ORGANIC ACIDS

I. OVERVIEW.

II. TYPES OF ORGANIC ACIDS.

I. OVERVIEW.

 Large volumes.

 Production.
Citric acid – 900 ktons/year.
Gluconic acid – 60
ktons/year. Lactic acid – 50
ktons/year.
L-ascorbic acid – 60
ktons/year. Malic acid – 40-60
II. TYPES OF ORGANIC ACIDS
 CITRIC ACID
II. TYPES OF ORGANIC ACIDS
Citric acid.
 TCA cycle intermediate.
 Fermentation with Aspergillus
niger.
 Western Europe (41%).
 North America (28%).
 Biochemistry.
 2 moles of pyruvate from
glycolysis.
 Pyruvate to acetyl-CoA.
II. TYPES OF ORGANIC ACIDS
Citric acid.
 Aspergillus niger.
II. TYPES OF ORGANIC
ACIDS
CITRIC ACID.
II. TYPES OF ORGANIC ACIDS
Citric acid.
 Biochemistry.
 Mitochondria - acetyl-CoA +
oxaloacetate to
citrate (citrate synthase).
 Cytoplasm – pyruvate
carboxylase converts pyruvate
+ CO2 to
oxaloacetate.
 Oxaloacetate converted to
malate by malate
II. TYPES OF ORGANIC ACIDS
Citric acid.
 Biochemistry.
 Malate crosses mitochondrial
membrane.
 Citrate synthase forms
citrate.
 Citrate accumulation -
deregulated synthesis.
 Mutant isolation – A.
niger.
II. TYPES OF ORGANIC ACIDS
Citric acid.
Mutant isolation – A. niger.
Respiratory inhibitor resistance.
Fluoroacetate resistance.
Low pH.
Decreased byproduct
formation (oxalic acid &
gluconic acid).
Yeast citric acid producers.
Substrates – alkanes and
II. TYPES OF ORGANIC ACIDS
 Yeast citric acid producers.
 Candida parapsilosis ATCC 7330.
 Candida guilliermondii ATCC 9058.
II. TYPES OF ORGANIC ACIDS
Citric acid.
 Yeast citric acid producers.
 Candida species.
 Yarrowia lipolytica.
 Nutrient control of A. niger citric acid
accumulation.
 Carbon source.
o Utilizes sucrose, maltose or
glucose.
o Industrial substrates =
beet/sugar cane molasses.
o High glucose concentration (100
–
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
Citric acid.
 Nutrient control of A. niger citric
acid accumulation.
Trace metal ions.
o Fe, Zn, Cu, & Co essential
for growth.
o Mn2+, Fe3+ & Zn2+ at growth-
limiting
concentration stimulates citric acid
production.
o Cu stimulates citric acid
2+
II. TYPES OF ORGANIC ACIDS
Citric acid.
Nutrient control of A. niger citric
acid accumulation.
Trace metal ions.
o Mn2+ limitation has multiple
effects.
o Increases glycolytic carbon flux,
alters plasma membrane, impairs
DNA synthesis and respiration
and causes cell morphology
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
Citric acid
 Nutrient control of A. niger citric acid
accumulation.
 pH.
o Citric acid accumulation
produces low pH (< 2.5).
o Low pH prevents oxalic acid (no
oxaloacetate) and gluconic acid
production (no glucose oxidase).
 Dissolved oxygen tension.
oneed strong aeration.
oinduces alternate NADH reoxidation
– heat release.
II. TYPES OF ORGANIC ACIDS
Citric acid.
Nutrient control of A. niger citric
acid accumulation.
Nitrogen source.
o Ammonium salts, nitrates and
urea.
o Ammonium salts may stimulate
citric acid synthesis.
Phosphate kept at low
concentration
in medium.
II. TYPES OF ORGANIC ACIDS
Citric acid.
 Production process.
 Surface process (7-
15 days). oLow power
requirements. oHigh
reproducibility.
o100 liters aluminum trays with
flow of sterile air.
oSpores distributed on tray.
oYeast citric acid producers
II. TYPES OF ORGANIC ACIDS
 Citric acid.
 Production process.
 Submerged process usually involves
7-10 days of acid
production.
oUses large-volume fermentors.
oDifficult to control due to trace metals
and oxygen flow.
oBulbous hyphae.
 Solid-state fermentation (7-15 days of
incubation)
oProcessing wastes of fruit and
sugarcane bagasse used as
II. TYPES OF ORGANIC ACIDS
 Citric acid.
 Production process.
 Solid state fermentation (7-15 days of
incubation).
o70-80% moisture solids used for
spore inoculation.
oSolids may be moistened using a
carbon source such as glucose.
oEthanol or methanol stimulates citric
acid production by A. niger.
oCan be performed in flasks, trays or
bioreactors.
oWater extraction.
II. TYPES OF ORGANIC ACIDS
 Citric acid.
 Production process.
 Recovery.
o Broth filtration.
o Wash mycelial cake (up to
15% citric acid) several times with
water to remove citric acid from
mycelium.
o Add lime for oxalate removal.
o Precipitation with Ca(OH)2.
o Solvent extraction.
o Ion-exchange adsorption.
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
Citric acid.
 Applications.
 Beverages (45%).
 Food & sugar confectionery
(21%).
 Detergent & cleaning industry
(19%).
 Pharmaceutical industry
(8%).
 About 900,000 tons of citric acid
produced per year with most
being produced by A. niger
II. TYPES OF ORGANIC
ACIDS
 Gluconic
acid.
II. TYPES OF ORGANIC ACIDS
Gluconic acid.
 Glucose dehydrogenation by bacteria
and fungi.
 Bacteria.
o Gluconobacter.
o Acetobacter.
o Vibrio.
o Pseudomonas.
 Fungi.
o Aspergillus.
o Penicillium.
o Gliogladium.
o Aureobasidium.
II. TYPES OF ORGANIC ACIDS
Gluconic acid.
 Biochemistry.
 Bacteria.
o Utilizes glucose dehydrogenase
with pyrroloquinoline quinone as
coenzyme.
o Pentose phosphate pathway.
o High glucose (>15 mM) and low
pH represses pathway.
 Fungi (2-step process).
o Glucose oxidase oxidizes
(H2O2) glucose to
II. TYPES OF ORGANIC
ACIDS
 Gluconic acid.
Biochemistry.
Fungi (2-step process).
oGlucose oxidase oxidizes
(H2O2) glucose to
gluconolactone.
o Oxidase induced by high
glucose, aeration and
pH>4.
ogluconolactone hydrolyzed
to
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
Gluconic acid.
 Production process.
 Fermentation produces >60,000
tons/year.
 Producer organisms.
o Aspergillus niger.
o Gluconobacter oxidans.
 Media (initial pH of 6.5).
o Up to 350 g/l glucose.
o Limiting phosphate &
nitrogen levels.
o High manganese levels.
o High oxygen levels.
opH>3 and pH maintained with calcium
II. TYPES OF ORGANIC ACIDS
Gluconic acid.
 Production process.
 Recovery.
o Sulfuric acid precipitation-Ca.
o Precipitate by increasing pH to 7.5-
Na.
o Ion-exchange for free acid.
 Applications.
 Acidulant in foods/beverages.
 Pharmaceutical chelator.
 Removes deposits from metal surfaces.
 Textile sizing agent.
 Additive to concrete
II. TYPES OF ORGANIC
 LACTIC ACID.
ACIDS
II. TYPES OF ORGANIC
ACIDS
Lactic acid.
 Biochemistry.
 2 isomeric forms D & L.
 Lactic acid bacteria (Lactobacillus).
 Heterofermentative = lactic acid,
ethanol, acetic and formic acids
from glucose – hexose
monophosphate pathway.
 Homofermentative = only one
isomer
of lactic acid from glucose (used
for bulk of lactic acid production).
II. TYPES OF ORGANIC ACIDS
•Lactic acid
 Production process (85-95% yield)
 Carbon source = glucose syrup (corn
starch), maltose syrup
(maltodextrin) and lactose (whey).
 Source of nitrogen and phosphate.
 Malt sprouts (contains amino
acids and B vitamins).
 Stainless steel fermentors.
o45oC and slow stirring for 4-6 days
(anaerobic conditions).
opH 5.5-6.0 – maintained by calcium
carbonate or ammonium addition.
II. TYPES OF ORGANIC ACIDS
Lactic acid.
 Production process.
 Recovery.
o Heat calcium lactate to 80-100oC
and increase to pH 10.
o Precipitate calcium sulfate using
sulfuric acid.
o Filter and concentrate.
o Remove heavy metals by ion-exchange
or chelating agents.
o Decolorization – oxidizing agents or
activated charcoal.
 4 grades – technical (20-80%), food
(>80%),
II. TYPES OF ORGANIC ACIDS

