Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)

M Moresi, Universit della Tuscia, Viterbo, Italy

E Parente, Universit della Basilicata, Potenza, Italy; and Istituto di Scienze dellAlimentazione, Avellino, Italy
2014 Elsevier Ltd. All rights reserved.

Several organic acids are used in a variety of food and nonfood

applications. Table 1 lists the main acids that are produced
commercially by chemical (C) or biotechnological (fermentation, F, or enzymatic, E) methods or extracted from winemaking residues (L).
Citric, acetic, lactic, propionic, tartaric, fumaric, and malic
acids are among the most versatile ingredients in the food and
beverage industry because of their valuable properties, such as
solubility, hygroscopicity, acidity, buffering capacity, and
chelation (see Preservatives: Traditional Preservatives Organic
Acids).
Citric acid accounts for around 80% of the food acidulant
usage, whereas the use of phosphoric or acetic acids is limited,
being almost exclusively utilized in cola soft drinks or in
vinegar (see Vinegar), sauces, and condiments, respectively.

Citric Acid
Citric acid (2-hydroxy-1,2,3-propanetricarboxylic acid: C6H8O7)
is widely distributed in natural raw materials (such as lime,
lemon, and raspberry) and is commercially available in the
monohydrated form (molecular mass of 210.13 Da, relative
density of 1.542 at 20
C, and heat of combustion of
1962 kJ mol1 at 25
C). It is a strong tricarboxylic acid
(TCA; its dissociation constants being K1 7.45  104,
K2 1.73  105, and K3 4.02  107 at 25
C), highly
soluble in water with pleasant acid taste.
Citric acid was rst isolated in 1784 by Scheele, who
precipitated it as calcium citrate by adding calcium
hydroxide (lime) to lemon juice. Before 1920, it was almost
exclusively produced in Sicily by pressing lemons: The rm
Arenella (Palermo, Italy) essentially established a monopoly
until the advent of the citric acid fermentation technique in

Table 1

Belgium (Societ des Produits Organiques de Tirlemont) in

1919 and in the United States (Chas. Pzer & Co., New
York) in 1923.
About 10 years later, about 80% of the worlds citric acid
was produced by the surface fermentation process. The
submerged fermentation process began to be applied only after
World War II.
From 1950 to 1980, citric acid was mainly used in pharmaceutical or health products. In fact, in the early 1980s, its
two largest manufacturers were Pzer and Miles/Bayer, both
suppliers of prescription drugs. Thereafter, as citric acid began
to be used in the food and beverage sector in industrial and
developing countries, its market size experienced signicant
growth and several new manufacturers were established in
Europe and North America, as well as in China where several
small-scale fermentation units have produced citric acid from
sweet potatoes or cassava since the 1970s.
In the early 1990s, a few manufacturers gave rise to the socalled citric acid cartel. The overcharges imposed on US buyers
was estimated in the range of $116309 million and, on
January 29, 1997, Haarmann & Reimer Corp., a subsidiary of
Bayer AG (D), pled guilty and paid a $50 million criminal ne.
In March 1998, even Archer Daniels Midland Co. (ADM)
agreed to pay $36 million to four citric acid customers that had
opted out of the July 1997 civil class-action antitrust settlement. At that time, the global citric acid capacity was about
840 000 Mg (mega grams) per year with a growth rate of 5%
per year. Afterward, the world citric industry became less
concentrated and numerous new manufactures, especially in
China, as well as Brazil, India, Indonesia, and Thailand, have
entered the market, thus making the formation of cartels less
probable.
Moreover by the early 2000s, almost all citric acid
manufacturing was globally integrated into the corn wet-milling

Main organic acids: molecular formulas, world output, production methods, and organisms

Acidulant

Chemical formula

World output (metric tons)

Production methods a,b

Organism

Acetic acid (vinegar)

Lactic acid

C2H4O2
C3H6O3

190 000
150 000

F 100%
F 100%

Propionic acid
Fumaric acid
Malic acid

C3H6O2
C4H4O4
C4H6O5

130 000
12 000
10 000

Tartaric acid
Itaconic acid
Citric acid
Gluconic acid

C4H6O6
C5H6O4
C6H8O7
C6H12O7

28 000
15 000
1 800 000
87 000

C 100%
C 100%
C 70%
E 30%
L 100%
C 100%
F 100%
F 100%

Acetobacter aceti
Lactobacillus spp.
Rhizopus spp.
Propionibacterium acidipropionici
Rhizopus arrhizus

Aspergillus terreus
Aspergillus niger
Aspergillus niger

Note: Because of the lack of published data, the production gures are approximate.
a
Percentage of total production for food uses.
b
F, fermentation; C, chemical synthesis; E, enzymatic synthesis; L, leaching.

