Thesis for the Degree of Master in Science
with a major in Textile Engineering
The Swedish School of Textiles
2014-06-26
Report no. 2014.14.04
Extraction of
β-carotene from orange
peel and carrot waste
for cotton dyeing
Susan Hecker
Visiting adress: Skaraborgsvägen 3  Postal adress: 501 90 Borås  Website: www.hb.se/ths
II
Master thesis by Susan Hecker
Description: Thesis submitted for the degree of Master in Science in Textile
Engineering
Title: Extraction of β-carotene from orange peel and carrot waste for cotton dyeing
Author: Susan Hecker
Supervisor: Mats Johansson, Vincent Nierstrasz
Cooperation partners:
Brämhults Juice AB, School of Engineering at the
University Borås, Almedahls
Examiner: Anders Persson
The Swedish School of Textiles Report no. 2014.14.04
Master thesis by Susan Hecker
III
Acknowledgement
At the School of Textiles in Borås I would like to thank Maria Björklund and Catrin
Tammjärv who helped me and had always an open ear for all my questions and thoughts.
For the good feedbacks and discussions I want to thank my supervisors Vincent Nierstrasz
and Mats Johansson. Special thanks for the help with the HPLC analysis to Jorge Ferreira
at the Engineer school at the University of Borås.
Thanks to Stina Haglund at Brämhults Juice AB for the allocation of the orange peel and
carrot residue.
Anna Frisk and Karin Eklund at Almedahls made it possible that I could make the Xenon
test there.
And of course I would like to thank my family and friends who supported me during my
whole studies and especially during my master thesis work.
Borås, June 2014
Susan Hecker
IV
Master thesis by Susan Hecker
Abstract
The further usage of vegetable and plant waste from juice pressing industry as textile dyes is
presented in this thesis. The thesis is focused on β-carotene (C40H56) dyestuff extracted from
orange peel and carrot residue. The three organic solvents; ethyl acetate, petroleum ether
and hexane/acetone (1:1 v/v) were used for the extraction. The analysis of the extract was
done by RP-HPLC with a C18 column. The yield and the purity of the extracted β-carotene
were determined. The highest yield was achieved with petroleum ether whereas the other
two solvents were nearly as good. The highest and purest amount on β-carotene was found
in the extracts of carrots. The dyeing process was continued with β-carotene dyestuff of
orange peel and carrot residue extracted from 27 g of residue on 0,8 g cotton in the dyeing
ratio 1:50. Unmordant and post mordanted bleached and mercerized cotton fabric was dyed.
10% alum of the weight fraction of cotton was used as mordant. Colour measurements (K/S,
C*, L*, a*, b*, h and ΔE) and fastness properties as light- (ISO-Norm B02) and wash fastness
(ISO 105 – C) were tested. Fair light fastness grades were achieved by β-carotene dyestuff
of orange peel residue dyed on cotton fabric. Poorer were the grades for β-carotene dyestuff
of carrot residue for both unmordanted and mordanted samples. The wash fastness couldn’t
achieve reasonable results neither for β-carotene dyestuff from carrots nor orange peel
residue, dyed on cotton fabric.
Keywords:
β-carotene, carrot and orange peel residue, organic solvent extraction, cotton
dyeing
Master thesis by Susan Hecker
V
Popular abstract
Brämhults Juice AB a juice pressing company in Borås, Sweden has every year 8100 tons of
orange peel waste and 570 tons of carrots waste from juice pressing that still contains a lot of
natural resources, such pigments and colorants. Using these pigments and colorants further
would increase its efficiency. A market for the further usage would be the food or textile
dyeing industry. The main colorant in carrots and orange peel residues is β-carotene
(C40H56). β-carotene is a water insoluble yellow-orange pigment. Nowadays it is used in the
food industry as food colorant and pro vitamin A precursor. In this paper the further usage of
dying cotton fabric is investigated. Due to solvent extraction β-carotene can be obtained in a
quite pure form from carrots residue and in less pure form from orange peel residue. The
dyed cotton fabric is red-orange for carrot extract. The orange peel extract contains besides
β-carotene also other carotenes; flavonoids, phenolic acids, pectin and waxes. The dyed
cotton is light yellow-orange. The fastness properties to stand washing and light exposure
were tested. The light fastness properties got fair results for β-carotene dyestuff of orange
peel residue dyed on cotton fabric and less fair results for the carrot extract. The wash
fastness properties couldn’t achieve reasonable results.
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Master thesis by Susan Hecker
Content List
ACKNOWLEDGEMENT
IV
ABSTRACT
V
POPULAR ABSTRACT
VI
TABLE LIST
IX
FIGURE LIST
X
LIST OF ABBREVIATIONS
XI
1.
INTRODUCTION
2
2.
LITERATURE REVIEW
4
2.1.
2.2.
2.3.
2.4.
2.5.
2.6.
3.
3.1.
3.2.
4.
Β-CAROTENE
CARROTS AND ORANGE PEEL
EXTRACTION METHODS AND SOLVENT PROPERTIES
DYEING WITH NATURAL SOURCES
MORDANTS
DYE AND DYEING METHOD
PROBLEM DESCRIPTION
AIM
RESEARCH QUESTION
MATERIALS AND METHODS
4
7
8
10
11
12
14
14
14
16
4.1. MATERIALS
4.2. DRYING OF THE CARROT AND ORANGE PEEL RESIDUE
4.3. EXTRACTION METHOD
4.4. ANALYSIS OF THE DYESTUFF, PURITY AND YIELD
4.5. COTTON DYEING
4.6. SPECTROPHOTOMETRIC MEASUREMENTS
4.7. FASTNESS PROPERTIES TESTS
4.7.1. XENON TEST (ISO-NORM B02)
4.7.2. WASH FASTNESS TEST (ISO 105 – C)
16
17
17
18
18
20
21
21
21
5.
22
5.1.
5.2.
5.3.
5.4.
5.5.
5.6.
RESULTS AND DISCUSSION
DETERMINATION OF Β-CAROTENE SPECTRUM
RESULTS OF THE RP-HPLC ANALYSIS
MORDANTING AND DYEING
SPECTROPHOTOMETRIC COLOUR MEASUREMENTS
RESULTS OF THE WASH FASTNESS TEST (ISO 105 – C)
RESULTS OF THE XENON TEST (ISO-NORM B02)
Master thesis by Susan Hecker
22
23
26
28
30
31
VII
6.
CONCLUSIONS
33
7.
FUTURE RESEARCH
34
REFERENCES
APPENDIX
APPENDIX I
AQUEOUS EXTRACTION
ORGANIC SOLVENTS EXTRACTION WITH ETHANOL
RESULTS OF THE HPLC ANALYSIS
APPENDIX II
CHEMICAL STRUCTURES OF THE SOLVENTS USED
APPENDIX III
VAT DYE
APPENDIX IV
HPLC RESULTS
APPENDIX V
SPECTROPHOTOMETRIC AND FASTNESS PROPERTY RESULTS
VIII
Master thesis by Susan Hecker
35
I
II
II
III
IV
VI
VI
VII
VII
VIII
VIII
XII
XII
Table List
TABLE 1: TOTAL CAROTENOIDS AND THE CONTENT OF DIFFERENT CAROTENE AND XANTHOPHYLLS IN CARROTS
AND ORANGE PEEL IN MG/G CARROTS/ORANGE PEEL ............................................................................ 7
TABLE 2: SOLVENT PROPERTIES OF THE USED SOLVENTS ............................................................................ 9
TABLE 3: AMOUNTS OF THE DRIED RESIDUES (G) AND SOLVENTS (ML) THAT WERE USED FOR THE HPLC
SAMPLES AND THE DYESTUFF EXTRACTION FOR THE COTTON DYEING.................................................. 17
TABLE 4: COMPOSITION OF THE USED Β-CAROTENE DYESTUFF OF CARROT AND ORANGE PEEL RESIDUE THAT
WAS USED FOR THE DYEING OF 0,8 G COTTON FABRIC. ....................................................................... 19
TABLE 5: FINAL DYE RECEIPT. AMOUNT PER LITRE WATER AND THE CALCULATED AMOUNT THAT WAS USED FOR
THE DYEING OF 0,8 G COTTON FABRIC IS LISTED. ............................................................................... 20
TABLE 6: THREE DIFFERENT CONCENTRATIONS USED FOR THE HPLC-STANDARD CURVE ............................ 23
TABLE 7: Β-CAROTENE CONTENT IN CARROTS AND ORANGE PEEL (MG/G OF CARROTS OR ORANGE PEEL)...... 25
TABLE 8: COLOUR COORDINATION OF THE DYED COTTON FABRICS WITH CARROT EXTRACT .......................... 28
TABLE 9: COLOUR COORDINATION OF THE DYED COTTON FABRICS WITH ORANGE PEEL EXTRACT ................. 29
TABLE 10: RESULTS OF THE WASH FASTNESS TEST OF UNMORDANTED AND POST MORDANTED (ALUM) DYED
COTTON FABRICS OF ORANGE PEEL AND CARROT EXTRACT................................................................. 30
TABLE 11: RESULTS OF THE XENON LIGHT FASTNESS TEST OF UNMORDANTED AND POST MORDANTED (ALUM)
DYED COTTON FABRICS OF ORANGE PEEL AND CARROT EXTRACT. ....................................................... 31
TABLE 12: Β-CAROTENE CONTENT IN CARROTS AND ORANGE PEEL (ΜG/G) ................................................... IV
TABLE 13: DIFFERENT PARAMETERS FOR VAT DYEING ON COTTON FABRIC .................................................. VII
TABLE 14: RESULTS OF THE DETERMINATION OF Β-CAROTENE BY HPLC ................................................... VIII
TABLE 15: COLOUR COORDINATION OF THE ALL DYED COTTON FABRICS WITH CARROT EXTRACT .................. XII
TABLE 16: COLOUR COORDINATION OF THE ALL DYED COTTON FABRICS WITH ORANGE PEEL EXTRACT ......... XII
TABLE 17: RESULTS OF THE WASH FASTNESS TEST OF ALL UNMORDANTED AND POST MORDANTED (ALUM)
DYED COTTON FABRICS OF ORANGE PEEL AND CARROT EXTRACT. ...................................................... XIII
TABLE 18: RESULTS OF THE LIGHT FASTNESS TEST OF ALL UNMORDANTED AND POST MORDANTED (ALUM)
DYED COTTON FABRICS OF ORANGE PEEL AND CARROT EXTRACT. ...................................................... XIII
Master thesis by Susan Hecker
IX
Figure List
FIGURE 1: FRUITS AND VEGETABLE WASTE FROM JUICE PRESSING OF BRÄMHULTS JUICE AB IN TONS PER
YEAR. ................................................................................................................................................ 2
FIGURE 2: W ASTE HANDLING OF BRÄMHULTS JUICE AB. W ASTE IN TONS PER YEAR. ..................................... 3
FIGURE 3: ALL-TRANS Β-CAROTENE ............................................................................................................ 4
