Dr. Hoang Hai Ha for enthusiasm,
insightful comments, teaching me on the HPLC analytical technique and useful
laboratory skills.
I am grateful to Research and Teaching Higher Education Academy – Committee
on Development Cooperation (ARES – CCD) for awarding the scholarship grant. I give
my thanks to Dr. Nguyen Thi Thanh Thuy for her great support during my study.

My sincere thanks are also sent to my friends especially, special thanks to my
juniors Than Thi Huong, Nguyen Thi Hien and Pham Thi Bich for their assistance in the
experimental work of this thesis.
Last but not least, I owe more than thanks to my family, my parents, my elder
sister and my younger brother for their love, support, patience and inspiration.
Hanoi, May 10th, 2017
Master candidate

Nguyen Thi Kim Thanh

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TABLE OF CONTENTS
Declaration ......................................................................................................................... i
Acknowledgements ........................................................................................................... ii
Table of contents .............................................................................................................. iii
List of abbreviations ......................................................................................................... v
List of tables..................................................................................................................... vi
List of figures .................................................................................................................. vii
List of figures .................................................................................................................. vii
Thesis abstract................................................................................................................ viii
Chapter 1. Introduction ................................................................................................. 1
1.1.

Introduction ........................................................................................................ 1

1.2.

AIM .................................................................................................................... 2

1.2.1.

General objective ................................................................................................ 2

1.2.2.

Specific objectives ............................................................................................... 2

Chapter 2. Literature review ......................................................................................... 3
2.1.

Tomato ................................................................................................................ 3

2.1.1.

Origin and distribution of tomato ....................................................................... 3

2.1.2.

Tomato composition ............................................................................................ 5

2.1.3.

Tomato processing waste .................................................................................... 6

2.2.

Lycopene ............................................................................................................ 7

2.2.1.

Source of lycopene .............................................................................................. 7

2.2.2.

Role of lycopene in the human health ............................................................... 10

2.2.3.

Physical and chemical properties of lycopene .................................................. 11

2.3.

Lycopene extraction ......................................................................................... 14

2.3.1.

Solvent extraction method ................................................................................. 15

2.3.2.

Other methods of lycopene extraction............................................................... 17

Chapter 3. Materials and methods .............................................................................. 20
3.1.

Materials ........................................................................................................... 20

3.1.1.

Sample collection and preparation .................................................................... 20

3.1.2.

Equipment ......................................................................................................... 20

3.1.3.

Chemical............................................................................................................ 21

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3.2.

Research contents ............................................................................................. 21

3.3.

Methodology..................................................................................................... 21

3.3.1.

Experimental design .......................................................................................... 21

3.3.2.

Analytical methods ............................................................................................ 25

3.3.3.

Data analysis ..................................................................................................... 28

Chapter 4. Results and discussion ............................................................................... 29
4.1.

Selection of the suitable organic solvent for lycopene extraction .................... 29

4.2.

Selection of treatment regime of tomato waste for lycopene extraction .......... 31

4.3.

Response surface methodology for optimization of lycopene extraction......... 36

Chapter 5. Conclusions and recommendations .......................................................... 42
5.1.

Conclusions ...................................................................................................... 42

5.2.

Recommendations ............................................................................................ 42

References ....................................................................................................................... 43
Appendix ......................................................................................................................... 48

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LIST OF ABBREVIATIONS
Abbreviation

Description

DPPH

1,1-diphenyl-2-picrylhydrazyl_C18H12N5O6

DW

Dry weight

HPLC

High performance liquid chromatography

w/v

Weight/ volume

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LIST OF TABLES
Table 2.1.

World tomato area, production and productivity, 2013 ............................... 3

Table 2.2.

World leading tomato producing countries in the world ............................. 4

Table 2.3.

Tomato area, production and productivity of some region
in Viet Nam, 2009 ........................................................................................ 4

Table 2.4.

Typical composition in 100 gram of a ripe tomato fruit .............................. 5

Table 2.5.

Carotenoid composition of tomato fruit, tomato processing wastes
and tomato paste (mg/100g wet sample) ..................................................... 7

Table 2.6.

Lycopene content of common fruit and vegetables ..................................... 9

Table 2.7.

Lycopene content in common tomato –based food ................................... 10

Table 2.8.

Physical properties of lycopene ................................................................. 11

Table 2.9.

Total lycopene and Cis-isomer content in the dehydrated tomato............. 14

Table 3.1.