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FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)
industry either by acquisition (Pzer and Miles/Bayer were
bought by ADM and Tate & Lyle, respectively) or by new process
development (Cargill).
In the years 198789, US list prices for citric acid anhydrous
remained unchanged at $1.79 kg1. By late 1989, the list price
reduced to $1.65 kg1, while in the fall 1990 it was as low as
$1.39 kg1. Then, thanks to the citric acid conspiracy in the
years 199396, the citric acid cartel accomplished its main goal
of raising and keeping list price at $1.87 kg1. Then, it lowered
from $1.76 kg1 in November 1994 to $1.54 kg1 in the early
1997.
Thereafter, the severe competition resulted in selling prices
of anhydrous citric acid decreasing to $0.70$0.80 kg1, thus
forcing the smaller manufacturers, unable to benet from the
economy of scale, to exit the business. As a consequence, the
panorama of organic acid manufacturers has profoundly
changed over the last decade. In 2010, China approximately
accounted for more than 50% of global citric acid production
capacity, while Europe and North America covered the 19
and 24%, respectively, and as much as 6570% of global
consumption.
Several substrates are used as fermentation substrates
depending on the local availability. Maize starch is mainly used
in the United States and China, while sugarcane or sugar beet
molasses prevail in the Brazilian and Indian or European
markets, respectively.
Cellulosic materials are currently unused in citric acid
production, even if there are projects to assess the technical
feasibility of such feedstock materials in the citric acid
industry.
From January 2008 to January 2009, export prices of
Chinese (anhydrous) citric acid oscillated in the range of US
$0.7$0.8 kg1; thereafter, they steadily increased to reach
a peak of $1.1 kg1 in June 2011, as a direct result of the
increase in the market prices for agricultural raw materials,
particularly corn. The present economic crisis in Europe and the
United States has newly reduced the market prices of (anhydrous) citric acid to US $0.70$0.96 kg1 depending on the
amount ordered.
In conclusion, the global citric acid production capacity
reached almost 1.8 million metric tons (Mg) in 2010, while
it was about 1.5  106 Mg in 2005.

Organisms and Metabolic Pathways Involved

Several molds (Penicillium spp., Aspergillus niger, Aspergillus
wentii, Trichoderma viride; see Aspergillus and Penicillium andTalaromyces: Introduction), yeasts (Yarrowia lipolytica, Candida
guillermondii), and bacteria (Arthrobacter) produce citric acid
from a variety of substrates (glucose, sucrose, n-alkanes), but
industrial processes have been developed only for the
production of citric acid from sugars (glucose, sucrose) with
A. niger and from sugars and n-alkanes with yeasts. Industrial
strains are not freely available, but citric acidproducing
strains (A. niger, NRRL 2270, NRRL 599, ATCC 11414, ATCC
9142; Y. lipolytica ATCC 20346, ATCC 20390, NRRL Y-7576,
NRRL Y-1095) can be obtained from international culture
collections.
Metabolic pathways involved in citric acid overproduction
by A. niger are shown in Figure 1.

805

A high ux through the glycolysis, decreased activity of TCA

cycle reactions that degrade citrate, and an anaplerotic reaction
to replenish the oxaloacetate (OAA) used for the synthesis of
citrate are all essential (see Metabolic Pathways: Release of
Energy (Aerobic)). Key regulatory steps in the process include
glucose transport and phosphorylation, citric acid export from
the mithocondria and cell, phosphofructokinase, pyruvate
carboxylase (PC), citrate synthase (CS), and a-ketoglutarate
dehydrogenase (KDH).
The metabolic changes necessary for citric acid overproduction in A. niger are induced by high sugar concentration,
low pH, and manganese (Mn2) deciency. Other factors (i.e.,
phosphate and nitrogen concentrations, high dissolved oxygen
(DO) concentration, trace metals), however, are important.
Very low concentrations of Mn2 (<10 mg m3) are critical.
They result in decreased activity of the pentose phosphate
pathway and increased glycolytic ux, increased intracellular
NH
4 pool and turnover of nucleic acids and proteins, changes
in membrane lipid composition and cell wall composition,
and morphological changes.
Improvement of strains for citric acid production traditionally has been carried out by mutagenesis and screening.
Overexpression of proteins critical to acidogenesis (hexokinase,
glucose carrier) or inactivation of genes encoding enzymes that
produce allosteric inhibitors of hexokinase has been attempted,
but with limited success, in the additional production of citric
acid. It has been postulated that the activity of seven glycolytic
enzymes needs to be increased to obtain increased production
of citric acid. The availability of the complete genome sequence
of A. niger is likely to allow for the design of overproducing
mutants.
Citric acid overproduction in yeast is relatively insensitive to
trace metals concentration and is triggered by nutrient (N, S, P,
or Mg) limitations coupled with a high rate of glucose
utilization, which results in a high adenosine triphosphate/
adenosine monophosphate ratio and, in turn, in inactivation of
nicotinamide adenine dinucleotide (NAD)-specic isocitrate
dehydrogenase (IDH). The main anaplerotic reactions include
the synthesis of OAA from pyruvate catalyzed by PC during
production from glucose and the glyoxylate cycle during
growth on n-alkanes. Accumulation of isocitrate (1050% of
the citrate produced) in excess with respect to the predicted
equilibrium of aconitase probably is due to the high permeability of yeast mitochondria to isocitrate compared with
citrate. Low cytoplasmic levels of citrate may be responsible for
reduced feedback inhibition of glycolysis. Improvement of
yeast for citric acid production is directed to obtain strains with
reduced isocitrate dehydrogenase and aconitase activities.

