Production of pigments by microorganisms.

1. PRODUCTION
OF MICROBIAL
COLORS OR
PIGMENTS
PRESENTED BY :
RITU L V

2. Introduction
•Color is the most vital attribute of
any article particularly food.
•Pigments from natural sources have
been obtained since long time ago,
and their interest has increased due
to the toxicity problems caused by
the synthetic pigments. In this way
the pigments from microbial
sources are a good alternative.

3. The various microorganisms such
as Micrococcus, Bacillus,
Rhodotorula, Monascus, Phaffia,
Sarcina and Achromobacter have
the capability to produce different
pigments. These colors have
number of beneficial properties
like anti-cancerous,
immunosuppressive, antibiotic,
anti-proliferative, bio-degradability
etc.

4. Microbial sources of natural
color
Microorganisms are the most versatile tools in biotechnology to
produce variety of molecules including enzymes, antibiotics,
organic acids and pigments. The presence of pigments has been
reported among the entire microbial world including Bacteria,
Fungi, Yeast, algae & protozoa These microorganisms can be
isolated/cultured/purified from various environmental sources such
as water bodies, soil, plants, insects and animals.

5. Production
•Advancements in fermentation
techniques have lead to the
easy production and isolation of
color pigments.
•Microbial pigments can be
produced either by
solid substrate fermentation
or
submerged fermentation.

6. Fermentation strategy
In solid substrate fermentation (SSF), the cultivation of microbial
biomass occurs on the surface of a solid substrate. This SSF technique
has many potential advantages including savings in wastewater and
yield of the metabolites.
In submerged fermentation, microorganisms are cultivated in liquid
medium aerobically with proper agitation to get homogenous growth
of cells and media component.
The influence of various process parameters such as carbon source,
temperature, pH, aeration rate on pigment production as well.

7. However, due to the high cost of using
synthetic medium, there is a need to
develop new low cost process and
extraction procedure for the production
of pigments. Efforts are on to utilize the
waste organic material for large scale
production of microbial pigments.
Some studies have focused on
production of carotenoids from waste
material such as whey, apple pomade
and crushed pasta. Such kind of waste
utilization procedures not only lower
down the production cost but also act as
potent waste management tool as well.

8. Microorganism Pigments
Achromobacter Creamy
Bacillus sp Brown
Brevibacterium sp. Orange, yellow
Corynebacterium michigannise Greyish to creamish
Pseudomonas sp Yellow
Rhodococcus maris Bluish red
Streptomyces sp. Yellow, red, blue
Microbial pigments and their colour shades
1. Bacteria

9. Microorganism Pigments
Aspergillus sp. Orange, red
A. glaucus Dark red
Blakeslea trispora Cream
Helminthosporium catenarium Red colour
H. gramineum Red
H. cynodontis Bronze colour
H. avenae Bronze colour
H. catenarin Dark maroon
Monascus purpureus Yellow, orange, red
P. cyclopium Orange
P. nalgeovensis Yellow
Microbial pigments and their colour shades
2. Molds

10. Microorganism Pigments
Cryptococus sp. Red
Phaffia rhodozyma Red
Rhodotorula sp. Red
Yarrowia lipolytica Brown
Microorganism Pigments
Dunaliella salina Red
Microbial Pigments and their colour shades
3. Yeast
Microbial Pigments and their colour shades
4. Algae

11. Monascus purpureus (fungi)
• The common names of this fungal product
are Red Yeast Rice (RYR),
• Monascus spp. produce many pigments of
industrial importance and these pigments
are mainly of three types i.e. red colorants,
orange colorants and yellowish colorants
• Monascus pigments are used in traditional
oriental foods as a natural colouring agent.
• These are extracted from M. purpureus
grown on steamed rice by solid state
fermentation.

12. Monascus fungi, can convert starchy
substrates into several metabolites
such as alcohols, antibiotic agents,
antihypertensives, enzymes, fatty
acids, flavor compounds,
flocculants, ketones, organic acids,
pigments and vitamins.
Red yeast rice

13. • .
Medium (broth) (maltose, yeast extract, glucose,
KH2P04, MgS04, CaCh)
Sterilization Medium (yeast extract, sucrose, agar)
Inoculation
Plates (5 +mm disc)
Incubation (28°C for 14 days in darkness)
Harvesting of mycelium
Mycelium coloured
Drying
Grinding
Extraction with solvent (methanol)
Shaking (1 min.)
Filtration
Drying (800
𝐶 for 24
hrs.)
Pigment Packaging
Flowsheet for the production of Monascus
pigment

14. Red Monascus pigment production by Monascus purpureus was carried out under
submerged fermentation using soybean meals as nitrogen sources to replace yeast
extracts.