Effect of solvent system on lycopene extraction from tomato waste ........ 21

Table 3.2.

Experimental design for drying of tomato waste ....................................... 22

Table 3.3.

Box- Behnken experimental design for lycopene extraction ..................... 23

Table 4.1.

Results of optimization treatment regimens for tomato waste .................. 31

Table 4.2.

Summary of effect of independent factors to the output variables ............ 32

Table 4.3.

Results of the analysis of variance on lycopene content ........................... 32

Table 4.4.

Result of the analysis of variance antioxidant capacity of
lycopene extract ......................................................................................... 34

Table 4.5.

Results of optimization condition for lycopene extraction ........................ 37

Table 4.6.

Summary of effect of independent factors to the output variables ............ 38

Table 4.7.

Results of the analysis of variance of lycopene content ............................ 38

Table 4.8.

Result of the analysis of variance of antioxidant capacity of
lycopene extract ......................................................................................... 39

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LIST OF FIGURES
Figure 2.1. Structure of trans and cis isomeric forms of lycopene .............................. 13
Figure 3.1. A. Tomato ‘Chanoka F1’ fruit, B. Fresh tomato waste,
C. Dried tomato waste ............................................................................... 20
Figure 3.2. HPLC chromatogram of (A) lycopene analytical standard at
0.25mg/ml and (B) CT5 sample in section 3.3.1.2 .................................... 26
Figure 3.3. HPLC calibration curve for lycopene standards dissolved in n-hexan
and dichloromethane (1:1) ......................................................................... 27
Figure 3.4. Trolox calibration curve............................................................................. 28
Figure 4.1. Effect of solvents systems on lycopene concentration .............................. 29
Figure 4.2. Effect of solvents systems on antioxidant capacity of lycopene extract.... 30
Figure 4.3. Profiler showing the optimal drying conditions of tomato waste .............. 35
Figure 4.4. Profiler showing the optimal extracting conditions of lycopene
extraction ................................................................................................... 40

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THESIS ABSTRACT
Master candidate: Nguyen Thi Kim Thanh
Thesis title: Optimization of some factors influencing lycopene extraction from tomato
processing waste using response surface methodology
Major: Food technology

Code: 24180557

Educational organization: Vietnam National University of Agriculture (VNUA)
Research Objectives: The aim of this research is to optimize some factor
(solvent/material ratio, temperature and time) influencing lycopene extraction process
from tomato waste which could be used to produce functional foods.
Materials and Methods:
- Materials: The red ripe tomato cv. Chanoka F1 was harvested in Bac Ninh
province. Tomato waste was obtained by removing the juice. Tomato paste was passed
through a fruit pulper to obtain waste. Tomato waste was dried by a convective oven
after that they were ground to use as material for lycopene extraction
- Methods: Suitable organic solvent for lycopene extraction from tomato waste
was studied ranging from single solvent (acetone, ethanol, ethyl acetate), double solvent
(acetone: ethanol) and triple solvent system (acetone: ethanol: ethyl acetate). The
treatment regime (moisture content and drying temperature) of tomato waste for
lycopene extract also investigated. Tomato waste, which was dried in the oven at the
optimal temperature and moisture content, was used to optimize of several factors (ratio
of solvent/dried tomato waste, temperature, time) influencing extraction of lycopene
with the most suitable solvent by response surface methodology.
- Analytical methods: Moisture content of tomato waste (%) was measured
using fast moisture detector (MA37, Germany). Lycopene content was quantified by
HPLC. Antioxidant capacity of lycopene content was quantified by DPPH radical
scavenging test.
Main findings and conclusions
The results of the present study indicated that ethyl acetate solvent proved to be
the most efficient compared to other solvents for lycopene extraction. The optimal
conditions for drying of tomato waste is temperature of 65oC until the moisture content
of the material reached 23%. The optimal extraction conditions for lycopene were:
+ Ratio of solvent/waste 40/1 (v/w),
+ Temperature 55oC and

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+ Extraction time 120 min.
Under this optimization condition lycopene content in the extract was 7.391 mg/g DW
and antioxidant capacity of extract was 10.384 µmol TE/g DW.