FIGURE 4: CAROTENOIDS CLASSIFICATIONS ................................................................................................ 5
FIGURE 5: Β-CAROTENE SYNTHESIS BY W ITTIG REACTION ............................................................................ 6
FIGURE 6: 9-CIS-Β-CAROTENE .................................................................................................................... 6
FIGURE 7: DESICCATOR SILICA UNDER VACUUM CONTAINING ORANGE PEEL AND CARROT PIECES ................. 17
FIGURE 8: GRINDING DRIED ORANGE PEEL AND CARROTS IN A MORTAR....................................................... 17
FIGURE 9: EXTRACTION FROM CARROTS IN PETROLEUM ETHER .................................................................. 17
FIGURE 10: GRAPH OF THE DETERMINED WAVELENGTH OF Β-CAROTENE AT 450 NM. ................................... 22
FIGURE 11: HPLC-STANDARD CURVE OF Β-CAROTENE .............................................................................. 23
FIGURE 12: HPLC GRAPH AT 450 NM SHOWING Β-CAROTENE, Α-CAROTENE AND ZEAXANTHIN EXTRACTED
FROM CARROT RESIDUE.................................................................................................................... 24
FIGURE 13: HPLC GRAPH AT 450 NM SHOWING Β-CAROTENE, Α-CAROTENE AND ZEAXANTHIN EXTRACTED
FROM ORANGE PEEL RESIDUE ........................................................................................................... 24
FIGURE 14: Β-CAROTENE DYESTUFF OF CARROT RESIDUE, DYED ON COTTON, MORDANTED AND UNMORDANTED
....................................................................................................................................................... 27
FIGURE 15: Β-CAROTENE DYESTUFF OF ORANGE PEEL RESIDUE, DYED ON COTTON, MORDANTED AND
UNMORDANTED ................................................................................................................................ 27
FIGURE 16: HPLC FROM CARROT EXTRACT IN AQUEOUS SOLUTION AT 450 NM ............................................ IV
FIGURE 17: HPLC FROM ORANGE PEEL EXTRACT IN AQUEOUS SOLUTION AT 450 NM.................................... IV
FIGURE 18: HPLC FROM CARROT EXTRACT IN 80 % AQUEOUS ETHANOL SOLUTION AT 450 NM ..................... IV
FIGURE 19: HPLC FROM ORANGE PEEL IN 80 % AQUEOUS ETHANOL SOLUTION AT 450 NM............................ V
FIGURE 20: HPLC GRAPH OF SAMPLE 1 Β-CAROTENE FROM ORANGE PEEL, EXTRACTED IN ETHYL ACETATE VIII
FIGURE 21: HPLC GRAPH OF SAMPLE 7 Β-CAROTENE FROM ORANGE PEEL, EXTRACTED IN ETHYL ACETATE .. IX
FIGURE 22: HPLC GRAPH OF SAMPLE 3 Β-CAROTENE FROM ORANGE PEEL, EXTRACTED IN HEXANE/ACETONE
........................................................................................................................................................ IX
FIGURE 23: HPLC GRAPH OF SAMPLE 9 Β-CAROTENE FROM ORANGE PEEL, EXTRACTED IN HEXANE/ACETONE
........................................................................................................................................................ IX
FIGURE 24: HPLC GRAPH OF SAMPLE 2 Β-CAROTENE FROM ORANGE PEEL, EXTRACTED IN PETROLEUM ETHER
........................................................................................................................................................ IX
FIGURE 25: HPLC GRAPH OF SAMPLE 8 Β-CAROTENE FROM ORANGE PEEL, EXTRACTED IN PETROLEUM ETHER
......................................................................................................................................................... X
FIGURE 26: HPLC GRAPH OF SAMPLE 4 Β-CAROTENE FROM CARROT, EXTRACTED IN ETHYL ACETATE ............ X
FIGURE 27: HPLC GRAPH OF SAMPLE 10 Β-CAROTENE FROM CARROT, EXTRACTED IN HEXANE/ACETONE....... X
FIGURE 28: HPLC GRAPH OF SAMPLE 6 Β-CAROTENE FROM CARROT, EXTRACTED IN HEXANE/ACETONE......... X
FIGURE 29: HPLC GRAPH OF SAMPLE 12 Β-CAROTENE FROM CARROT, EXTRACTED IN HEXANE/ACETONE...... XI
FIGURE 30: HPLC GRAPH OF SAMPLE 5 Β-CAROTENE FROM CARROT, EXTRACTED IN PETROLEUM ETHER ...... XI
FIGURE 31: HPLC GRAPH OF SAMPLE 11 Β-CAROTENE FROM CARROT, EXTRACTED IN PETROLEUM ETHER .... XI
X
Master thesis by Susan Hecker
List of Abbreviations
Abbreviation
Unit
Å
a*
b*
C*
FTIR
Red-green
Blue-yellow
h
HPLC
K/S
L*
NaOH
Owf
RP-HPLC
TLC
UV
v
Chroma
Fourier transform infrared spectroscopy
Hue
High Performance Liquid Chromatography
Kubelka Munk, colour strength or colour uptake
Lightness
V
ΔE
Sodium hydroxide
Of the weight fraction
Reverse-Phase High Performance Liquid Chromatography
Thin Layer Chromatography
Ultra violet
Volume
Volt
Colour difference between uncoloured and coloured fabric
μg
λ max
Definition
Ångström
Microgram
Maximal absorption
Master thesis by Susan Hecker
XI
1. Introduction
Plant material like fruits, berries, roots, barks, vegetables, leaves etc. have been used for
textile dyeing over hundreds of years (Cardon, 2007). With the industrialisation and the
invention of synthetic dyes, natural dyes disappeared almost entirely in the textile industry
(Cardon, 2007, Hardman and Pinhey, 2009). With their well-known structures synthetic dyes
became easier in handling, higher in process safety and better in reproducibility compared to
natural dyes and so a success for dye houses. Nevertheless different researchers
investigated in the recent years again in new methods on dyeing with natural material and
their extracts. The used extraction methods are mostly organic-solvent extraction (Saleh et
al., 2013, Yi and Yoo, 2010). The article by Saleh et al. (2013) shows great potential of
dyeing cotton fabric with β-carotene containing dyestuff extracts, extracted from banana
leaves by organic solvent extraction. The results evidenced high tensile strength, high colour
strength, and high fastness properties for the dyed cotton fabrics.
Brämhults Juice AB, Borås has 8100 tons of orange peel waste and 570 tons of carrots
waste from juice pressing each year (Figure 1). In this state after the juice pressing the
vegetable and fruit residues still contain a lot of natural resources such as colorants and
pigments (Brämhults Juice AB, 2014). A further use of the waste could be the textile dyeing
industry (Guinot et al., 2007, Saleh et al., 2013). With extraction and purification of the
pigments, dyes comparable to synthetically textile dyes can be achieved.
570
630
Orange peel
Carrots
Others
8100
Figure 1: Fruits and vegetable waste from juice pressing of
Brämhults Juice AB in tons per year.
2
Master thesis by Susan Hecker
β-carotene is a natural yellow-red pigment. Its chemical formula is C40H56. Its sources are
mainly plants, vegetables and fruits with a yellow-red colour as carrots and oranges. It is an
important precursor for vitamin A since it cannot be synthesised in the human body. The
intake has to be over the diet (Schlieper, 2005). As colorant it is used in the food industry
under the numbers E160 a-f (Domke et al., 2004).
The total amount of residue of the juice pressing of all vegetables and fruits from Brämhults
Juice is 9300 tons per year.
500
500
Biogas production
and fodder
Fodder
Waste disposal
8300
Figure 2: Waste handling of Brämhults Juice AB. Waste in
tons per year.
This is composed of 8300 tons that is used for the biogas production and fodder, 500 tons for
fodder and 500 tons for waste disposal (Figure 2) (Brämhults Juice AB, 2014). A suitable
extraction method of β-carotene from orange peel and carrot residue would gain an
additional value to their waste. The further use of the residues can be economically beneficial
for both the juice pressing and the textile dyeing industry. Synthetically produced β-carotene
and the extracted β-carotene from natural sources have the same chemical structure and are
therefore comparative with each other.
Master thesis by Susan Hecker
3
2. Literature review
The following literature review gives a look into the physical and chemical properties of βcarotene, its synthesis and uses today. It gives an overview on the existing research in the
field of extraction of β-carotene and other natural pigments and their appliance in different
dyeing methods. It is shown how much β-carotene other researchers could extract from
orange peel and carrots. An investigation on different solvents and their suitability for βcarotene extraction was done. Different dyeing methods used for water insoluble dyes and
on cotton and the usage of mordants is described as well.
2.1.
β-carotene
β-carotene is a natural yellow-red coloured pigment with the chemical structure C40H56,
(Figure 3). It occurs mainly in plants, fruit and vegetables. It belongs to the group of
carotenes that together with xanthophyll belongs to the upper-level grouping of carotenoids
(Bergmann, 2004). Chemically β-carotene is classified as tetraterpene (Koskinen, 2012).
Carotenoids are divided in oxygen containing molecules (xantophylls) and non-oxygen
containing molecules (carotene) (Domke et al., 2004), Figure 4. The lack of hydroxyl groups
makes β-carotene hydrophobic. Due to its two cyclic rings at each end of the molecule chain,
β-carotene is a dicyclic compound, composed of 8 isoprene-units (C5H8). The high amount of
conjugated double bonds is called chromophor and is responsible for the colour impression
(Bergmann, 2004). β-carotene absorbs light of the wavelength 450 nm of the visible part of
the spectrum (Bauernfeind, 1981).
Figure 3: All-trans β-carotene
In the nature carotenoids have an indispensable protective role for chlorophyll and the