15. Torularhodin
•Orange-red
•Produced by Rhodotorula spp.
(yeast)
• Antioxidant, Anti-microbial
• Apple pomace, a waste from apple
juice processing industry, which is
cheap, rich in sugar, acid and
minerals, has been employed in
pigment production.
Rhodotorula spp

16. • The production of Rhodotorula glutinis by
batch fermentation in rotatory shaker has
resulted in good yield of carotenoids. The
Rhodotorula cells are grown at 30°C for 5
days and, after centrifugation, washed
with distilled water . The optimum
conditions in the fermenter for the
carotenoid production are: fermenter
capacity, 1 litre; temperature, 30°C; air
flow rate, 1 minI ; magnetic stirrer, 220
rev/min; and, antifoaming agent, 0.3%
w/v.

17. Phaffia rhodozyma ( yeast)
•Phaffia rhodozyma ferments
glucose and other sugars to
produce pigment, astaxanthin.
The pigment is produced in
shake flask at 210
𝐶 and an
increase in glucose
concentration in the medium
gave higher yield of pigment
rather than growth only.

18. IN FOOD INDUSTRY
•β –Carotene production
known as pro-vitamin A
acts as antioxidant and has potential positive properties against
certain diseases
Following microbes are mainly using for β –Carotene production:
Blakeslea trispora
Mucor circinelloides
Phycomyces blakesleeanus

19. β-carotene from Blakeslea
trispora
Inocula of spores of mating types + and − are used in
the ratio in a suitable culture medium, such as (in g/L) d
-glucose 50, olive oil 5.4, yeast extract 1.0, l -
asparagine 2.0, KH2PO4 1.5, MgSO4 ·7H2O 0.5, BHT
0.02, and thiamine-HCl 0.005. After 8 days in aerated
culture with moderate agitation at 26 °C, the cells are
separated from the broth by filtration, partially
dehydrated and slowly extracted with a suitable
solvent such as chloroform. The solvent is separated
from the biomass and evaporated; the dry raw extract is
further processed. From 62 kg of corn steep liquor and
38 kg of glucose, there should be produced 20 kg of
biomass which, extracted with 50 L chloroform, would
give approximately and 8 kg extract containing 90 g of
β-carotene.

20. β –Carotene
Blakeslea trispora
Mucor circinelloides

21. Riboflavin (Vitamin B2) Production
• yellow food colourant
• used mainly for cereal-based
products.
• Applications - limited due to its bitter
taste and naturally slight odour.
• many microorganisms have potential to
produce riboflavin by fermentation.
• Fermentation with Ashbyagossypi.
Corn steep liquor, peptone, soybean
oil as culture medium
Ashbya gossypi

22. S.No. Producer microorganism Amount
produced
Examples
1 Weak overproducer 100 mg/L or less Clostridium acetobutylicum
2 Moderate overproducer Upto 600mg/L Candidaguilliermundiior
Debaryomycessubglobosus
3 Strong overproducer Over 1g/L Eremotheciumashbyii and Ashbyagossypi
Different level of Riboflavin production from microbes

23. Lycopene Production
• red open-chain unsaturated carotenoid
• also known as psi-carotene
• very sensitive to heat and oxidation and is
insoluble in water
• Genetically modified fungus Fusarium
sporotrichioides was used by Jones et al to
manufacture the colourant and antioxidant
lycopene.
They used the cheap corn fiber material as the
substrate.
Cultures in lab flasks produced 0.5 mg (lycopene)/g
ofdry mass within 6 days and such a production will be
increased within the next years

24. In parmaceutical industry
•Prodigiosin:
 potential pigment having many
pharmacological properties.
 broad range of cytotoxic
activity is produced by Vibrio
psychroerythrus, Serratia
marcescens, Pseudomonas
magnesiorubra, and other
eubacteria as Maltose and peanut
oil based, Powdered peanut
medium, Sucrose and peptone
based culture medium.

25. Violacein
• violet pigment violacein is an indole derivative
• isolated mainly from bacteria Chromobacterium violaceum, which
exhibits important antitumoural, antiparasitary, antiprotozoan,
anticancer, antiviral, antibacterial and antioxidant activities.

27. Factor affecting Microbial pigment
production
•Temperature
Depends on the type of microorganism
The growth of Monascus spp. requires 25-28˚C,
Pseudomonas requires 35-36˚C for its growth and pigment
production.