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CHAPTER 1. INTRODUCTION
1.1. INTRODUCTION
Lycopene is one of 750 carotenoids found in nature and is responsible for
the red color of fruits. It is present in high concentration in red fruit and
vegetable, such as tomato, gac, carrot, watermelon… (Britton, 2004). In the food
industry, lycopene is used as a natural pigment in the dyeing of food product.
Besides, lycopene is also known as a potential antioxidant which is believed to be
responsible for protecting cell against oxidative damage and thereby decreasing
the risk of chronic diseases (Rao et al., 2006). Thus, lycopene demands on using
in pharmaceutical, food, feed and cosmetic industries calls more attention
nowadays.
Tomato, Lycoperisicon esculentum, is one of the most widely cultivated
vegetable in worldwide and known as one of fruits which are rich in lycopene.
World tomato production in 2013 was about 163 million tons of fresh fruit from
an estimated 4.7 million hectares (Faostat, 2014). Tomato contains a wide variety

of antioxidants including vitamin E, ascorbic acid, carotenoids, flavonoids,
phenolic compounds (Sathish et al., 2009). Lycopene represents about 80-90% of
total carotenoids in tomato. Lycopene is located in different fractions of tomato
such as tomato skin, water insoluble fraction, and fibrous fraction including fiber
and soluble solids. Tomato processing industry produces large amounts of solid
waste. It is about 10–40% of the total tomato processed in the facility and
includes 33% seeds, 27% skin and 40% pulp (Topal et al., 2006). Toor and
Savage (2005) indicated that 70–90% of the lycopene was associated with the
water insoluble fraction and the skin. In Vietnam and other countries, the waste is
usually used for animal feed or for organic fertilizer but it is not used for human
consumption (Knoblich et al., 2005). Therefore, large quantity of carotenoids is
lost as waste. In addition, this waste has a high moisture content that makes it
susceptible to microbial proliferation and spoilage. Therefore, it can be preserved
by drying or other methods and then for lycopene extraction.
Tomato carotenoids are liposoluble. Recently, there are several methods
used for lycopene extraction. Sabio et al. (2003) studied a lycopene extraction
process based on supercritical CO2, which allows the extraction of over 60% of

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the lycopene from tomato waste. Xi (2006) reported that the lycopene yield from
high pressure processing treatment of tomato paste waste for 1 min was much
higher than from solvent extraction for 30 min. However, lycopene is commonly
extracted with organic solvents due to the cheap cost of technology and better
recovery as compared to other methods. There are a lot of organic solvents,
which are usually used in several studies to non-polar carotenoid extraction such
as ethyl acetate, ethanol, acetone, etc. However, their parameters

(solvent/material ratio, temperature and time) are largely influence on lycopene
extraction. Therefore in the current study, we conduct research entitled
“Optimization of some factors influencing lycopene extraction from tomato
processing waste using response surface methodology”.
1.2. AIM
1.2.1. General objective
The aim of this research is to optimize some factors (solvent/material ratio,
temperature and time) influencing lycopene extraction process from tomato
waste which could be used to produce functional foods.
1.2.2. Specific objectives
- To select suitable organic solvent for lycopene extraction process from
dried tomato waste;
- To optimize the moisture content and drying temperature by convective
drying of tomato waste for lycopene extraction;
- To optimize several factors (ration of solvent/material, temperature, time)
influencing on extraction of lycopene from dried tomato waste;
- To characterize the extracted lycopene in term of lycopene content and
anti-oxidant activity.

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CHAPTER 2. LITERATURE REVIEW
2.1. TOMATO
2.1.1. Origin and distribution of tomato
Tomato (Lycopersicon esculentum Mill.) is one of the most widely
cultivated vegetable worldwide. Tomatoes are members of the family Solanaceae
(also known as the nightshade family), genus Lycopersicon, subfamily

Solanoideae and tribe Solanceae (Taylor, 1986). It was originated in the coastal
highlands of Andean region that includes parts of Chile, Colombia, Ecuador,
Bolivia and Peru (Sims, 1979). The Spanish introduced tomato into Europe in the
early 16th century (Harvey et al., 2002). European acceptance of tomato as a
cultivated crop and its inclusion in the cuisine were relatively slow. Tomatoes
were initially grown only as ornamental plants: the fruits were considered to be
poisonous, because of the closely related deadly nightshade (Solanum
dulcamara). Since the mid-16th century tomatoes have been cultivated and
consumed in southern Europe, though they only became widespread in northwestern Europe by the end of the 18th century (Harvey et al., 2002). In 17th
century, European took the tomato to South, Southeast Asia and China. In the
18th century, tomato came to Japan and the USA. The production and
consumption of tomato expanded rapidly in the USA in the 19th century, and by
the end of that century, processed products such as soups, sauces and ketchup
were regularly consumed (Harvey et al., 2002).
Table 2.1. World tomato area, production and productivity, 2013
Location
Africa
America
Asia
Europe
Ocean
World