e
II. TYPES OF ORGANIC ACIDS
 Lactic acid.
 Applications.
 Food industry (emulsifier, acidulant and
dough conditioner).
 Pharmaceutical/cosmetic industry,
intestinal treatment, moisturizer,
emulsifier.
 Polymer industry (polylactides
biodegradable plastic coating – Cargill
Natureworks®).
 60,000-65,000 tons/year produced and
food grade sells between $0.71-
$1.15/pound.
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC
ACIDS
 Itaconic
acid.
II. TYPES OF ORGANIC ACIDS
Itaconic acid.
 Biochemistry.
 Synthesized by Aspergillus terreus or
Aspergillus itaconicus and excreted.
 Enzyme aconitate dehydrogenase
forms itaconitate from cis-aconitate in
cytoplasm.
 Excretes acid into medium.
 Production (up to 85% yield).
 Carbon sources =
glucose syrup, hydrolyzed starch
or molasses.
 High oxygen tension.
II. TYPES OF ORGANIC ACIDS
 Itaconic acid.
 Production.
 Metal ion limitation by Cu
ions or ferric hexacyanoferrate.
 pH between 2.8-3.1.
 Surface, submerged or solid-state
fermentation.
 5 days at 39-42 o C.
 Recovery.
 Evaporation, activated carbon
treatment, crystallization and
recrystallization.
 Grades.
o Refined (higher quality).
o Industrial.
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
 Ascorbic acid (vitamin C).

 Production (60% yield).

 5 synthetic steps and 1 bioconversion
step.
 Acetobacter xylinum converts D-
sorbitol
to L-sorbose at 30-35oC at pH 4-6.
 80,000 tons/year (3-4% growth).
II. TYPES OF ORGANIC ACIDS
II. TYPES OF ORGANIC ACIDS
 Malic acid.

 Aspergillus niger, Aspergillus wentii.

 Thin stillage as a substrate
using surface fermentation.
 Biochemistry: Malate
dehydrogenase.
 Applications include use in the food
and beverage industries as well
as in
metal cleaning, pharmaceuticals and
II. TYPES OF ORGANIC
ACIDS
 Other organic acids.
II. TYPES OF ORGANIC
ACIDS
 Other organic acids.

Tartaric acid

Fumaric acid

trans-2,3-epoxysuccinic
acid
II. TYPES OF ORGANIC ACIDS
 Other organic acids.

Succinic acid

Kojic acid

Gallic acid