human eyes by absorbing and dissipating excessive light energy that would damage them
4
Master thesis by Susan Hecker
(Campbell, 2003). β-carotene is the most common carotene (Schlieper, 2005). β-carotene is
very sensitive to light, heat and oxygen. It can change its chemical structure due to oxidation,
degradation or isomerization. The latter doesn’t have any effect on the colour impression of
β-carotene because the double bonds do not break (Liaaen-Jensen, 1989). The handling of
carrots and orange peel residue for the β-carotene pigment extraction and the later dyeing
process therefore has to be handled with care. Too high exposures to light, heat and oxygen
have to be avoided. The storing is recommended under frozen conditions. According to Qian
et al. (2012) the β-carotene stability against degradation is higher at a pH between 4-8.
Natural β-carotene occurs in trans- and cis-isomers whereas synthetically produced βcarotene is mostly all-trans-form, due to its higher absorbance for the human body
(Bundesinstitut für Risikobewertung, 2013).
Carotenoids
Carotene
(hydrophobic)
Xantophylls
(hydrophillic)
β-carotene
α-carotene
Lutein
Lycopene
etc.
etc.
Zeaxanthin
Figure 4: Carotenoids classifications
The β-carotene synthesis is either produced by employing a Wittig reaction or a Grignard
reaction (Koskinen, 2012). The following reaction (Figure 5) is by Wittig. It shows a transselective Wittig olefination of aldehydes II—synthesis of β-carotene from a dialdehyde.
Master thesis by Susan Hecker
5
Figure 5: β-carotene synthesis by Wittig reaction
As vitamin A precursor, synthetically produced β-carotene is especially important for the use
in the food industry as completion to the natural, in the food existing β-carotene. As food
additive with the purpose of a food colorant it is known under the numbers E160 a-f. (Domke
et al., 2004)
Figure 6: 9-cis-β-carotene
6
Master thesis by Susan Hecker
2.2.
Carrots and Orange peel
Carrots and orange peels are a rich source for carotenoids. Carotenoids, especially βcarotene is present in both of them and is responsible for their yellow-orange colour. Main
compounds in carrots and orange peel are β-carotene, α-carotene, lutein and zeaxanthin
(Table 1), (Heinonen, 1990, Wang et al., 2008, Curl and Bailey, 1956). Additionally orange
peel contains flavonoids, phenolic acids, pectin and waxes (Wang et al., 2008). The
compounds in orange peels extracts are by some researches tested on their UV-protective
properties and antimicrobial activity (Hou et al., 2013, Yi and Yoo, 2010). The total
carotenoids content and the amount of some xantophylls and carotenes of carrots and
orange peel are listed below.
Table 1: Total carotenoids and the content of different carotene and xanthophylls in carrots
and orange peel in mg/g carrots/orange peel
Carrots
Total
carotenoids
0,16 – 0,38
Article
Mustafa et al.
(2012)
Orange peel
Article
0,45
Wang et al. (2008)
Carotenes (hydrophobic carotenoids)
β-carotene
α -carotene
0,046 – 0,10
0,022 – 0,049
Heinonen
(1990)
Heinonen
(1990)
Dumbravă
0,05 – 0,056
et
al.
(2010), Wang et al.
(2008)
0,017 – 0,019
Curl
and
Bailey
(1956)
Xanthophylls (hydrophilic carotenoids)
Lutein
0,0011 – 0,0056
Zeaxanthin
0,0574
Heinonen
(1990)
Curl and Bailey
(1956)
0,029
Wang et al. (2008)
0,027
Wang et al. (2008)
Master thesis by Susan Hecker
7
2.3.
Extraction methods and solvent properties
In literature different extraction methods for β-carotene (carotenes) from orange peel, carrots
and other fruits, vegetables and plants are described. The most common method is the
organic solvent extraction. It was described by several different researches for the extraction
of β-carotene from orange peel (Ghazi, 1999), (Dumbravă et al., 2010) and carrots (Biswas
et al., 2011), (Fikselova et al., 2008), (Livny et al., 2003), (Marx et al., 2000), but also from
other vegetables and plants as tomato, paprika (Levy et al., 1995) and from the algae
Dunaliella salina (Marchal et al., 2013), (Mojaat et al., 2008).
Several other extraction methods for β-carotene were researched. So were investigations on
supercritical fluid extraction of β-carotene done by Kaur et al. (2012), Chandra and Nair
(1997) and Benelli et al. (2010) for carrots and the latter one for orange peel. “Ultrasound
assisted extraction of β-carotene from Spirulina platensis”, a cyanobacteria was researched
by Dey and Rathod (2013). A microwave-assisted extraction of β-carotene from carrots was
done by Hiranvarachat et al. (2013) and the effect of enzymes on carotene extraction in
carrots was investigated by Jaramillo‐Flores et al. (2005).
The “Relative solubility, stability, and absorptivity of lutein and β-carotene in organic solvents
were tested by Craft and Soares (1992). The solubility of both carotenoids (β-carotene and
lutein) was best in tetrahydrofuran, whereas the least solubility for lutein was in hexane and
for β-carotene methanol and acetonitrile. Organic solvents can be classified due to their
chemical structure, according their solubility in water they are grouped into polar and nonpolar solvents (Smallwood, 1996). A good solubility for β-carotene was shown by some
organic solvents of the category hydrocarbons (same chemically classification as βcarotene), ethers, esters, chlorinated solvents and ketones (Craft and Soares, 1992). For the
selection of a suitable solvent for β-carotene extraction a low boiling point is advantageous.
The solvent has to be removed by rotary evaporator after the extraction to achieve pure βcarotene, with a low boiling point the thermal sensitive β-carotene can be preserved.
Two articles used the organic solvent mixture of acetone and hexane (1:1 v/v) for the
“Extraction of β-carotene from orange peel” (Ghazi, 1999) and from carrot juices (Marx et al.,
2000). Ghazi (1999) concluded that the organic solvent mixture of acetone and hexane
achieves the highest extracted yield of β-carotene. Biswas et al. (2011) extracted amongst
others β-carotene from carrots with four different organic solvents; acetone, diethyl ether,
8
Master thesis by Susan Hecker
acetonitrile and methanol. With acetone the highest yield was achieved followed by diethyl
ether. In another study β-carotene was extracted from orange (Citrus sinensis L.) fruits peel
extracts by three different organic solvents; ethanol, benzene and petroleum ether. Only in
the petroleum ether extract, β-carotene could be identified. The amount in the ethanol and
benzene extract was too little (Dumbravă et al., 2010). Ishida and Chapman (2009)
compared ethyl lactate as an environmentally friendly solvent with the, for the food industry
commonly used solvents ethyl acetate and ethanol for the carotenoid extraction. It was
mentioned that ethyl acetate is a good solvent for β-carotene and lutein and less good for alltrans isomer of lycopene. That could help to achieve a higher purity of the extracted βcarotene. Therefore ethyl acetate was chosen instead of ethyl lactate, which was slightly
better in the extraction of β-carotene.
The following table are the solvents that were used in this thesis for the β-carotene extraction
from orange peel and carrot residue. They are listed after their solvent classification. The
solubility of β-carotene (mg/L) and the stability after 10 days in % of the initial absorbance of
β-carotene at λmax (Craft and Soares, 1992), values of the boiling point (°C) and the polarity
of the solvents are given (Smallwood, 1996).
Table 2: Solvent properties of the used solvents
Solvent
Solubility of
β-carotene
1
mg/L
Stability after 10 days
% of initial absorbance
1
of β-carotene at λmax.
Boiling
2
pt. °C
Polarity
2
(H2O=100)
Article that uses
the solvent
Hydrocarbons
Petroleum
ether
-
-
36
0,9
(Dumbravă et al.,
2010)
Hexane
600
92
69
0,9
(Ghazi,
1999,
Marx et al., 2000)
500
05
77
6,02
(Ishida
and
Chapman, 2009)
200
93
56
20,6
(Ghazi,
1999,
Marx et al., 2000)
Esters
Ethyl
acetate
Ketones
Acetone
1
(Craft and Soares, 1992); 2 (Smallwood, 1996)
Master thesis by Susan Hecker
9
2.4.
Dyeing with natural sources
There are different researches in the field of textile dying with natural plant and vegetable
extracts. Some of them are based on the extraction of pigments from orange peel and
carrots but are not specialized on the extraction of β-carotene. The extraction methods used
are depending on the solubility properties of the natural extract are mostly either aqueous
extraction or organic solvent extraction.
Hou et al. (2013) and Yi and Yoo (2010) investigated UV-protective properties and
antimicrobial activity of orange peel extracted dye. They used water and organic solvent
extraction to achieve dyestuff for wool and cotton fabric. Hou et al. (2013) analysed the water
extracted main colorant compounds in the extracted dyestuff as phenolic colorants and
pectin by UV-visible spectroscopy and FTIR (Fourier transform infrared spectroscopy). A
direct dyeing was done on wool with pre-, one-bath and post-mordanting. As mordants alum
and ferrous sulphate were used to achieve good colour, wash and acceptable light fastness
properties. Yi and Yoo (2010) identified flavonoids as the main colorant extracted by ethanol.
They concluded that the best dyeing conditions for cotton with orange peel dyestuff were a
pH of 3 at 60°C for 60 min, with a dye concentration of 800% owf.
With focus on by-product or waste further usage, dyestuff extracts of carrots and banana
leaves were analysed. Under aqueous extraction flavonoid aglycones were extracted from
