BIOCOLOURS The New Generation Additives-1 (Final)

BIOCOLOURS; The
New Generation Additives
By Vinish & Vasanthakumar

ABSTRACT
Biocolorants are those colouring agents, which are obtained from the
biological sources,Biocolors can be classified into three main classes: natural
colours,browning colours and additives. There are various microbial sources
of biocolour as carotene production by a Pigmented strain of bacterium
Bacillus sp., carotenoid Astaxanthin produced by yeast phaffia xhodoxyma
which are considered as an important source of the natural pigment for
colouring foods. Biocolours as additive can be used in various industries as in
pharmauceutical industry, textile industy, dairy industry etc. Technological
limitations are the major bottleneck for the commercial exploitation of the
microbes for biocolour production while designing of proper bioreactors would
help to ease out the situation.
INTRODUCTION
Colour is one of the most important qualities of foods. In the past, consumers did not
care about the kind of pigments used in food colouring (natural or synthetic). But with
reference to food colorants recently there is an aversion towards synthetic pigments
owing to the belief such as "synthetic pigments are associated with several illnesses"
and "natural pigments have pharmacological benefits" (Clydesdale,1993). Natural
organic pigments are generally extracted from fruits, vegetables,seeds, roots and
microorganisms and they are sometimes called biocolours because of their
biological origin (Pattnaik et al., 1997). Biocolours that are permitted for human foods
are very limited and there is difficulty in getting approval for new sources for the
reason that the U.S. Food and Drug Administration (FDA)
CLASSIFICATIONS
1 Natural colours
2 Browning colours
3 Additives
1.NATURAL COLOURS
The principal natural colours used as additives are the

green pigment chlorophyll, the carotenoids which give
yellow to red colours and the flavonoids with their
principal subclass the anthocyanins, which impart red to
blue colours to flowers and fruits. In recent years, there
has been much interest in carotenoids, especially –
carotene as it is converted in the body to vitamin A and
has antioxidant properties. It has a beneficial effect in
reducing the risk of some cancers and perhaps heart
diseases. It can be produced commercially using
microorganisms like Dunaliella salina and Blakeslea
trispora.
2. BROWNING COLOURS
These are produced during

cooking and processing and thus
may not be of any direct
importance in foods. For e.g., as
produced during sugar
caramelization, baking etc.
3.ADDITIVES
Food additive colours are based on

anthocyanins derived from sources such as
red grapes or beet but the first additive colour
were synthetic dyes which were extensively
used as food colorants in nineteenth century
and early 1900 s. Anthocyanins are
polyphenolic group of compounds which have
been named Vitamins of the 21 st Century
due to their impressive medical and health
benefits.
APPLICATIONS
There are various applications of Biocolours in different industries as

described below:
A Pharmaceutical industry
Hepatitis C virus (HCV) infects approximately 170 million people worldwide, and is often associated with
chronic hepatitis, leading to liver cirrhosis and hepatocellular carcinoma (Brown, 2005). Currently,
interferon (IFN) and the nucleoside analogue ribavirin are used as the standard therapy to treat chronic
HCV infection. However, IFN- alone or in combination with ribavirin often leads to a range of side effects.
The 65 kDa HCV NS5B protein has RNA- dependent RNA polymerase (RdRp) activity and is key player
in HCV RNA replication. Moncascus purpureus produces pigments that are responsible for inhibiting
hepatitis C virus replication by interfering with viral RNA polymerase activity (Sun et al., 2011). A group of
Monascus orange pigment
(MOP) derivatives effectively inhibited NS5B RdRp activity and interfered with the mevalonate synthesis
pathway, thereby suppressing HCV replication in cells. A double-hit strategy, including inhibition of HCV
RdRp activity and interference with the mevalonate synthetic pathway, to inhibit HCV amplification may
provide the basis for successful antiviral therapy using the MOP AADs derived from this microbial
secondary metabolite.
B Dairy industry
Among the various pigment-producing microorganisms,

Monascus sp. Is reported to produce non-toxic pigments,
which can be used as food colorant. Besides a colouring
agent it enhances the flavour of the food and acts a food
preservative. Monascus ruber has been used widely in
the preparation of flavoured milk by utilization of rice
carbohydrate for its metabolism and production of
secondary metabolite namely pigment. Red, orange and
yellow pigments are produced using solid- state
fermentation and rice broken as a substrate. Monascus
Fermented Rice (MFR) 1.2 % is used in the preparation
of flavoured milk.
C Fish industry
Aquaculture is a rapidly growing global industry, comprising

cultivation of various freshwater and marine species of finfish,
shellfish, molluscs and ornamental fish (Garcia and Maurilio,
2013). Pigmentation is one of the important quality attributes of the
aquatic animal for consumer acceptability. As fishes cannot
synthesize their own colouring pigments de novo, the colouring
agents which are synthesized by some plants, algae and
microorganisms, need to be incorporated in the diet (Johnson and
An, 1991). Most promising pigment proved to be successful
in enhancing skin colour is Astaxanthin.Commercially available
products of astaxanthin (carotenoid) rich yeast Phafia rhodozyma
and fermentation product of Xanthophyllomyces dendrorhous has
been widely used. Microalgae Chlorella vulgaris imparts yellow
blue hues, yielding both muscle and skin pigmentation effects.
D Textile industry
The textile industry discharges large proportion of effluent that