Methods of Manufacture
Citric acid production is mainly accomplished by the
submerged fermentation process, probably because of the
smaller contribution of investment and labor costs to its overall
production costs. The surface fermentation process currently
accounts for only 510% of the world supply. In Europe, all
surface fermentation plants have been shut down during the
past decade. Smaller amounts of citric acid (<1%) are reported
to be extracted from citrus fruits in Mexico and South America
and to be produced by the solid-state process in Japan.

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FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)

807

Both submerged- and surface-culture fermentation processes use beet molasses or glucose syrup as the main raw
material and use A. niger as the fermenting organism.
Submerged fermentation is carried out either in 150200 m3
stirred-tank reactors or in 300500 m3 (up to 1000 m3 as claimed
by some manufacturers) bubble-column reactors. The main
advantages of these techniques are improved asepsis during
fermentation, automatic control of inoculation and fermentation
procedures, shorter fermentation times, and greater product
yields.
In spite of the old and renewed interest in citric acid
production by yeast grown submerged in sugar- or hydrocarbon-based media to overcome the main disadvantages of
traditional mold fermentation (i.e., high sensitivity to trace
metals and low production rates), no yeast-based process is
currently known to be operating worldwide.

Table 2
Composition for the production media used in the
laboratory- and industrial-scale production of citric acid by A. niger

Production Media

Media sterilization is carried out batch wise at 121
C for

1530 min at the laboratory or pilot scale or continuously
using a plateheat exchanger unit at the industrial scale.

Citric acid is produced from media containing high concentrations of simple sugars (molasses or glucose syrup) (see
Fermentation (Industrial): Media for Industrial Fermentations), in which mycelium growth is restrained by nutrient
(phosphorous, manganese, iron or zinc) limitation. Their range
of composition is given in Table 2.
Nitrogen is usually added as ammonium nitrate or sulfate.
Metals are removed by pretreatments of raw materials, especially molasses, with cation-exchange resins or the addition of
potassium hexacyanoferrate (HCF). The optimal iron concentration seems to depend on the fungal strain, but iron levels of
200 mg m3 were found to inhibit citrate production. The
inhibitory effect of Fe can be counterbalanced by the addition of copper and zinc salts during the inoculum development
or during early mycelium growth in the production medium.
Manganese concentration has to be kept as low as possible
(<10 mg m3).
Some ingredients (methanol, 36% w/v; corn, peanut, and
olive oils, 0.10.5% w/v; starch, 0.0250.5% w/v) have been
claimed to enhance the citric acid yield.

Component
Sucrose or glucose
NH4NO3 (or other
NH
4 salt)
KH2PO4
MgSO47H2O
Fe2
Zn2
Cu2
Mn2
Initial pH

Range of
concentrations

Typical values

Unit

125225
0.53.5

180
1.5

kg m3
kg m3

0.52
0.12.0
21300
02900
110 200
046
2.56.5

0.5
0.25
<200
2001500a
2001500a
<2
2.2

kg m3
kg m3
mg m3
mg m3
mg m3
mg m3

To overcome the detrimental effects of iron and manganese on mycelium structure

and restore proper morphology.