Area
(1000 ha)

Production
(1000 tons)

Productivity

(tons/ha)

902.16
455.84
2 821.82
500.87
7.64
4 688.34

18 118.82
24 264.84
99 205.50
20 965.20
554.36
163 108.72

20.08
53.23
35.16
41.86
72.56
34.79

Source: FAOSTAT (2014)

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Nowadays, tomatoes become the most important vegetable in the world.
According to Faostat (2014) tomato grows more than 175 countries around the
world. World tomato production in 2013 was about 163 million tons of fresh fruit
from an estimated 4.7 million hectares (Faostat, 2014) (Table 2.1). China leads
world tomato production with about 50 million tons with 30.1% of world
production followed by India with 18.2 million tons (11.1% global production)
(Table 2.2).
Table 2.2. World leading tomato producing countries in the world
No.

Country

Tomato production
(tons)

Share of world
production (%)

1
2
3
4
5

China
India
United States
Turkey
Egypt

50 552 200
18 227 000
12 574 550
11 820 000
8 533 803

30.10
11.10
7.70
7.20
5.20
Source: FAOSTAT (2014)

In Vietnam, tomato was first cultivated about 100 years ago, from the
French colonial period. Until now, tomato is still the main crop which prioritized
for development by United State. In the 2010, tomato is estimated to be grown on
more than 23,000ha with a production of nearly 460,000 tons per year (FAO,
2010).
Table 2.3. Tomato area, production and productivity of some region
in Viet Nam, 2009
Regions
Red River Delta
North Eastern
North Western
North Central
South Central
Tay Nguyen
South Eastern
Mekong River Delta

Area (ha)

Production (tons)

Productivity (tons/ha)

7 828
2 670
248
1 636
1 650
4 879
1 142
3 080

172 304
33 026
3 079
13 991
18 599
145 590
10 368
52 224

22.01
14.62
12.42
8.55
11.27
29.84

9.08
16.96
Source: FAO (2010)

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Tomatoes are mainly cultivated in the winter season. Besides, it is also
grown in the summer- autumn, autumn-winter, spring-summer season on the rice
land in order to bring high profit to farmers. Tomatoes are grown popularly in
Red Delta region (Table 2.3), concentrated in Ha Noi, Hai Duong, Hung Yen,
Bac Ninh… also in the Southern such as An Giang, Tien Giang, Lam Dong
provinces…) (Dang, 2014).
2.1.2. Tomato composition
Tomato is a kind of vegetable which had high nutrition value to human
diet and subsequent importance in human health. They are rich in minerals,
vitamines, essential amino acids, sugars and dietary fibers. Tomato contains
much vitamin B and C, iron and phosphorus (Ayandiji et al., 2011). The Table
2.4 gives the main nutrients and their quantities that can be derived from
consuming 100 gram of ripened tomatoes.
Table 2.4. Typical composition in 100 gram of a ripe tomato fruit
Nutrient

Unit

Amount

Water

Energy
Fat
Protein
Carbohydrates
Dietary fiber
Potassium
Phosphorus
Magnesium
Calcium
Vitamin C
Vitamin A
Vitamin E
Niacin
β- Carotene
α-carotene
Lycopene

G
Kcal
G
G
G
G
Mg
Mg
Mg
Mg
Mg
IU
Mg

Mg
µg
µg
µg

93.76
21.00
0.33
0.85
4.46
1.10
223.00
24.00
11.00
5.00
19.00
623.00
0.38
0.63
449.00
101.00
2573.00
Source: USDA nutrition database (2010)