carrot by-products and dyed on pre-mordanted hemp fibres. Analysis of the vegetable
colorant was done by TLC (Thin layer chromatography) and HPLC (High performance layer
chromatography). Two solvents for extraction were applied on banana leaves. In the alkaline
extract luteolin and apigenin and in the acetone extract chlorophyll a, and b and β-carotene
were analysed with HPLC. With ferrous sulphate, copper sulphate and potassium dichromate
premordanted cotton fabric was dyed in a closed bath for 5 min. at 56°C. The results showed
high tensile strength, high colour strength, and high fastness properties. (Guinot et al., 2007,
Saleh et al., 2013)
There are different investigations from the University of Innsbruck, Austria on the topic of
natural textile dyes from plant extracts. In all their articles aqueous extraction is applied to
10
Master thesis by Susan Hecker
assure an environmental friendly extraction process. In this way the residues of the
extraction can be further used as animal food or combustion. They registered the extraction
method for a patent. The plant materials used are amongst others Canadian golden rod,
onion skin, dyer´s weed and nutshells. The extracted dyes are analysed by a diode array
spectrophotometer. The dyestuff compounds analysed are flavonoids, tannins et al. Solid
dyestuff was produced from the extract due to the better handling and storing for the dye
houses. Exhaust dyeing was the employed dyeing method. The substrates used were linen,
polyamide and wool. To bind the dyestuff better to the fibres mordanting with alum, ferrous
sulphate and ferrous chloride was applied. (Bechtold et al., 2007, Leitner et al., 2012,
Wallner and Wenisch, 2011)
2.5.
Mordants
Dye extracts of natural sources often have none or little amount of functional groups. Due to
that, their fastness properties are weaker. To overcome that disadvantage mordants are
often used to fix the colorant to the textile substrate. Mordants are metal or mineral salts that
function as a bond between the textile substrate and the dyestuff (Flint, 2008). There are
many different mordants used to help natural dyestuffs binding to a textile substrate. Some
have only the function of fixing the dyestuff to the textile substrate (alum) whereas others can
also influence or change the colour impression and saturation of the dyed textile (ferrous
sulphate), depending on the nature and concentration of the mordant (Cardon, 2007). Most
of them are heavy metal salts such as copper sulphate, ferrous sulphate and potassium
dichromate others are alum, stannous chloride and sodium chloride. Among them alum,
potassium aluminium sulphate (KAl(SO4 )2 ×12H2O) and ferrous sulphate (FeSO × 7H O)
4
2
are very common used mordants due to their lower environmental pollution (Hou et al., 2013,
Bechtold et al., 2007, Choo and Lee, 2002, Meksi et al., 2012). There are different methods
when the mordant is added to the dye bath. Pre-, simultaneous- or post-mordanting. In premordanting, the to dyeing fabric is mordanted before the actual dyeing in a separated
mordanting bath. In simultaneously mordanting the mordant is added to the dye bath. This
brings the risk that the dyestuff and the mordant bind together before either of them has
attached to the fibre but its quick and saving resources as water and energy. The postmordant method is used to fixate the colours permanently and intensifies them. It is done in a
separate mordant bath after the dyeing process (Flint, 2008). Usually mordanting is done on
wool or other animal fibres. Cotton mordanting is rarely found in literature. One article that
Master thesis by Susan Hecker
11
was investigating in the mordanting of cotton fabric was by Ali et al. (2009). They used preand post-mordanting of cotton fabric in a mordant-bath ratio of 1:15 (goods/liquid). The
mordants used were alum and ferrous sulphate. The best results were achieved for postmordanted cotton fabric with alum in a concentration of 10% (owf).
2.6.
Dye and dyeing method
The difference between natural dyes to synthetic dyes is that natural dyes often don´t consist
of a single colouring molecule. Often they consist of several different colorants that belong to
different chemical groups (Cardon, 2007). A good example therefore is the orange peel.
Besides hydrocarbons (were β-carotene belongs to) orange peel consists of diole,
monoether diole, diether diole and polyole (Curl and Bailey, 1956). This and the changing
amount of the various compounds due to seasonal deviations make the repeatability of those
dyes more difficult compared to synthetic dyes. The classifications of textile dyes are either
done by their chemical structure and properties or by the fibre that will be dyed with it. The
most prominent property of the β-carotene is its water insolubility. This was the factor after
what the dye method was chosen. The other factors were the used fabric material, cotton. As
water insoluble textile dyes vat, sulphur and acid disperse dyes for polyester and metal
complex dyes are known. The substrates dyed with them are mostly cellulosic and polyester
fibres. Reactive dyeing is the most common dyeing method for cellulosic fibres as cotton and
viscose.
Cellulose fibres can be dyed by sulphur dyes. Similar to the β-carotene dyestuff they are
both water insoluble dyes. The difference is the containing sulphur group in the sulphur dye.
The water insoluble sulphur dyes becoming water soluble through alkali reduction.
Vat dyes are also used for the dyeing of cellulosic fibres. Vat dyes are absolutely water
insoluble and due to reduction they are transformed to the water-soluble leuco form. After the
dyeing process they are getting water insoluble again by oxidation.
The highest similarities in the dye structure have β-carotene with disperse dyes for
polyester. Both are low water-soluble due to the lack of existing water-soluble making
groups and reactive groups in the dye molecule. The dye molecules are relatively small and
non-polar (Marx, 2011). Some disperse dyes hydrolyse under alkaline conditions and form
free carboxylic acids that are soluble in the alkaline solution (Broadbent et al., 2001).
12
Master thesis by Susan Hecker
Hydrolysis of a disperse dye for polyester under alkaline conditions:
Dye - NR - CH2CH2CO2CH3 (s) + HO- (aq)
¬¾
®
Dye - NR - CH2CH2CO-2(aq) + CH3OH(aq)
The synthetically form of β-carotene is absolutely water insoluble, but nature has not
bounded the natural β-carotene to lipoidal systems. The protein complexes of which the βcarotene is surrounded in carrots and orange peels and colloidal structures allow the βcarotene to be dispersed in primarily aqueous systems. (Bauernfeind, 1981)
Reactive dyeing is the most common used dyeing method in the industry. Reactive dyes are
quite small (comparable to β-carotene) and water soluble molecules. The dyeing medium is
usually alkali and a huge amount of sodium chloride is used as electrolyte addition. The
chemical binding between reactive dyes and the fibres are an atom binding. (Marx, 2011)
The fact that polyester requires dyeing temperatures of 130°C to open up the fibre structure
and let the dye molecules migrate into fibre, cotton was chosen. Cotton can be dyed at
temperatures around 60°C. The high temperatures of polyester dyeing would destroy the βcarotene double bonds and so the colour impression. The only possibility the dye polyester
at lower temperatures is with the use of carriers. But they are highly environmentally
unfriendly. (Marx, 2011)
Master thesis by Susan Hecker
13
3. Problem description
Brämhults Juice AB is producing 8670 tons of orange peel and carrot waste every year from
the total waste of 9300 tons. In the state after the juice pressing the vegetable and fruit waste
is still a valuable source for different natural pigments that can be extracted and used as
dyestuff for the textile dyeing industry. Alone from the carrots and orange peel waste more
than 200 kg of β-carotene could be extracted and used as a new dyestuff source for the
textile dyeing industry. There is no previous research found which focus on the extraction of
β-carotene dyestuff from orange peel and carrots as further use for cotton dyeing.
The extracted β-carotene dyestuff has to be a process stable product for dye houses. The
dye process has to be secured and reproducible with the same colour result to be applicable
for a dye house. Therefore an, as possible, pure dye has to be extracted from the natural
source, orange peel and carrots residue.
3.1.
Aim
The aim of this master thesis is a comprehensive study. It is about the extraction of βcarotene dyestuff from orange peel and carrot residue from the juice pressing industry and
the analysis of the extract by HPLC on purity and yield of β-carotene. Further the usage for
the dyeing of cotton fabric will be investigated. The extract on yield and the purity of the
extract have to be competitive with synthetically produced β-carotene. The dyed cotton
fabrics are tested on their wash and light fastness to see if the dyestuff is competitive with
synthetic dyes. The aspect of environmentally friendly solvents for the extraction is neglected
in the study. This might be interesting for the further work. For the present report this was out
of scope.
3.2.
Research question
 Which organic solvent or mixture is most suitable for β-carotene extraction from
carrots and orange peel
14
Master thesis by Susan Hecker