Fermentation Process
Inoculation is generally carried out by transferring aseptically
the stock culture maintained on agar slants on other working
slants. After w24 h incubation at 30
C, the conidia crop is
inoculated in starch-rich seed-production media to yield up to
1011 spores cm3. This culture may be directly transferred into
1020 m3 seed fermenters to obtain a pellet-type inoculum
consisting of 15  105 pellets dm3 (0.10.2 mm in diameter), which in turn is used as inoculum (510% v/v) for the
industrial-scale production medium.
The fungus will develop different morphological forms
(Figure 2).
The formation of a loose mycelium with long, unbranched
hyphae is to be avoided because this results in an enormous
increase in the apparent viscosity of the culture broth, thus
limiting the effective oxygen transfer rate with little or no citric

Figure 1 Metabolic pathways for citric acid overproduction in Aspergillus niger. Only relevant enzyme activities, substrate, products, and effectors
(, inhibitor; , activator) are shown. Enzymes and transport systems: INV, membrane bound invertase; GC, low-afnity glucose carrier; FC: Fructose
carrier; PP, proton pump; CC, putative citrate carrier; HK, Hexokinase; PGI, phosphoglucose isomerase; PFK1, phosphofructokinase; PFK2,
6-phosphofructo-2-kinase; ALD, aldolase; PK, pyruvate kinase; PC, pyruvate carboxylase; MDH, malate dehydrogenase; PT, pyruvate transport system;
TCC, citrate transport system; PDH, pyruvate dehydrogenase; CS, citrate synthase; ACT, aconitase; IDH, isocitrate dehydrogenase; KDH, a-ketoglutarate
dehydrogenase; AOX, alternative oxidase system. Substrates and products: glu, glucose; glu6P, glucose-6-phosphate; fru6P, fructose-6-phosphate;
fru1,6 dP, fructose-1,6-bisphosphate; fru2,6 dP, fructose-2,6-bisphosphate; gly, glycerol; dhp, dihydroxiaceton phosphate; pep, phosphoenolpyruvate;
pyr, pyruvate; oaa, oxaloacetate; mal, malate; cit, citrate; acCoA, acetyl-coenzyme A; aco, cis-aconitate; ica, isocitrate; a-kg, a-ketoglutarate; sucCoA,
succinyl-coenzyme A; tre, trehalose.
The most important steps in controlling glycolytic ux are glucose transport (simple diffusion is the main mechanism at high sugar concentrations,
although A. niger has both low-afnity and high-afnity carriers for glucose) and hexokinase (HK) activity, which initially is inhibited by trehalose-6phosphate. PFK1 is feedback inhibited by citrate, but the inhibition is counteracted by the presence of high levels of NH
4 and by fructose-2,6-biphosphate
(FBP). A phosphorylated fragment of PFK1, which is insensitive to citrate inhibition, may be responsible for acidogenesis. PFK2 activity is increased at high
substrate concentration: its product, FBP lowers the MichaelisMenten constant (Km) of PFK1, counteracts citrate inhibition, and inhibits gluconeogenesis,
thus increasing carbon ux through glycolysis during acidogenesis. CS activity in A. niger is regulated by the level of OAA, which is produced in the
anaplerotic reaction catalyzed by PC. Malate is produced from OAA by cytosolic MDH and acts as a counterion for citrate efux from the mitochondrion,
where it is oxidized back to OAA. Low activity of NADP-specic IDH and KDH are a consequence of the effective removal of citrate, which has a higher
afnity than ACT.
A salicylhydroxamic acidsensitive, alternate oxidase system is used during acidogenesis to reoxidize the NADH produced during glycolysis.
Malfunction of the normal respiratory chain is due to diminished activity of NADH ubiquinone reductase and other respiratory chain enzymes.

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FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)

Figure 2 Pellet morphology of A. niger NRRL 2270 during citric production in a laboratory 2 dm3 stirred fermenter: (a) young pellet (100); (b) stubby,
bulbous hyphae with frequent branching, which are characteristic of citric acid production (400); and (c) degenerating pellet with pointed unbranched
hyphae protruding from the pellet core (100).

acid production. On the contrary, small spherical, dense

pellets (Figure 2(a)) with short stubby hyphae (Figure 2(b))
are generally regarded as the best morphological form
for optimal citrate yields. Frequent observation of pellet
morphology during this early stage of the fermentation using
a microscope allows hyphae proliferation to be controlled by
the addition, in case of adverse development (Figure 2(c)), of
appropriate amounts of inhibiting compounds, such as HCF
or zinc and copper sulfates. Mycelial clumps (whose structure
is less compact than pellets) also may develop under high
agitation speed.
Fermentation is exothermic and temperature has to be kept
in the range 2835
C. Assuming that the overall heat transfer
coefcient and effective temperature difference between the
fermenting medium and cooling water are of the order of
500 kJ m2 h1 and 5
C, respectively, the heat transfer surface
required to keep the fermentation temperature constant would
be w3.2 m2 per m3 of fermentation medium.
Low pH and high DO concentration are essential for citric
acid production. Initial decrease of pH is due to ammonium
uptake. Extreme pH values (<1.6) limit productivity, and the
addition of alkali (NH3) is used to control pH at 2.22.6 once
production of citric acid has started. The typical industrial-scale