28. pH
•pH of the medium affects the growth of the
microbe and type of pigment production.
•Optimum pH for Monascus sp. – 5.5-6.5
• Rhodotorula is 4.0- 4.5.
•Neutral to slight alkaline pH favours lycopene
formation
• acidic pH favours β-Carotene synthesis

29. Carbon source
•The mycelial growth of pigment producing
microorganism is affected by the type of carbon
sources like glucose, fructose, maltose, lactose,
galactose, etc.

30. Type of Fermentation
•The solid state fermentation
yields 3 fold more pigment
than submerged fermentation
•In M. purpureus, yields are
superior in solid cultures
than submerged, though
media composition, pH and
agitation also affect pigment
production

31. 5. Minerals
6. Nitrogen source
Ammonium chloride is the best for production of Monascus
pigment followed by ammonium nitrate and then glutamate.
Potassium nitrate is the poorest nitrogen source, while glutamate
proved outstanding for the pigment production.
7.Moisture content
In solid state fermentation, the higher level of pigment production
by Monascusruber occurs at the 70% initial moisture level in
substrate i.e. rice
8. Aeration rate

32. APPLICATIONS

34. Application
1) Pharmaceutical industry:
• Treating various diseases. These have many
properties like antibiotic, anticancer and
immune-suppressive properties.
• Adonirubin and astaxanthin act as
nutraceuticals, prevent carcinogenesis,
prevent problems like heart attacks and
strokes.
• Monascus purpureus produce pigments which
help in the inhibition of hepatitis virus
replication by interfering with viral RNA
polymerase activity.
• Prodigiosins are known for their immune
suppressive anticancer properties.
Dairy industry:
• Monascus species are known to produce
nontoxic pigments, which can be used as
food colorants, flavour enhancers and as
a food preservative.
• Monascus ruber is used to prepare
flavoured milk by utilizing rice
carbohydrate for its metabolism and
produces pigment as a secondary
metabolite.

35. •Textile industry:
• These synthetic dyes are used in industries,
due to their easy and cheap synthesis,
stability towards light, temperature and
advanced colors covering whole color
spectrum.
• However, these synthetic dyes have many
drawbacks like toxicity, mutagenicity and
carcinogenicity properties leading to
various health problems like skin cancer and
allergies.
• Hence, consumers demand for dyes of
natural origin as colorants.

36. • Nutritional supplements - β-carotene, Riboflavin, Yellow β-
xanthins ( essential dietary amino acids)
• Food colorants: An important goal of food industry is to produce food
with an attractive appearance. Food producers are opting for natural food
colors, as artificial ones shows many negative impacts on health when
consumed.
Penicillium oxalicum is a fungus which produces red color used in cosmetic
and food industry. These colors are useful in different products like baby
foods, breakfast cereals, pastas, sauces, processed cheese, fruit drinks,
vitamin-enriched milk products, and some energy drinks, as probiotcs
and so on.

37. S.No Name of dye Source Application
1 Prodigiosin Vibrio spp. Wool, nylon, acrylics and silk
2 Red Prodigiosin Serratia marcescens Acrylics, polyester, polyester microfiber, silk
and cotton
3 Pigments from Janthinobacterium lividum Silk, cotton and wool (bluish-purple, all
natural fibers) and nylon and vinylon (both
synthetic fibers)
4 Violacein (violet pigment) Chromobacterium violaceum Pure cotton, pure silk, pure rayon, jacquard
rayon, acrylic, cotton, silk stain and polyster
5 Pink pigment Roseomonas fauriae Cotton fabrics
6 Pigments from Fusarium oxosporum,
Trichoderma viride and
Alternaria spp
Cellulosic fibers
7 Anthraquinone Dermocybe sanguine Wool fibers
8 Pigment from Trichoderma spp Wool and silk fibers
Sources and uses of dyes in textile industry

38. CONCLUSION
•Microbial pigments are not only used as food colorant,
flavoring agent and dying agents they are widely applied
in medicinal aspects.
•lower blood cholesterol concentration, Anti-Diabetic
Activity, Anti-Inflammation etc.

39. References
• http://www.ijpcbs.com/files/volume5-1-2015/24.pdf
• https://ijpsr.com/bft-article/microbial-pigments-as-a-natural-color-a-
review/?view=fulltext
• https://www.frontiersin.org/articles/10.3389/fnut.2019.00007/full
• https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4519520/
• https://www.researchgate.net/profile/Shashi_Bhushan6/publication/228730339_Mic
robial_Pigments/links/0fcfd5090a422deb9b000000/Microbial-
Pigments.pdf?origin=publication_detail