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Tomatoes are widely known for their outstanding antioxidant content,

including their high concentration of lycopene and excellent amounts of other
conventional antioxidants like vitamin C and tocopherols, additional carotenoid
(β- carotene, lutein, and zeaxanthin) (Arab and Steck, 2000). According to Naika
et al. (2005), yellow tomatoes have higher vitamin A content than red tomato, but
red tomato contain lycopene, an anti-oxidant that may contribute to protection
against carcinogenic substances.
2.1.3. Tomato processing waste
According to FAOSTAT (2014), world tomato productions are huge about
163 million tons of fresh fruit in 2012 and on the rise in recent years. More than a
third of these were used for processing industry such as juice, soup, concentrate,
dry- concentrate, sauce, salsa, puree, dry-tomato, ketchup or paste (Kaur et al.,
2008). Commercial processing of tomato produces large amounts of solid waste
or by-products, namely tomato seeds and peels, representing 10-40% of total
processed tomato (Al-Wandawi et al., 1985; Topal et al., 2006).
The importance of utilization of the waste is no revenue from the sale and
dumping of this processing waste at the nearest landfill site will add to the
processing cost. On the other hand, if this waste remains unutilized, they not only
add to the disposal problem but also aggravate environmental pollution (AlWandawi et al., 1985). These wastes can be used for animal feed. Wet tomato
processing wastes can be ensiled with corn plants and the resulting silage
supported good milk production (Weiss, 1997).
One way of avoiding this problem would be to re-use the tomato
processing wastes to take advantage of the large quantity of potentially beneficial
compounds they contain. Tomato processing waste contain a variety of
biologically active substances being a promising source of dietary fibers,
proteins, carotenoids, tocopherols, polyphenols and other compounds (Vagi et
al., 2007). Among these bioactive compound polyphenol, carotenoid and
vitamins have a lot of physiological properties such as anti-inflammatory, antiallergenic, antimicrobial, anti-thrombotic, cardio-protective anti antioxidant
effects (Yang et al., 2008). The results of Al-Wandawi‘s (1985) research showed
that tomato skins yield about 71% of the lycopene found in tomato pastes.
Finally it is clear that a large quantity of nature color is normally disposed in

tomato processing “as waste” (Al-Wandawi et al., 1985). The Table 2.5 gives

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carotenoid composition of tomato fruit, tomato processing waste and tomato
paste that can be derived from consuming a 100 gram of wet sample (AlWandawi et al., 1985).
Table 2.5. Carotenoid composition of tomato fruit, tomato processing wastes
and tomato paste (mg/100g wet sample)
Carotenoid
phytoene
Phytofluene
β-carotene
Lycopene

Tomato processing wastes

Whole
mature fruit

Tomato skin

Tomato seed

Tomato
paste

Trace

Trace
0.13
3.35

Trace
Trace
0.30
11.98

0
0
0
0

2.29
1.86
1.06
16.79

Source: Al-Wandawi et al. (1985)

In recent years, a number of food scientists proposed that utilization of
tomato processing waste can be as source of lycopene for food, in order to
increase the intake of this carotenoid in the diet. Calvo et al. (2008) indicated that
tomato peel could be added to dry fermented sausages to produce a meat product
enriched in lycopene. Dry tomato peel was added to the meat mixture used in
sausage manufacture with ratio from 0.6 to 1.2% (w/w). After 21 days ripening,
lycopene level remained between 0.26 and 0.58 milligram per 100 gram of
sausage (Calvo et al, 2008). Garcia et al. (2009) also indicated that the direct
adding of dry tomato peel to hamburger could be useful both to obtain a new

product enriched in lycopene and to provide a use for this by-product from the
tomato industry. The addition of dry tomato peel to 4.5% w/w in hamburgers
with good overall acceptability and a lycopene content of 4.9 mg/100 g of cook
hamburger (Garcia et al. 2009). In addition, dry tomato waste is not only added
to meat product (sausage, hamburger, minced meat, dry-cured, beef patties and
frankfurters) but also add to bread to enrich in lycopene and other bioactive
compounds (Nour et al., 2015).
2.2. LYCOPENE
2.2.1. Source of lycopene
Lycopene belongs to carotenoid family that is a natural pigment synthesized
exclusively by plant and microorganisms. Lycopene is a natural pigment widely
used in the food industry as a food additive due to its strong color and non-