productivities of 11.5 kg m3 h1 result in microbial oxygen

demand rates of 0.30.5 kg O2 m3 h1, that are met by
sparging 0.10.4 volumes of air per medium volume per
minute (vvm) at pressures at the sparger section of the
fermenter not less than 0.30.4 MPa and at the tank top,
ranging from 0.250.35 MPa to 0.120.15 MPa, depending on
the (stirred or air-lift) fermenter type used. Foaming is
controlled by adding food-grade antifoam agents.
Temporary interruption to the air supply during fermentation does not seem to affect the performance of the culture on
the condition that the DO level is greater than 20% of the
saturation value. DO values of about 0 for as long as 85 min,
followed by restoration of the air supply, do not inhibit
permanently mycelial growth and citrate production, but they
do reduce the product yield coefcient up to 20%.
Figure 3 shows the evolution of a typical batch citric acid
fermentation in glucose-based media by A. niger NRRL 2270 in
2-dm3 stirred fermenter and by a mutant strain of A. niger in
400 m3 bubble-column fermenter.
Two distinct phases are evident: during the primary growth
phase (trophophase), no acid production occurs; during the
second growth phase (idiophase), acid production by almost
nongrowing cells is observed.

FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)

809

A microscopic description of this fermentation using

A. niger pellets also has to account for oxygen diffusion
phenomena from the bulk of the fermenting medium to the
pellet surface and through the porous structure of the pellet
itself. The low effective diffusivity of oxygen within the pellet
limits mycelial activity to a peripherical spherical shell only,
with the oxygen penetration depth ranging from 110 to 300 mm
in 2 mm pellets.

Recovery and Purication Processes

Figure 3 Time course of a typical batch citric acid fermentation from

glucose-based media by A. niger NRRL 2270 in a 2 dm3 stirred fermenter
(closed symbols) and by an industrial strain in a 400 m3 bubble-column
fermenter (open symbols): Concentrations of mycelial biomass (X: A, >),
glucose (S: l, B), citric acid (P: n, ,), and ammoniac nitrogen (N: D)
versus time (t). The industrial-scale trial was gently provided by Dr A.
Truno c/o Palcitric SpA, Calitri, Italy, and the laboratory-scale trial was
performed by the authors at the University of Basilicata (Potenza, Italy).

Table 3 shows the simplied overall stoichiometric reactions occurring during the trophophase and idiophase of the
fermentation examined. In particular, it was assumed that
during the trophophase the microorganism (represented by
a raw formula based on elemental analysis: CHnOpNq) replicates itself at the expanses of a generic carbon source in the
presence of ammonia as the only nitrogen source; during the
idiophase, it undergoes further growth while decreasing
progressively its intracellular nitrogen content and excreting
citric acid in a medium practically devoid of any nitrogen
source (Figure 3).
Assuming that no carbon atom of sugar is converted into
biomass, carbon dioxide, or other by-products as shown by the
reaction in Eqn [2] (i.e., when y and d are equal to 0), the
theoretical molar yield (z) of citric acid would be one or two if
glucose or sucrose is used. This would be equivalent to 1.17 (or
1.23) kg of citric acid monohydrate per kilogram of glucose
(or sucrose) consumed. In practice, the industrial yields range
from 57 to 81% of this theoretical value, with the smaller
gure generally being associated with the surface fermentation
technique.
The citric acid fermentation may be mathematically
described by means of the set of kinetic equations shown
in Table 3.
In accordance with the HerbertPirt maintenance concept,
both product formation (rP) and substrate consumption (rS)
rates may be linearly related to cell growth rate (rx) and cell
concentration (X). In this way, the well-known Luedeking
Piret kinetics for product formation has to be regarded as
a special case: In fact, the rst term in Eqn [10] may be
described as the product formation rate in association with the
mycelial growth rate, whereas the second term may be regarded
as the nongrowth-associated product formation rate. In both of
the fermentation trials shown in Figure 3, citric acid fermentation may be classied to be of the mixed-growth-associated
product formation type.