mainly consists of synthetic dyes. Synthetic dyes have been
extensively used in the textile industries due to their ease and
cost effectiveness in synthesis, high stability towards light,
temperature and technically advanced colours covering the whole
colour spectrum. However, these synthetic dyes are often toxic,
mutagenic and carcinogenic leading to several human health
problems such as skin cancer and allergic reactions
(Srikanlayanukul et al., 2006; Gurav et al., 2011). Thus, the
worldwide demand for the dyes of natural origin is increasing
rapidly in the textile industry.
E Printing industry
It has become important to reuse and recycle

used papers in offices, etc. for the purpose of
the conservation of forest resources and
reduction of wastes. Reuse of papers in offices
is preferable, but it requires that letters and
images easily disappear from printed papers.
Decolorable ink for inkjet printing contains a
Monascus pigment. The Monascus pigments
are easily discoloured and finally lose their
colours by the irradiation of visible and/or
ultraviolet light.
METHODS OF EXTRACTION OF PIGMENTS
Solvent extraction is the conventional method that is followed to

extract colours. Anthocyanin and betalain pigments, which are water
soluble, are extracted from the raw material with water and
sometimes with aqueous methanol.For carotenoids extraction,
hexane is the solvent of choice. After thorough extraction, the
extract is concentrated and subjected to purification steps by using
column chromatography. Identification and quantification of the
pigment is performed by spectrophotometry or by high pressure
liquid chromatography
Salient features of some important biocolours
PIGMENT SOURCE COLOUR SHADE REMARKS
β-carotene Dunaliella salina, Yellow to orange depending It is sparingly oil soluble andcomprises of all
Euglena, Blakeslea upon colour formulations the trans isomers and possesses pro vitamin A
trispora activity
Astaxanthins Haematococcus Pluvialis Orange pink to red Astaxanthin belong to the Carotenoids family.
Astaxanthin can protect against chemically
induced cancers and is very strong
antioxidant.
Phycobiliproteins Algae belonging to Red and blue These have good long term stability when
Rhodophyta and stored refrigerated (2- 5°C) as ammonium
Chlorophyta sulphate precipitates. These are relatively
stable at room temperature and neutral pH.
Monascus Monascus purpureus Yellow, orange and red Pigment production and quality is good when
pigments and M. anka the organism is provided with carbon source
such as maltose, fructose and glucose and
yeast extracts as nitrogen source. Pigments
are stable to pH change in temperatures.
For carotenoids extraction, hexane is the solvent of choice. After
thorough extraction, the extract is concentrated and subjected to
purification steps by using column chromatography. Identification and
quantification of the pigment is performed by spectrophotometry or by
high pressure liquid chromatography (Naidu and Sowbhagya, 2012).
The current advance techniques followed
in colour extraction are as follows:
1. High Hydrostatic Pressure (HHP) and Pulsed Electric Field (PEF):
These methods are environment friendly and energy efficient

technologies that enhance mass transfer processes within cellular
tissues, as the permeability of cytoplasmic membranes can be
increased which in turn enhances extraction of valuable cell
components. PEF is reported to be an ideal method to enhance juice
production, increase the extraction of valuable components better
than the yields obtained by enzymatic maceration (Mason and
Zhao,1994).
2. Sonication-assisted extraction:
It is one of the most commonly used methods to enhance mass
transfer phenomena by cavitation forces, where bubbles in the
liquid/solid extraction can explosively collapse and generate
localized pressure, causing plant tissue rupture and improving the
release of intracellular substances into the solvent. (Nayak et al.,
2007).
3. Gamma irradiation:
Gamma-irradiation, as a pre-treatment to a plant material,
increases cell wall permeabilization, resulting in enhanced
extraction of cell constituents in higher yield (Sowbhagya and
Chitra, 2010).
4. Enzymatic extraction:
Enzyme assisted extraction of pigments is another new technology. Enzyme
pre-treatment cannot be a complete substitute for conventional solvent
extraction, but can result in increased yield of value added cell components and
a reduction in time of extraction and amount of solvent consumption (Rodriguez
et al., 2001).
5. Membrane technology:
Membrane processing is a fast and emerging technique for the concentration and
separation of macro and micro molecules based on molecular size and shape in
biotechnology and food processing industries (Downham and Collins, 2001).
Advantages of membrane processing are many which include improved product quality
with higher yield, utilization of by products, temperature and pH sensitive products can
easily be extracted without alteration and lastly is environmental friendly as no harmful
chemicals are being used and less energy is consumed. (Spence et al.,2010).
Future prospects:
A giant leap forward incolour production could be achieved by

combining genetic manipulation and fermentation. Technological
limitations are the major bottleneck for the commercial exploitation
of the microbes for biocolour production and designing of proper
bioreactors would help to ease out the situation.
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BIOCOLOURS; The

New Generation Additives

By Vinish & Vasanthakumar

The principal natural colours used as additives are the

These are produced during

Food additive colours are based on

There are various applications of Biocolours in different industries as

Among the various pigment-producing microorganisms,

Aquaculture is a rapidly growing global industry, comprising

The textile industry discharges large proportion of effluent that

It has become important to reuse and recycle

Solvent extraction is the conventional method that is followed to

PIGMENT SOURCE COLOUR SHADE REMARKS

1. High Hydrostatic Pressure (HHP) and Pulsed Electric Field (PEF):

These methods are environment friendly and energy efficient

A giant leap forward incolour production could be achieved by

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