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toxicity. It is approved for food use with registering as 160d (i) - synthetic
lycopene, 160d (ii) – lycopene extract from tomato, 160d (iii) – lycopene
Blakeslea trispora (tracuuphugia.vfa.gov.vn).
 Chemical synthesis
Synthetic lycopene is the final product of a series of reactions, starting from
synthetic reagents and using chemical solvents. The final product often contains
trace of chemical solvents, impurities and by-products, which could be toxic and
very dangerous for health. This industrial production of synthetic lycopene has a
negative environmental impact due to high amount of chemical solvents utilized
(Lycopene, 2015).
Synthetic lycopene is highly concentrated, it has a purity of 90-95% and it
could not be directly used for human consumption. It is used for soaps, creams

and cosmetics. In fact, it has a very low bio-availability and is very labile to air
and light. The dietary supplements including this kind of lycopene are obtained
by diluting lycopene up to 1-10% with vegetable oils, preservatives and other
exogenous chemicals (Lycopene, 2015).
 Biological sources
Lycopene belongs to a group of naturally-occurring pigments known as
carotenoids. Lycopene is a natural constituent of red fruits and vegetables and of
certain algae and fungi (Olempska – Beer, 2005)
Microbial source:
Blakeslea trispora is a fungal plant pathogen. It is known as a source to
produce lycopene and is also the microbe used for producing commercial βcarotene for dietary supplements and food additives (Wikipedia: Blakeslea
trispora, 2016).
Lycopene from B. trispora is produced by mating and co-fermentation of
two non-pathogenic and non-toxigenic strains. Lycopene is extracted by
isopropanol and isobutyl acetate from the biomass and purified by filtration and
crystallization. Pure lycopene crystals are unstable when exposed to oxygen and
light so lycopene is stored under nitrogen or other inert gases in light-proof
containers. Olempska-Beer (2005) indicated that lycopene from B. trispora
contains at least 90% of all-trans-lycopene and minor quantities of 13-cislycopene and β- and γ-carotene.

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Plant sources:
Lycopene and other carotenoid are responsible for red color of many kinds
of fruit and vegetable in nature. Many fruit and vegetables are known to contain
lycopene such as tomato, watermelon, pink guava, pink grapefruit, papaya,
apricot and so on (Table 2.6).

Table 2.6. Lycopene content of common fruit and vegetables
Fruit/ vegetables

Lycopene (µg/g of weight)

Tomatoes
Watermelon
Pink guava
Pink grapefruit
Papaya
Apricot

8.8-42.0
23.0-72.0
54.0
33.6
20.0-53.0
<0.1
Source: Rao and Agarwal (1999)

Tomatoes, especially deep-red fresh tomato fruits, are considered the most
important source of lycopene in many human diets (Schwartz et al., 2002).
Lycopene can represent about 80-90% of total carotenoids in tomatoes. The
amount of lycopene in tomato fruits depend on variety, maturity, and the
environmental conditions under which the fruit matured (George, 2004). Most of
lycopene compound (70-90%) is located in the insoluble fraction of the tomato
such as peel. According to Al- Wandawi et al. (1985) tomato skin contains 12 mg
lycopene per 100 gram skin (wet basis), while whole mature tomato contain only
3.4 mg lycopene per 100 gram (wet basis). Skin tomato is a rich source of
lycopene, as they contain about five times more lycopene than the whole tomato

pulp (Sharma and Le Maguer, 1996).
Besides, the product was made from in tomato also contain large amount of
lycopene (Table 2.7). They are also mainly source of lycopene in human diets.

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Table 2.7. Lycopene content in common tomato –based food
Tomato product

Lycopene (µg/g dry matter)

Fresh tomato
Cooked tomato
Tomato sauce
Tomato paste
Tomato soup (condensed)
Tomato powder
Tomato juice
Pizza sauce
Ketchup

8.8 -42.0
37.0
62.0
54.0 - 1500.0
80.0
1126.3 -1264.9

50.0 – 116.0
127.1
99.0-134.4
Source: Rao and Agarwal (1999)

2.2.2. Role of lycopene in the human health
Human and animals cannot synthesize lycopene, and thus amount of
lycopene in their body depends on dietary sources (Shi and Magure, 2000). Of
more than 700 carotenoids in nature, there are 50 type may be absorbed and
metabolized by the human body. Only 14 carotenoids have been identified in
human serum, and lycopene is the most abundant (Xianquan et al., 2005).
Lycopene is particularly high concentration in the prostate gland, adrenal glands,
skin, liver and kidneys (Shi and Magure, 2000). Unlike other carotenoids,
lycopene cannot be converted into vitamin A (Rao and Agrawal, 1999). The
ability of lycopene to act as a potent antioxidant is thought to be responsible for
protecting cells against oxidative damage and thereby decreasing the risk of
chronic diseases (Rao and Agrawal, 1999).
Several recent studies have been shown that dietary intake of tomatoes
and tomato products containing lycopene associated with a decreased risk of
chronic diseases (Agrawal and Rao, 2000). Dorgan et al. (1998) also indicated
that serum and tissue lycopene levels have been found to be inversely related to
the incidence of several types of cancer, including breast cancer and bladder
cancer. Lower serum lycopene levels were found associated with increased risk
and mortality from coronary heart disease (Kristenson et al., 1997). Similarly,
Coodle et al. (1995) and Periquet et al. (1995) also reported that lower serum
lycopene levels were in human immunodeficiency virus (HIV) positive women
and also in children infected with HIV.