Citric acid may be recovered from the broths resulting from

either the surface- or submerged-culture fermentations, using
almost the same three methods namely, direct crystallization
upon concentration of the ltered liquor, precipitation as
calcium citrate tetrahydrate, or liquid extraction (see Fermentation (Industrial): Recovery of Metabolites). Direct crystallization cannot be applied unless rened raw materials, such as
sucrose syrups or crystals, are used. Liquid extraction is used by
Tate & Lyle Co. (formerly Haarmann & Reimer Co., a subsidiary
of Bayer Co.) in the Dayton (OH, USA) and Elkhart (IN, USA)
plants. The precipitation process is used by the great majority of
world citric acid manufacturers, including ADM in the United
States.
A simplied process owsheet of this method is shown
in Figure 4.
Mycelia and suspended particles are separated by
continuous belt lters under vacuum. Citric acid is precipitated as calcium citrate by the addition of lime to the ltrate.
Liming temperature is critical. Amorphous tricalcium citrate
tetrahydrate generally is obtained at w70
C, while crystalline dicalcium acid citrate is obtained at 90
C. No removal
of oxalic acid is needed if the submerged-culture fermentation is used.
The residual citrate in the ltrate is precipitated as tricalcium
citrate by further addition of lime to set the pH to 5.8. The
crystals are recovered using another continuous belt lter and
then recycled back to the liming step, while the ltrate has to be
disposed.
Precipitation of dicalcium acid citrate results in one-third
less consumption of lime and consequently of sulfuric acid for
the subsequent regeneration of citric acid, in greater lterability
and washability because of its crystalline structure, but 1025%
of the expected product yield is needed as seed. The precipitate
is washed to remove the impurities adsorbed onto it (i.e.,
residual sugars and contaminants from the raw carbon source
and soluble proteins from the autolysis of the fungus). The
washed crystals and 98% w/w sulfuric acid are simultaneously,
but separately, fed to a mixer containing a 40% citric acid
solution at pH 0.50.6, to free the citric acid with the formation
of a precipitate of calcium sulfate dihydrate (gypsum). Final
rening of the ltrate is performed by decolorization on activated carbon and removal of residual calcium sulfate and iron
and nickel salts on strong cation exchange and weak anion
exchange (demineralization step). The resulting solution
(250280 kg m3 of citric acid anhydrous) is concentrated
using multiple-effect evaporators to about 700 kg m3, before
feeding a vacuum crystallizer operating at temperatures below
(35
C) or above (62
C) the transition temperature (36.6
C)
between the monohydrate and anhydrous forms, depending

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FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)
Table 3
Citric acid fermentation: overall stoichiometric reactions, kinetic equations, and instantaneous concentrations of
mycelia, product, sugar, and nitrogen sources
Equation or reaction
Trophophase reaction

C6 H12 O6 a NH3 b O2 / y CHn Op Nq d CO2 e H2 O

glucose

Idiophase reaction
Kinetic equations

C6 H12 O6 b O2 / y CHn Op Nq z C6 H8 O7 d CO2 e H2 O

glucose

rX

dX
mX X
dt

rN


dN
dX
YN=X
dt
dt

rN

dN
0
dt

3

6

for t  tlim

7

for t > tlim

rP

dP
0 for t  tlim
dt

rP


dP
dX
YP=X
mP X
dt
dt

8

9

for t > tlim

10


dS
dX
mS X
YS=X
dt
dt

X X0

11

for t  to

X X0 e mM
X

5

for N < Nlim


X
m mM 1 
XM

rS 

4

for N  Nlim

for t  to

m mM

t t0

12
for t  tlim

XM

XM
1
 1 e mM t tlim
X0

13
for t > tlim

14

for t < t0

N N0

2

citric acid

mycelium

m 0

Integral solutions of the

differential kinetic equations

1

mycelium

N N0  YN=X X  X0

15
for t  tlim

16

for t > tlim

17

P P0 mP At YP=X X  X0

18

S S0  mS At YS=X X  X0 

19

At 0

20

N Nlim

At

for t  tlim


XM
X h
ln 1  M 1  e mM
mM
mM

t tlim

i

for t > tlim

21

Nomenclature: A(t), cumulative nongrowth contribution to product formation; b, y, z, d, and e, stoichiometric coefcients; mP (mS), specic rate of
product formation (or substrate consumption) at zero cell growth rate; Nlim, critical concentration of nitrogen at the onset of citric acid production; P,
citrate concentration; ri, conversion rate of any reagent or product; tlim, overall duration of the citrate lag phase; to, overall duration of the cell lag phase;
S, substrate concentration; X, mycelium concentration; XM, maximum mycelium concentration; m, specic cell growth rate; mM, maximum specic cell
growth rate; YN/x, YP/x, and YS/x, yield factors for ammoniac nitrogen, citrate, and substrate on unit cell biomass. Subscripts: lim, referred to limiting
concentration of the nitrogen source; N, nitrogen; P, citric acid; S, glucose; X, mycelium; 0, referred to the initial value.

FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)
811

Figure 4 Process owsheet of a typical citric acid fermentation from glucose-based media by A. niger. Equipment and utility identication items: AE, anion exchanger; AF, antifoam agent; AL, alkaline reagent; BD,
uidized-bed drier; BF, vacuum belt lter; C, centrifuge; c, Condensate; CA, activated carbon adsorber; CE, cation exchanger; CR, vacuum crystallizer; cw, cooling water; CY, cyclone; D, holding tank; dcc, dicalcium
citrate; DW, demineralized water; E, heat exchanger; EA, exhausted air; EV, evaporator; F, production-bubble fermenter; FI, sterile pressure lter; GR, grinder; HT, holding tube; LS, lime slurry; HA, hot air;
HS, sulfuric acid; M, mixer; NA, nutrients and additives; PC, centrifugal pump; PE, plateheat exchanger; S, low-pressure steam; SA, sterile compressed air; Se, dicalcium citrate seed; SF, seed-bubble fermenter;
tcc, tricalcium citrate; WE, water evaporated.

812

FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)

on the form manufactured. Crystals are separated by centrifugation and dehydrated in a two-stage uidized-bed dryer, the
rst one using hot air at 90
C and the second one using air
conditioned at 20
C and relative humidity (RH) of 3040%
because of crystal hygroscopicity. The mother liquor is partly
(w20%) diluted with equipment-cleansing waters, decolorized, and fed back to liming; the remainder is in sequence
decolorized and demineralized before being recycled to the
crystallization unit. In this way, citric acid crystals do not need
additional purication steps to meet specications for U.S.
Pharmacopeia or Food Chemical Codex material.
In the liquid extraction process, citric acid may be extracted
from the fermentation broth using a highly selective, low-price,
and nontoxic food-grade solvent (i.e., water-insoluble amines,
namely trilaurylamine, n-octanol, C10 or C11 isoparafn, tri-nbutyl phosphate, alkysulphoxides). The extract is then heated
and washed countercurrently with water, resulting in about
90% recovery yield and an aqueous citric acid concentrated
solution, which is passed through a granular activatedcarbon
column before undergoing the aforementioned evaporation
and crystallization steps.

Future Developments
Production of citric acid has not been much in the focus of
modern molecular biology presumably because it is considered
a mature area. Any improvement of strains of A. niger usually
were carried out by mutagenesis and selection, but metabolic
and genetic engineering are likely to improve acidogenesis.
The replacement of the current batch fermentation with
semicontinuous processes, to increase volumetric productivity
and reduce specic production costs, presently is hampered in
fungal processes by the deterioration of mycelial structure, the
mechanism of which still is unknown. Although the effect of
nitrogen deciency on citric acid accumulation by A. niger is
well known, a low level of ammonium ions (i.e., 30 g m3,
equivalent to 2 mmol of intracellular NH
4 per gram of dry cell)
was found to inhibit the morphological degeneration of pellets
and postpone sporulation. NH
4 ions simply are not deposited
into the cell to form the so-called ammonium pool, but they
enter the cell to combine with glucose and form glucosamine,
that is straight away released in the medium. The effective
relationship between the different compounds of the TCA cycle
is to be studied further and controlled before the present batchproduction technology may be converted effectively into
a prolonged fed-batch or continuous production process.
Similarly, the possibility of maintaining microbial cells
active and controlling their growth and production processes
for several weeks or months by immobilization within organic
or inorganic matrices represents a further challenge to the
technological modernization of this sector.
The traditional recovery technology results in several
problems because of disposal of liquid efuents (their chemical oxygen demand being about 20 kg m3) and solid
by-products (i.e., about 0.15 kg of dried mycelium and 2 kg of
gypsum per kilogram of citric acid anhydrous). Several process
alternatives have been suggested thus far to minimize the
overall environmental impact of this process. The replacement
of molasses with raw or hydrolyzed starch- or raw sucrosebased materials would simplify only the downstream

processing. On the contrary, the recovery of tricalcium (or

trisodium) citrate from claried, decolorized fermentation
broths by electrodialysis, as well the adsorption of citric acid
onto weakly basic anionic-exchange resins or zeolites using the
simulated-moving bed chromatographic technology (Citrex
Process, UOP, Des Plaines, IL, USA) followed by desorption
with water or dilute acidic solutions, or the use of liquid
membranes, would allow the citric acid to be separated in
a single step and to be recovered without the formation of solid
wastes for disposal.
The environmental aspects of citric acid production have
been assessed. Despite the fact that most raw materials are of
biological origin, many ingredients, such as ammonium
nitrate, lime, and sulfuric acid, are hazardous chemicals. For
instance, it was found that the environmental impact of citric
acid production using whey was smaller than that using corn
starch.