Does the extracted β-carotene conform to synthetically produced β-carotene?

Which yield and purity can be extracted?
 Which dyeing method is most suitable to dye cotton with the extracted β-carotenedye?

Can acceptable wash and light fastness be achieved on the dyed cotton
fabric?

Can a better result be achieved with using a mordant?
Master thesis by Susan Hecker
15
4. Materials and methods
All the materials and methods that were used in this work are explained and listed in the
following chapter. There were always 2 samples done for the HPLC analysis and for the
dyeing with its fastness and colour measurements, to value the results in the arithmetic
average of 2 samples.
4.1.
Materials
The residue of orange peels and carrots were from the local juice company Brämhults Juice
AB, Borås. Orange peel Valencia from Egypt and small amounts from Mexico, Tunisia and
Spain and carrots from the Nantes family from Gotland, Sweden were used for the βcarotene extraction. The orange peel residues were 0,5 – 2 cm and the carrot residue was
0,1– 0,2 cm big when received from the juice company. The organic solvents used for the βcarotene extraction were: ethanol absolute (>99,8%) from VWR International (preliminary
tests), ethyl acetate (≥99.9%) and a mixture of acetone (≥99.9%) and hexane (≥97.0%), all
HPLC grade and petroleum ether, spectrophotometric grade (all chemicals if not other
mentioned were from Sigma Aldrich). A bleached and mercerized cotton weave fabric (150
g/m2) was used for the dyeing process.
The mordants that were used are potassium aluminium sulphate (potassium alum)
(KAl(SO4 )2 ×12H2O) and ferrous sulphate (FeSO × 7H O) from Sigma Aldrich. Auxiliaries
4
2
used for the dyeing process were sodium chloride as electrolyte addition, Zenit Levelling
agent S from Zenit AB Sweden, dispersing agent Lyocol RDN fl. from Clariant Germany,
sodium hydroxide 25% (NaOH) and acetic acid 25% for the pH regulation.
16
Master thesis by Susan Hecker
4.2.
Drying of the carrot and orange peel residue
For drying a desiccator silica with silica gel
was used. The drying was done to remove
the water from the residue to make sure
that only the residue without any water
was used for the further extraction. The
samples were dried in a dark place to
prevent degradation of the β-carotene due
to
light
exposure.
The
drying
was
continued until the dry matter weight was
Figure 7: Desiccator silica under vacuum
containing orange peel and carrot pieces
not changing anymore to be sure all
moisture was removed from the samples.
To achieve a higher surface area the dried
The orange peel and carrot were dried
orange peel and carrot pieces were
under vacuum with a vacuum pump RV12
ground in a porcelain mortar after the
from Edwards of 200 V before extracted.
drying.
Figure 8: Grinding dried orange peel and carrots in a mortar
Master thesis by Susan Hecker
17
4.3.
Extraction method
ethyl acetate in a sample/solvent ratio of
1:15. The amounts of dried residue and
solvent used for the HPLC samples and
the dyestuff extraction were listed in the
table below. The dried ground samples
were mixed together with the solvent. The
mixture was kept at room temperature
under magnetic stirring for 30 min. After
filtration (Munktell filter paper, grade 3) the
Figure 9: Extraction from carrots
in petroleum ether
solvent
was
evaporated
using
rotary
evaporator Büchi Rotavapor R-114 with
Organic solvent extraction was used to
Büchi Waterbath B-480 and an Edwards
extract
from
vacuum pump R12 of 200 V. First the
orange peels and carrots residue. The
HPLC samples were done and after their
organic solvent extraction was carried out
analysis the dyestuff extraction for the
with the solvents: hexane and acetone
dyeing was continued. From each sample
mixture (1:1 v/v), petroleum ether and
2
the
β-carotene
dyestuff
duplicates
were
done.
Table 3: Amounts of the dried residues (g) and solvents (ml) that were used for the HPLC
samples and the dyestuff extraction for the cotton dyeing.
Solvents
HPLC samples
Dyestuff extraction
Ethyl acetate
5 g dried orange peel or carrot
Ethyl acetate was not used for the dyestuff
extraction due to the lowest yield of β-carotene
75 ml solvent
Petroleum ether
Hexane/acetone
determined by HPLC.
5 g dried orange peel or carrot
27 g dried carrot
75 ml solvent
400 ml solvent
5 g dried orange peel or carrot
27 g dried orange peel
75 ml solvent
400 ml solvent
mixture
Master thesis by Susan Hecker
17
4.4.
Analysis of the dyestuff, purity and yield
The spectrum for β-carotene was determined at 450 nm. For its standard curve 4 different
concentrations of β-carotene (β-carotene standard, synthetic ≥95%, HPLC grade, crystalline
from Sigma Aldrich) in hexane/ acetone (1:1 v/v) were made spectrophotometrically in a
Biochrom Libra from Nordic Biolabs. The concentrations for the standard curve were 0,033,
0,0056, 0,00093 and 0,00015 mg/ml of β-carotene.
Reverse Phase High Performance Liquid Chromatography (RP-HPLC) was used to
analyse β-carotene in the extracts and to determine the purity and the yield. 2 x 6 samples
were analysed. β-carotene analysis was realized by a Waters Alliance Seperation Module
2695, with a UV detector, Refractive Index Detector 2414 at 450 nm. The column was a C18
Sunfire reverse phase column from Waters, 4,6 mm × 250 mm; 5,0 µm particle size and pore
size 100 Å. The mobile phase was a methanol and acetonitrile (9:1 v/v) mixture. The flow
rate was 1,0 ml/min. β-carotene standard, synthetic ≥95%, HPLC grade, crystalline from
Sigma Aldrich was used. The standard for the HPLC was prepared in the concentrations
0,033, 0,025 and 0,0167 mg/ml. With help of the standard curve the concentration of βcarotene in the extracts was calculated to draw conclusions from how much β-carotene could
be extracted per grams of carrots or orange peel and how much β-carotene was used for the
dyeing of cotton fabric. The 12 samples for the RP-HPLC analysis were diluted with 6 ml of
the used solvents for each extraction.
4.5.
Cotton dyeing
A reactive dyeing method in a dye bath ratio of 1:50 (sample/dye bath g/ml) was applied. The
sample size was 10 cm x 5,5 cm. The size was big enough to cut a sample for the washing
test of the size 10 cm x 4 cm and a sample for the light fastness test of the size 7 cm x 1,5
cm. The rest of the sample was used as a reference sample to compare the colour changes
after the fastness property tests.
The dye: The dye used for the cotton fabric dyeing is named β-carotene dyestuff of carrot or
orange peel residue in the following work. The extracts achieved by organic solvent
extraction were used without any modifications. The two highest yields of the β-carotene
extraction of orange peel and carrot residue were used for the dyeing. The petroleum ether
18
Master thesis by Susan Hecker
extract from carrots and the hexane/acetone mixture extract from orange peel. The dyes
consists of the extracted β-carotene in known yield (determined by the RP-HPLC analysis)
and the by literature determined compounds α-carotene and zeaxanthin. Besides this
allocated compounds, the β-carotene dyestuff of orange peel residue contains flavonoids,
phenolic acids, pectin and waxes.
Table 4: Composition of the used β-carotene dyestuff of carrot and orange peel residue that
was used for the dyeing of 0,8 g cotton fabric.
Dye name
β-carotene dyestuff
Containing β-
Containing
carotene amount
compounds
determined by RP-
determined by
HPLC analysis
literature
10,5 mg
α-carotene and
of carrot residue
β-carotene dyestuff
Other compounds
-
zeaxanthin
0,1 mg
of orange peel
residue
α-carotene and
Flavonoids, phenolic
zeaxanthin
acids, pectin and
waxes
After the evaporation of the solvent the dye was left in the rotary evaporator beakers. The
amount for both extracts (carrot and orange peel) was so less that it was impossible to
remove the dye by a spoon. The chosen way to remove the dye from the glass beaker walls,
was to solve it in the rotary evaporator beakers. Half of the water amount used for the dye
bath (20 ml) was added to the beakers together with the 8 ml sodium hydroxide. The sodium
hydroxide showed in pre tests that it helps the extract to solve in the water. It could be
observed that the β-carotene dyestuff of orange peel residue solved readily whereas βcarotene dyestuff of carrot residue needed to be warmed in a water bath together with the
glass beaker. Additionally the extract was scratch from the glass walls with a spattle. The
dyeing pistons were each prepared with the additives: 1,2 g sodium chloride, 4 ml levelling
agent, 0,2 ml dispersing agent and 0,8 g cotton fabric. Then the solved dye was added to the
piston. The rest of 20 ml water was to rinse the rotary evaporator bakers once more, to be
sure, that all dye was removed form the beaker. Acetic acid 25 % was added to the piston
until the pH between 4-5 was reached.
Master thesis by Susan Hecker
19
Table 5: Final dye receipt. Amount per litre water and the calculated amount that was
used for the dyeing of 0,8 g cotton fabric is listed.
Chemical/Additives
Amount per litre
Calculated amount used for the
water
dyeing of 0,8 g cotton fabric
Cotton
-
0,8 g
Water (tap water)
-
40 ml
β-carotene dyestuff of carrot
Defined in table 4
residue
β-carotene dyestuff of
Defined in table 4
orange peel residue
Salt
30 g/l
1,2 g
Levelling agent 10%
20 ml/l
4 ml
Dispersing agents, lyocol
5 ml/l
0,2 ml
200 ml/l
8 ml
RDN 10%
Sodium hydroxide 25 %
The samples were dyed in a dyeing machine, Pyro Tec MB 2 from Roaches International Ltd.
at 60°C for 60 min. with a rate of 2°C/min. After dyeing the samples were rinsed in cold water
and dried in a dryer.
4.6.
Spectrophotometric measurements
Spectrophotometric measurements of the dyed samples were done with a Datacolor CHECK
Pro and evaluated by the Datacolor TOOLS software from Datacolor. The measured values
were the average of 5 measurements. As standard illuminate D65/10 (daylight) was used
and a tolerance of CMC 2:1. The values measured were K/S Kubelka Munk, which can be
seen a colour strength or colour uptake, the CIE colour parameters for lightness-darkness L*,
red-green a*, blue-yellow b*, chroma C*, hue h and ΔE. Where ΔE describes the colour
20
Master thesis by Susan Hecker
difference between the uncoloured bleached and mercerized cotton weave with coloured
ones. The Datacolor TOOLS software automatically calculated all values.
4.7.
Fastness properties tests
Two fastness property tests were done on the dyed cotton samples. Xenon test (ISO-Norm
B02) and wash fastness test (ISO 105 – C).
4.7.1. Xenon test (ISO-Norm B02)
To determine the light fastness of the dyed cotton fabric xenon test according to ISO-Norm