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Lycopene is not only reducing the risk of chronic diseases but also used in
cosmetic products. It is a common ingredient in anti-aging creams due to its
potent antioxidant. Lycopene can decrease inflammation and help to protect the
skin from damage resulting from UV sun exposure (Stahl et al., 2001).
2.2.3. Physical and chemical properties of lycopene
The molecular formula of lycopene is C40H56, and is an acyclic open-chain
polyene with 13 double bonds. There are 11 conjugated double bonds arranged in
a linear array, making it longer than any other carotenoid. The acyclic structure
of lycopene causes symmetrical planarity, and therefore lycopene has no vitamin
A activity. Lycopene is more soluble in chloroform, benzene, carbon disulfide,
acetone, ethyl acetate, and other organic solvents than in water (Shi and Maguer,
2000). The solubility of lycopene in vegetable oil is about 0.2 g/L at room
temperature (Borel et al., 1996). In aqueous systems, lycopene tends to aggregate
and to precipitate as crystals. This behavior is suspected to lower lycopene
bioavailability in humans. In ripe tomato fruit, lycopene takes the form of
elongated, needle like crystals that are responsible for the typical bright-red color
of ripe tomato fruits (Shi and Maguer, 2000).
The physical properties of lycopene are shown in Table 2.8.
Table 2.8. Physical properties of lycopene
Molecular formula

C40H56

Molecular weight

536.85 Da

Melting point

172-175oC

Crystal form

Long red needle from mixture of carbon disulfide and ethanol

Powder form

Dark reddish-brown

Solubility

Soluble in chloroform, hexane, benzene, carbon disulfide,
acetone, petroleum ether and ethyl acetate
Insoluble in water, ethanol, methanol

Sensitivity

Light, oxygen, high temperature, acids.
Source: Shi and Maguer (2000)

The chemical name of lycopene is 2,6,10,14,19,23,27,31-octamethyl2,6,8,10,12,14,16,18,20,22,24,26,30-dotriacontatridecaene.

Common

include Ψ, Ψ-carotene, all-trans-lycopene, and (all-E)-lycopene.

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names

Lycopene


occurs in all-trans-lycopene and various cis configuration. In literature source,
all-trans-lycopene is referred to as (all-E)-lycopene and cis isomers are referred
to as Z isomers (Olempska-Beer, 2005).
The all-trans isomer of lycopene is most predominant geometrical isomer in
fruits and vegetables, and is the most thermodynamically stable form. In red
tomato fruit, lycopene presents about 94-96% of all-trans-lycopene (Schierle et
al., 1997). In nature, lycopene exists in all-trans form and may be expected to
undergo changes into mono-cis or poly-cis forms under the influence of heat,
light, or certain chemical reactions (Cole and Kapur, 1957). The 5-cis, 9- cis, and
15-cis isomers of lycopene have presented in various tomato –based foods. In
human serum and tissue, cis isomers of lycopene contribute more than 50% of
total lycopene (Shi and Maguer, 2000). Cis isomers of lycopene have physical
characteristics and chemical behaviors distinct from those of their all-trans
counterparts, including decreased color intensity and lower melting points; they
are more polar than their all-trans counterpart, less prone to crystallization, and
more soluble in oil and hydrocarbon solvents. Experimental results revealed that
cis isomers of lycopene are better absorbed by human than the all-trans form
(Boileau et al., 2002). On the other hand, the conversion of cis- isomer to transform is another reaction that can occur during the product storage. Cis-isomers
are in the unstable, wherease trans-isomers are in the stable. The chemical
structure of lycopene isometrics in tomatoes are shown in Figure 2.1.