B02 was done. The prepared samples had the size 1,5 x 7 cm. The samples were rayed by
xenon arc light up to grade 4 and analysed in D65 illuminate.
4.7.2. Wash fastness test (ISO 105 – C)
The wash fastness was ascertained by a wash fastness test according to ISO 105 – C. The
sample was together with an additional adjacent fabric, a multistripe with 6 different fabric
samples (diacetate, cotton, polyamide, polyester. acryl and wool) washed for 30 min at 60°C,
with 50 ml of detergent solution (4 g detergent /l). 25 steel balls were added. A common
household detergent was used as washing liquid. The washed samples were rinsed and
dried. Afterwards the dyed cotton samples were analysed on colour change and the
multistripe adjacent fabrics were analysed on staining with the 5 grades grey-white scale in
D65 illuminate.
Master thesis by Susan Hecker
21
5. Results and discussion
The following chapter lists the results and interprets them in graphs, tables and explanations.
Some of the results are further analysed to become clearer. The results are the arithmetic
average of 2 samples done for the HPLC analysis and for the dyeing with its fastness and
colour measurements.
Preliminary test showed that ethanol and water as solvents are not suitable for β-carotene
extraction due to their polar character (see Attachment I).
5.1.
Determination of β-carotene spectrum
The determination of the spectrum was done in the range of 190 – 500 nm to determine the
visible range of β-carotene. The identified wavelength was at 450 nm that is confirmed by
literature (Bauernfeind, 1981).
Figure 10: Graph of the determined
wavelength of β-carotene at 450 nm.
22
Master thesis by Susan Hecker
5.2.
Results of the RP-HPLC analysis
The following graph shows the HPLC-standard curve for β-carotene.
1600000
y = 41,479,150.34x
R² = 1.00
1400000
1200000
Area
1000000
800000
600000
400000
200000
0
0
0.01
0.02
0.03
Concentration (mg/ml)
0.04
Figure 11: HPLC-standard curve of β-carotene
The standard or calibration curve was done by three different concentrations of β-carotene in
mg per ml solvent. Those are listed in the table below together with the area. The standard
curve proceeds linearly.
Table 6: Three different concentrations used for the
HPLC-standard curve
Sample
Area
Concentration (mg/mL)
Standard 1
1389337
0,034
Standard 2
1019366
0,025
Standard 3
704341
0,0167
Master thesis by Susan Hecker
23
The RP-HPLC analysis identified clearly β-carotene in the extracts of carrot and orange
peel at a retention time around 55 min (Figure 12-13) and a purity of more than 50 % in the
carrot extract. The differences in the orange peel and carrot extract were the amount of other
compounds that were shown by the HPLC. An identification of the other main compounds
was done with help of literature.
RP-HPLC analysis in this work was based on the article by Ben-Amotz and Fishier (1998).
Due to their work other peaks besides β-carotene were allocated. They identified three main
peaks of carrots: β-, α-carotene and zeaxanthin. According to them β-carotene showed the
highest area and zeaxanthin the lowest. Another compound with a very low area was lutein.
With the determined retention times in the HPLC graphs of the carrot extract compounds,
these three compounds could be identified in the orange peel samples as well (Figure 12,
13). The purity of carotenes in the extract is with an area of β- and α-carotene of more than
90 % and less other compounds very high. Besides the allocated compounds (α-, β-carotene
and zeaxanthin) orange peel contains flavonoids, phenolic acids, pectin and waxes (Wang et
al., 2008).
Figure 12: HPLC graph at 450 nm showing β-carotene, α-carotene and zeaxanthin
extracted from carrot residue
Figure 13: HPLC graph at 450 nm showing β-carotene, α-carotene and zeaxanthin
extracted from orange peel residue
24
Master thesis by Susan Hecker
There was no significant difference in the purity and amount between the different solvents.
The lowest amount of β-carotene was extracted by ethyl acetate with 0,26 mg/g of carrots
and 0,0027 mg/g of orange peel. The highest amount of β-carotene from carrots was
extracted by petroleum ether with 0,39 mg/g and from orange peel respectively with 0,0036
mg/g from petroleum ether and hexane/acetone mixture, (Table 4).
Table 7: β-carotene content in carrots and orange peel (mg/g of
carrots or orange peel)
Organic solvent
Carrots
Orange peel
Ethyl acetate
0,26
0,0027
Petroleum ether
0,39
0,0036
Hexane/acetone (1:1 v/v)
0,30
0,0036
The extracted amount varies only minimal between the different solvents. The highest
extraction yield for both carrot and orange peel residue, achieved petroleum ether. Whereas
the difference to the lowest extraction yield with ethyl acetate is only minimal for carrot and
orange peel extracted β-carotene. The amount of extracted β-carotene increased with
decreasing polarity of the solvents.
Neither water as a polar solvent nor ethanol as an organic solvent is suitable for extracting
the non-polar β-carotene. No relevant peaks were allocated at a wavelength of 450 nm with
the HPLC of these extracts. Hence nor or too less β-carotene or other carotenes were
extracted. The same result was obtained by Dumbravă et al. (2010) the extracted amount
with ethanol from orange peel was to low for determination in a visible range. Ethanol has a
quite low relative solubility of ethanol for β-carotene with 30 mg/L (Craft and Soares, 1992)
compared with the used solvents (ethyl acetate, petroleum ether and hexane/acetone) were
acetone had the lowest relative solubility with 200 mg/L and hexane the highest with 600
mg/L. From the article by Yi and Yoo (2010) can be assumed that mostly flavonoids are
extracted by ethanol.
The selection of the solvents was done under the criteria of the polarity of the solvent and the
boiling point. For the evaporation of the solvent in the rotary evaporator it is important that
Master thesis by Susan Hecker
25
the solvents boiling temperature is as low as possible. Degradation of heat sensitive βcarotene is therefore avoided.
To solve the non-polar β-carotene, solvents of the same character were used with the
exception of the hexane and acetone mixture. A mixture of the non-polar solvent hexane and
the polar solvent acetone helped to extract the highest amount of natural β-carotene.
According to Ghazi (1999) “…naturally occurring carotenoids are found inside cells and
surrounded by an aqueous protoplasmic medium, so acetone can bind water and force
carotene to be out of solution.“
5.3.
Mordanting and Dyeing
Preliminary test showed that mordanting with ferrous sulphate adulterated the light shades of
β-carotene dye on cotton. The differentiation between ferrous sulphates own colour in
orange-red shades and orange shades of β-carotene dyestuff of carrot and orange peel
residue is difficult. This state of investigation is only based on the original colour of βcarotene dyestuff of carrot and orange peel residue on cotton. Therefore the research on
ferrous sulphate as mordant was not investigated further.
The results for the vat dyeing method (see appendix II) of cotton with β-carotene dyestuff of
carrot and orange peel residue extracted by the organic solvents ethyl acetate and
hexane/acetone did not show any colour uptake of the cotton. The vat dyeing bath was at
room temperature around 20°C. That had the consequence that the cotton fabric was barley
coloured because the colour pigments didn’t diffuse into the fibre. When changing the
method vat dyeing in reactive dyeing where the cotton fabric was dyed in the same
concentration and the ratio 1:40 at 80°C for 90 min, the fabric didn´t show any colour uptake.
The reason for this might be that at the temperature of 80°C the β-carotene degraded or
oxidized and the colour was destructed.
Regarding the results from the HPLC analysis the dyeing was done by the highest
concentration of β-carotene achieved. That was for carrots with petroleum ether as solvent
and for orange peel with hexane and acetone mixture. The extracts achieved by the organic
solvent extraction were not further modified. The composition of the dye is listed in table 4.
According to Ali et al. (2009) the best colour and fastness results for a mordanted cotton
26
Master thesis by Susan Hecker
fabric in colour coordinates L*, a*, b*, C*, h*, wash and light fastness was achieved by post
mordanting with 10% of alum owf of cotton.
Fair dyeing results were achieved with the dyeing at 60°C for 60min. The dye results on
cotton fabric with β-carotene dyestuff of carrot and orange peel residue are shown in figure
14-15. The same adjustments gave good results in the dyeing of cotton with unripe Citrus
grandis Osbeck extract done by Yi and Yoo (2010).
Figure 14: β-carotene dyestuff of carrot residue, dyed on cotton, mordanted and
unmordanted
Figure 15: β-carotene dyestuff of orange peel residue, dyed on cotton, mordanted and
unmordanted
The pH of the dye solution was adjusted to 4-5 were the β-carotene is according to Qian et
al. (2012) stable. The goods – dye bath ratio was 1:50. The dyed cotton fabric was for both
β-carotene dyes fairly even dyed. Even though the β-carotene concentration of the dyed
cotton with β-carotene dyestuff of orange peel residue is 100 times lower (0,1 mg β-carotene
is contained in the dyestuff) than the concentration in the β-carotene dyestuff of carrot
residue dyed samples (10,5 mg) their colour uptake ΔE is higher (Table 8-9). This might be
due to the different compounds that the β-carotene dyestuff of orange peel residue contains.
Master thesis by Susan Hecker
27
The different compounds might have more functional groups and a better absorbency on the
cotton fibres than the β-carotene itself has. The β-carotene that is mostly present in the βcarotene dyestuff of carrot residue has a lack of functional groups that makes it difficult to
absorb on the cotton fibres, which resulted in a lower colour uptake indicated by ΔE.
5.4.
Spectrophotometric colour measurements
The β-carotene dyestuff of carrot residue, dyed cotton fabric was dyed in a yellow-reddish
shade. The cotton fabric was evenly dyed. Compared to the lower concentrations of levelling
agent in some preliminary tests the evenness of the colour was good with 100ml/l for both βcarotene dyestuffs. The levelling agent lowered the rate of absorption of the β-carotene
dyestuff of carrot and orange peel residue on the cotton fabric, which resulted in a more
evenly dyed substrate.
The results for the colorimetric measurements of cotton weave fabric, dyed with β-carotene
dyestuff of carrot residue are given in table 5. The measurements were done by the CIE
colour system. The K/S values were for both unmordanted and mordanted samples the
same. The post mordanted samples were little bit lighter in colour due to the slightly higher L*
value. However the L* values for both unmordanted and mordanted samples were high
compared to the low values of C*. Considering both values L* and C* together the shades of
β-carotene dyestuff of carrot residue dyed cotton weave fabric were light and weak in colour
saturation. The a* and b* values indicate the shades of reddish yellow. The mordanted
samples were compared to the unmordanted samples slightly more colour saturated (C*) and
the mordant bind more dyestuff on the cotton fabric means more colour absorbed on the
cotton fabric (ΔE). The shade is more yellowish and less reddish.
Table 8: Colour coordination of the dyed cotton fabrics with carrot extract
Carrot
K/S
L*
a*
b*
C*
h*
ΔE
Unmordant
0,15
85,65
8,37
12,13
14,76
55,50
25,99
Alum post mordant
0,15
88,25
6,31
14,78
16,07
66,90
26,92
28
Master thesis by Susan Hecker
The ΔE shows that the colouration of β-carotene dyestuff of orange peel residue dyed on
cotton is higher than the colouration with β-carotene dyestuff of carrot residue. This might be
due to the higher amount of different compounds extracted from the orange peel. The
different compounds have different molecule sizes and functional groups and therefore a
different affinity to absorb on cotton fibres. Whereas in the β-carotene dyestuff of carrot
residue most compounds are - and β-carotene, which have due to the lack of functional
groups a lower affinity to cotton fibres.
Table 9: Colour coordination of the dyed cotton fabrics with orange peel extract
Orang-peel
K/S
L*
a*
b*
C*
h*
ΔE
Unmordant
0,31
90,32
-1,68
27,86
27,91
93,40
39,20
Alum post mordant
0,40
88,96
-3,16
27,49
27,67
96,54
39,25
The results for the colorimetric measurements of cotton weave fabric dyed with β-carotene
dyestuff of orange peel residue are given in table 6. The measurements were as well done
by the CIE colour system. The shade of orange peel extract dyed on cotton fabric was
greenish yellow. The cotton fabric was evenly dyed. The colour uptake (ΔE) was for the
mordanted samples again slightly higher than for the unmordanted samples. The mordant
alum increased the absorption of the dyestuff molecules on the cotton fabric. The colour
strength K/S is higher for the mordanted samples. The unmordanted samples were
compared with the post mordanted samples with alum slightly lighter due to their higher L*
value. The C* values for colour saturation were for the post mordanted samples little lower,
which means the colour saturation decreases with mordanting. Considering both values L*
and C* the shades of both unmordanted and mordanted cotton fabric dyed with orange peel
extract are light and weak in shade. The slight negative values of a* and the more positive b*
values indicate shades of greenish yellow and slightly more greenish and less yellowish for
the mordanted samples.
Master thesis by Susan Hecker
29
5.5.
Results of the wash fastness test (ISO 105 – C)
The results for the wash fastness test according to ISO 105-C are given in the table below.
Grade range from 1-5, where 1 is very bad and 5 very good.
Table 10: Results of the wash fastness test of unmordanted and post mordanted
(alum) dyed cotton fabrics of orange peel and carrot extract.
Orange peel
Carrots
Colour change
Staining
Colour change
Staining
Unmordant
1-2
4-5
1
4-5
Alum post mordant
1-2
4-5
1
4-5
The results for the colour change of unmordanted and mordanted samples dyed with βcarotene dyestuff of orange peel residue were of grade 1-2. Most of the colour was washed
away by the washing test. The samples were partially coloured after the test in much lighter
and weaker colour saturation. The results for the colour change of β-carotene dyestuff of
carrot residue dyed cotton fabrics were only a grade 1. Almost no colour was left on the
cotton samples after washing at 60°C for 30 min. Whereas the colour change gave poor
results for both β-carotene dyestuff of carrot and orange peel residue dyed on cotton fabric
the staining results were with the grades 4-5 for all samples very good. The poor results for
colour change of the washing test for both β-carotene dyestuffs of carrot and orange peel
residue dyed on cotton weave fabric is interpreted by the lack of functional groups and the
relative small molecule sizes of the β-carotene dye and their consequential low affinity
towards the textile fibres (Marx, 2011). Yi and Yoo (2010) achieved same results for orange
peel dyestuff. Their poorest washing grade was for a dye concentration of 100 % owf that
was increased to a grade 4 for a dye concentration of 800 % owf. A possibility to improve the
wash fastness properties could be the increase of dye concentration as done by Yi and Yoo
(2010).
30
Master thesis by Susan Hecker
5.6.
Results of the xenon test (ISO-Norm B02)
The xenon test samples are graded between the ranges from 1-8, where 1 is very bad and 8
is very good. The results are listed in the table below.
Table 11: Results of the xenon light fastness test of unmordanted and
post mordanted (alum) dyed cotton fabrics of orange peel and carrot
extract.
Orange peel
Carrots
Unmordant
3
2
Alum post mordant
4
3
The light fastness properties of β-carotene dyestuff of carrot residue dyed on cotton (2 for
unmordanted and 3 for mordanted samples) were poorer compared to the β-carotene
dyestuff of orange peel residue dyed samples. The results for β-carotene dyestuff of orange
peel residue were reasonable, with the grade 3 for unmordanted and 4 for mordanted
samples. The light fastness was improved by post mordanting the samples. The mordanted
samples of both β-carotene dyestuffs of carrot and orange peel residue were compared with
their unmordanted counterparts each with 1 grade better. The poorer results of the light
fastness for β-carotene dyestuff of carrot residue could be interpreted due to the high
sensitivity of β-carotene in light exposure and its higher and purer amount in this dye
compared with the β-carotene dyestuff of orange peel residue (Liaaen-Jensen, 1989). In its
natural form β-carotene is higher concentrated for example in carrots and it is surrounded by
proteins that can keep it more stable (Bauernfeind, 1981). Another reason is the small
molecule size and the resulting lower affinity towards the cotton fibres (Marx, 2011).
The research by Saleh et al. (2013) (explained in the literature on p. 10 ) showed great
promise that dyeing on cotton fabric with the dyestuff containing; chlorophyll a, and b and βcarotene extracted from banana leaves by acetone is possible. They concluded that the
dyeing resulted in high colour strength, and high fastness properties. Comparing their results
of unmordanted samples with the results in this report (also unmordanted samples) in colour
Master thesis by Susan Hecker
31
strength and fastness properties, a conflict arises. The K/S values were with 0,52 higher
compared to the results of β-carotene dyestuff of carrot residue with 0,15 and β-carotene
dyestuff of orange peel residue with 0,31. The lightness L* is lower with a value of 77,01
compared to 90,32, (β-carotene dyestuff of orange peel residue) and 85,65 (β-carotene
dyestuff of carrot residue). They concluded a light fastness of grade 4 whereas this report
graded 3 for β-carotene dyestuff of orange peel residue and grade 2 for β-carotene dyestuff
of carrot residue. The wash fastness test at 40°C gave a grade of 4 and in this report 1-2 (βcarotene dyestuff of orange peel residue) and 1 (β-carotene dyestuff of carrot residue) was
achieved. Neither high colour strength nor high fastness properties could be concluded in
this report and made the initial evidence that the dyeing on cotton with β-carotene dyestuff
could work invalid.
32
Master thesis by Susan Hecker
6. Conclusions
The work showed that the organic solvent extraction of β-carotene from orange peel and
carrots residue from the juice pressing industry is possible. The three different organic
solvents; ethyl acetate, petroleum ether and hexane/acetone mixture were chosen for the
extraction of β-carotene. Amongst them petroleum ether was slightly better for the βcarotene extraction from both orange peel and carrot residue due to its strong non-polar
character.
The dyeing with β-carotene dyestuff of carrot and orange peel residue is only reasonable
possible and might not be an alternative for the dyeing industry, due to its poor fastness
properties and the unstable character of the β-carotene molecule to light and heat exposure.
The lack of functional groups in the β-carotene molecule made it difficult or impossible for it
to absorb on the cotton fibres and to achieve fair fastness properties. The used mordant
alum could increase the light fastness of both dyes with 1 grade, but not the wash fastness.
In general the further use of residues of the juice pressing industry for textile dye extraction
might be a valuable source for textile dyestuffs. The further usage of the residues might be a
value-adding step for the waste that would otherwise only be used as fodder or biogas
production.
Master thesis by Susan Hecker
33
7. Future research
The work showed that dyeing with β-carotene dyestuff of carrot and orange peel residue
from the juice pressing industry is only reasonable possible. The lack of functional groups in
the β-carotene dyestuff molecule makes the dyeing on cotton fabric difficult. An HPLC
analysis of the dyed cotton fabric could detect, which of the extracted compounds were really
absorbed on the fabric. It would be interesting to investigate if other fibres can improve the
fastness properties. Fibres such as wool (protein fibres) that are commonly used for the
dyeing with natural extracts due to their acid and alkaline functional groups, polypropylene
that has a low Tg at 35°C and is mostly suitable for fat- and oil soluble dyestuffs or polyester
(dyed with carriers to keep a low dyeing temperature).
The extraction of other compounds from residues of the juice pressing industry, which own
more functional groups, might be possible for the further work. Compounds such as
xantophylls (lutein or zeaxanthin) that belong to the group of carotenoids with hydrophilic
character could be a better dyestuff for cotton dyeing than β-carotene containing dyestuffs
with hydrophobic character.
34
Master thesis by Susan Hecker
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Master thesis by Susan Hecker
37
APPENDIX
Master thesis by Susan Hecker
I
APPENDIX I
Aqueous extraction
a) Carrots
20 g carrots (ground in the Kenwood blender until the pieces were ca. 1 mm)
200 ml tap water