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Figure 2.1. Structure of trans and cis isomeric forms of lycopene
Source: Rao et al. (2006)

Lycopene has acyclic structure, large array of conjugated double and
extreme hydrophobicity. Therefore, lycopene exhibits many unique and distinct
biological properties. Although it has no pro-vitamin A activity, lycopene is able
to function as an antioxidant. Di Mascio et al. (1991) indicated that antioxidant
capacity of lycopene was more than double that of β-carotene and 10 times more
than that of α-tocopherol.

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2.3. LYCOPENE EXTRACTION
Lycopene is a pigment principally responsible for dark-red color of ripe
tomato fruit and tomato products (Montesano et al., 2008). Fresh tomato
industrial waste, which can be material for lycopene extraction, has high
moisture content about 82.9 % (Montesano et al., 2008). Drying process of the
waste may be affected lycopene content due to exposure of temperature, light and
oxygen. Favati et al. (2003) reported that the lycopene content of dried sample
was lower than that of fresh tomato industrial waste sample. The dried methods
of tomatoes make significantly increase in cis- isomers and simultaneously made
decrease in all-trans isomers (Table 2.8). Cis- isomers were not detected in the
fresh tomato sample after HPLC analysis (Rodriguez-Amaya and Tavares, 1992).

The highest amount of cis-isomers was found in air-dried tomato samples (Shi et
al., 1999).
Table 2.9. Total lycopene and Cis-isomer content in the dehydrated tomato
Sample

Total lycopene

Lycopene

All- trans

Cis- isomers

(µg/g dry basis)

loss (%)

isomers (%)

(%)

Fresh tomato

755

0

100

0

Osmotic treatment

755

0

100

0

Osmotic-vacuum dried

737

2.4

93.5

6.5

Vacuum- dried

731

3.2

89.9

10.1

Air –dried

726

3.9

84.4

16.6

Source: Shi and Le Mague (1999)

Lycopene is sensitive compound to light, oxygen, heat and acid.
Therefore, lycopene extraction from tomato waste has to be carried out under
controlled environmental factors to minimize lycopene degradation through
oxidation or isomerization. Actually, in the world, there are a lot of studies about
various methods, which are used for lycopene extraction from tomatoes such as
solvent extraction, supercritical fluid extraction using CO2, enzymatic aided
treatment… However, preferred method, which is used popularly, is solvent
extraction. The main reason for the adoption of solvent extraction method is the
cheaper cost of technology and better recovery as compared to other methods.

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2.3.1. Solvent extraction method
Lycopene is a fat-soluble substance therefore it is more commonly extracted

with single organic solvents, such as ethanol, acetone, ethyl acetate, petroleum
ether, hexane, benzene and so on or mixture of

various solvents. During

extraction process, whole extracted systems should be guaranteed safety to avoid
detonating combustion because the solvent has low boiling point, and easy to
volatile. Some main factor influencing lycopene extraction by solvent extraction
includes kinds of solvent, solvent/material ratio, temperature, extraction time,
particle size and number of extraction.
2.3.1.1. Solvent
The lycopene is used in production of healthy food, food additives,
medicines and cosmetics. One of the problems is the elimination of the residual
solvents to obtain a safe extract. Solvents should have not only low toxicity and
considered to be safe for foods but also low cost, high affinity toward the target
compound, stability during the extraction process, low detrimental effect on the
environment, low flammability (for safety). United Stated Department of Health
and Human Services (2012) has classified solvents into three categories. Among
of them, solvents belong to Class 3 such as acetone, ethanol, ethyl acetate,
propanol and propyl acetate are acceptable in small residual amount, and are used
in the food and pharmaceutical industries.
Acetone: is the organic compound with the formula (CH3)2CO. It is a
colorless, volatile, flammable liquid, and is the simplest ketone. Acetone can be
soluble in water and boils at 56.05oC (Allen et al., 1952).
Ethanol: also called alcohol, ethyl alcohol and drinking alcohol, is the
principal type of alcohol found in alcoholic beverages. It is a volatile flammable,
colorless liquid with a slight characteristic odor. Its chemical formula is C2H6O
or C2H5-OH. It boiling point is at 78.24oC.
Ethyl Acetate: is the organic compound with the formula C4H8O2. Ethyl
acetate is the ester of ethanol and acetic acid; it is manufactured on a large scale

for use as a solvent. Ethyl acetate is used primarily as a solvent and diluent, being
favored because of its low cost, low toxicity, and agreeable odor. In perfumes, it
evaporates quickly, leaving only the scent of the perfume on the skin. Ethyl
acetate is fairly volatile at room temperature and has a boiling point of 77oC.

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