All together was put in an Erlenmeyer flask and heated up in a water bath to 100°C
for 1 h.

Stirring with magnetic stirring at level 7.

After 1 hours filtration (Büchner funnel)
Rotary evaporator:

Evaporation of the water in the rotary evaporator at 40°C.

Storage of the residue (oily, caramelized liquid) in small glass bottles in no light.
b) Orange peel
20 g orange peel (ground in the Kenwood blender until the pieces were ca. 1 mm)
200 ml tap water

All together was put in an Erlenmeyer flask and heated up in a water bath to 100°C
for 1 h.

Stirring with magnetic stirring at level 7.

After 1 hours filtration (Büchner funnel)
Rotary evaporator:
II

Evaporation of the water in the rotary evaporator at 40°C.

Storage of the residue (oily, caramelized liquid) in small glass bottles in no light.
Master thesis by Susan Hecker
Organic solvents extraction with ethanol
a) Carrots
20 g carrots (as got from Brämhults juice; ground state)
200 ml 80 % aqueous ethanol solution (800 ml ethanol: 200 ml H2O)

All together was put in an Erlenmeyer flask and heated up in a water bath to 60°C
for 4 h.

Stirring with magnetic stirring at level 7.

After 4 hours filtration (Büchner funnel)
Rotary evaporator:

Evaporation of the solvent (ethanol) in the rotary evaporator at 40°C.

Solve the residue with distilled water and evaporate the water again in the rotary
evaporator.

Storage of the residue (oily, caramelized liquid) in small glass bottles in no light.
b) Orange peel
20 g orange peel (ground in the Kenwood blender until the pieces were ca. 1 mm)
200 ml 80% aqueous ethanol solution (800 ml ethanol: 200 ml H2O)

All together was put in an Erlenmeyer flask and heated up in a water bath to 60°C
for 4 h.

Stirring with magnetic stirring at level 7.

After 4 hours filtration (Büchner funnel)
Rotary evaporator:

Evaporation of the solvent (ethanol) in the rotary evaporator at 40°C.

Solve the residue with distilled water and evaporate the water again in the rotary
evaporator.

Storage of the residue (oily, caramelized liquid) in small glass bottles in no light.
Master thesis by Susan Hecker
III
Results of the HPLC analysis
The following table and graphs show the result of the HPLC analysis of β-carotene by
aqueous and ethanol extraction. For both solvents the β-carotene content was not
extracted or too low for the detection in the visible light at a wavelength of 450 nm.
Table 12: β-carotene content in carrots and orange peel (μg/g)
Solvent
Carrots Orange peel
Water
0
0
80 % aqueous ethanol solution 0
0
Figure 16: HPLC from carrot extract in aqueous solution at 450 nm
Figure 17: HPLC from orange peel extract in aqueous solution at 450 nm
Figure 18: HPLC from carrot extract in 80 % aqueous ethanol solution at 450 nm
IV
Master thesis by Susan Hecker
Figure 19: HPLC from orange peel in 80 % aqueous ethanol solution at 450 nm
Master thesis by Susan Hecker
V
APPENDIX II
Chemical structures of the solvents used
Chemical structure of ethyl acetate:
Chemical structure of acetone:
Chemical structure of hexane:
VI
Master thesis by Susan Hecker
APPENDIX III
Vat dye
Mercerized and bleached cotton was dyed as vat dye with the following parameters. First
the stock vat was made in a warm water bath to solve the sodium hydrosulphide. Then it
was add to the dye bath together with the fabric. Under magnetic stirring it was left for 10
min, then it was oxidized in the air for around 5 min. This procedure was repeated 2-3
times. All four samples didn´t show a recognizable colouration.
Table 13: Different parameters for vat dyeing on cotton fabric
Sample 1
Sample 2
Sample 3
Sample 4
Residue
Solvent
Carrot
Orange peel Carrot
Orange peel
Ethyl acetate Ethyl acetate Hexane/acetone Hexane/acetone
Cotton sample weight
2,9 g
1,7 g
0,4 g
0,5 g
Organic solvent (ethyl
acetate or
hexane/acetone (1:1 v/v)
β-carotene
Water 50°C
Sodium hydroxide 25 %
Sodium hydrosulphide
4 ml
1,3 mg/g
10 ml
1 ml
0,6 g
4 ml
0,014 mg/g
10 ml
1 ml
0,6 g
4 ml
1,5 mg/g
10 ml
1 ml
0,6 g
4 ml
0,018 mg/g
10 ml
1 ml
0,6 g
Dye bath
Water ca. 20°C
Sodium hydroxide 25%
Sodium hydrosulphide
Sodium chloride
Stock vat
300 ml
0,1 ml
0,1 g
10 g
ca. 15 ml
100 ml
0,1 ml
0,1 g
10 g
ca. 15 ml
100 ml
0,1 ml
0,1 g
15 g
ca. 15 ml
50 ml
0,1 ml
0,1 g
5g
ca. 15 ml
Stock vat
Heated in water
bath at ca. 60°C
Master thesis by Susan Hecker
VII
APPENDIX IV
HPLC results
The following table 20-31 show the results of the HPLC for the analysis of β-carotene. The
retention time is around 56 min. for all samples and the area is between 32.635 – 162.668
for orange peel and 8.634.783 – 16.041.328 for carrot. There are two samples of each
solvent. In the report the calculations for the concentration and the used β-carotene for
dyeing was done based on the average value of this two samples.
Table 14: Results of the determination of β-carotene by HPLC
Sample
1
7
3
9
2
8
Residue
Orange peel
Orange peel
Orange peel
Orange peel
Orange peel
Orange peel
Solvent
Ethyl acetate
Ethyl acetate
Hexane/acetone
Hexane/acetone
Petroleum ether
Petroleum ether
Retention
Time
55,40
56,22
56,62
56,06
56,65
56,10
Area
32.635
152.876
162.668
86.549
111.816
142.022
Concentration
(mg/mL)
0,001
0,004
0,004
0,002
0,003
0,003
4
10
6
12
5
11
Carrot
Carrot
Carrot
Carrot
Carrot
Carrot
Ethyl acetate
Ethyl acetate
Hexane/acetone
Hexane/acetone
Petroleum ether
Petroleum ether
56,45
56,07
56,32
56,03
56,31
56,03
8.634.783
9.135.152
10.856.273
10.454.314
10.747.621
16.041.328
0,207
0,219
0,260
0,247
0,258
0,393
The HPLC graphs of each sample at the wavelength 450 nm are the following:
Figure 20: HPLC graph of sample 1 β-carotene from orange peel, extracted in
ethyl acetate
VIII
Master thesis by Susan Hecker
Figure 21: HPLC graph of sample 7 β-carotene from orange peel, extracted in
ethyl acetate
Figure 22: HPLC graph of sample 3 β-carotene from orange peel, extracted in
hexane/acetone
Figure 23: HPLC graph of sample 9 β-carotene from orange peel, extracted in
hexane/acetone
Figure 24: HPLC graph of sample 2 β-carotene from orange peel, extracted in
petroleum ether
Master thesis by Susan Hecker
IX
Figure 25: HPLC graph of sample 8 β-carotene from orange peel, extracted in
petroleum ether
Figure 26: HPLC graph of sample 4 β-carotene from carrot, extracted in ethyl
acetate
Figure 27: HPLC graph of sample 10 β-carotene from carrot, extracted in
hexane/acetone
Figure 28: HPLC graph of sample 6 β-carotene from carrot, extracted in
hexane/acetone
X
Master thesis by Susan Hecker
Figure 29: HPLC graph of sample 12 β-carotene from carrot, extracted in
hexane/acetone
Figure 30: HPLC graph of sample 5 β-carotene from carrot, extracted in
petroleum ether
Figure 31: HPLC graph of sample 11 β-carotene from carrot, extracted in
petroleum ether
Master thesis by Susan Hecker
XI
APPENDIX V
Spectrophotometric and fastness property results
The following tables 12-14 show the results of all samples for the spectrophotometric
measurements of colour properties, the wash fastness test after ISO 105-C and light
fastness after ISO Norm B02. The values mentioned in the report are the average value of
the two samples.
Table 15: Colour coordination of the all dyed cotton fabrics with carrot extract
Carrot
K/S
L*
a*
b*
C*
h*
ΔE
Unmordant Sample 1
0,08
89,10
9,25
11,97
15,13
52,32
24,64
Unmordant Sample 2
0,22
82,21
7,48
12,29
14,39
58,68
27,34
Alum post mordant Sample 1
0,15
87,83
6,27
14,42
15,73
66,52
26,70
Alum post mordant Sample 2
0,15
88,68
6,34
15,14
16,42
67,28
27,14
Table 16: Colour coordination of the all dyed cotton fabrics with orange peel extract
Orange peel
K/S
L*
a*
b*
C*
h*
ΔE
Unmordant, sample 1
0,33
90,72
-2,19
29,61
29,69
94,23
40,92
Unmordant, sample 2
0,29
89,91
-1,17
26,11
26,14
92,57
37,49
Alum post mordant, sample 1
0,40
89,03
-3,39
27,50
27,71
97,02
39,28
Alum post mordant, sample 2
0,41
88,89
-2,92
27,48
27,64
96,07
39,22
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Master thesis by Susan Hecker
Table 17: Results of the wash fastness test of all unmordanted and post mordanted (alum)
dyed cotton fabrics of orange peel and carrot extract.
Orange peel
Carrots
Colour change
Staining
Colour change
Staining
Unmordant, sample 1
1-2
4-5
1
4-5
Unmordant, sample 2
1-2
4-5
1
4-5
Alum post mordant, sample 1
1-2
4-5
1
4-5
Alum post mordant sample 2
1-2
4-5
1
4-5
Table 18: Results of the light fastness test of all unmordanted and
post mordanted (alum) dyed cotton fabrics of orange peel and carrot
extract.
Orange peel
Carrots
Unmordant, sample 1
2-3
2
Unmordant, sample 2
4 colour darkend
2
Alum post mordant, sample 1
4
3
Alum post mordant sample 2
4
3
Master thesis by Susan Hecker
XIII