Gluconic Acid
D-Gluconic acid (2,3,4,5,6-pentahydroxy pentane-1-carboxylic
acid: C6H12O7) is an oxidation product of D-glucose, which, in
an aqueous solution, leads to a complex equilibrium between
gluconic acid and its two lactones: 1,5-lactone (D-gluconod-lactone) and 1,4-lactone (D-glucono-g-lactone).
D-Gluconic acid is commercially available as 50% aqueous
solution (density of 1230 kg m3 at 20
C and pH 1.82). This
acid and its derivatives are used in the pharmaceutical, food,
feed, and chemical industry because of their low toxicity and
their ability to form water-soluble complexes with metallic
ions (e.g., Ca2, Fe3), especially in the presence of 510% of
sodium hydroxide. Sodium gluconate is the main industrial
product and it is used as a sequestering agent (e.g., bottle
washing, metal surface cleaning, and rust removal) and to
plasticize and retard the curing process of cement mixes. The
calcium and iron gluconates are used in medicine to treat
diseases of calcium and iron deciency (such as osteoporosis
and anemia). D-Glucono-d-lactone is used as a latent acid in
baking powders for use in dry cake mixes, meat processing, and
instant chemically leavened bread mixes, whereas D-gluconog-lactone is made only in small quantities as a specialty
chemical.
The conversion of glucose to gluconic acid is a simple
oxidation process and may be carried out by a variety of
processes namely, microbial fermentation, chemical, electrochemical, or enzymatic catalysis. Currently, these processes
appear to be either more expensive, unstable, or less efcient
than the fermentation process, which presently is the only
method of choice.
After the rst isolation of calcium gluconate (1880) from
glucose fermentation in the presence of CaCO3 by a strain of
Mycoderma aceti, the Chas. Pzer & Co., Inc. (New York, USA)
started industrial-scale production of gluconic acid in 1923.
Further research at the U.S. Department of Agriculture in
cooperation with the Iowa State College led to the semicontinuous production of sodium gluconate from glucose
using A. niger NRRL 67.
Several lamentous fungi of the genera Penicillium and
Aspergillus, the yeastlike fungus Aureobasidium pullulans, and

FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)
bacteria (Acetobacter suboxidans, Pseudomonas ovalis, Gluconobacter spp.) produce gluconic acid from glucose-based media,
but industrial processes have been developed only for the
production gluconic acid from glucose syrups with A. niger and
Gluconobacter oxydans. Industrial strains are not freely available,
but a few gluconic acidproducing strains (A. niger NRRL 3,
NRRL 67) can be obtained from international culture collections. Penicillium spp. generally produce less gluconate than
Aspergillus, but they have the advantage of excreting the glucose
oxidase (an important by-product) into the medium, which
makes its recovery easier.
The formation of gluconic acid by A. niger is controlled by
the enzyme glucose oxidase, an omodimer containing two
avin adenine dinucleotide (FAD) moieties. Such enzyme
abstracts two hydrogen atoms from glucose, thus yielding the
glucono-d-lactone, which to some extent hydrolyzes to gluconic acid. The FADH2 reacts with oxygen to form hydrogen
peroxide, which is converted into oxygen and water by the
enzyme catalase. Both glucose oxidase and catalase are
constitutive endoenzymes in A. niger.
A highly productive process of gluconic acid using freegrowing cells of A. pullulans DSM 7085 recently has been
developed. Its high conversion yields (9098%) and rates
(1319 kg m3 h1) resulted in as high gluconate concentrations
as 504 or 230433 kg m3 in fed-batch or chemostat trials.
Although this novel fermentation process offers a new opportunity for commercial gluconic acid production, as well as many
advantages over the traditional microbial fermentation processes, it is still conned to laboratory-scale applications. Thus,
only the sodium process by batch-submerged fermentation from
glucose syrups using A. niger will be described in the following
paragraphs.
Glucose syrups of 70
Brix strength are generally used as
carbon source in the preparation of the fermentation medium.
Table 4 lists the typical composition of the seed and
production media used in laboratory- and industrial-scale
trials.
After the pH is adjusted at 4.5 with sulfuric acid, the
medium is sterilized at 121
C for 1530 min, cooled at 33
C,
and then transferred into the fermentation vessel. The pH is

Table 4
Composition for the production media used in the
laboratory- and industrial-scale production of gluconic acid by A. niger

Component
Glucose
NH4NO3 (or other
NH
4 salt)
KH2PO4
MgSO4$7H2O
Agar
Yeast extract
Corn-steep liquor
ZnSO4$7H2O
CuSO4$5H2O
FeCl3$6H2O
MnSO4$7H2O
Initial pH

Vegetative seed
culture media

Gluconic acid
production media

Unit

40
2.4

120350
0.40.5

kg m3
kg m3

1.5
3.57
1
1
0
100
20
300
0
6.5

0.10.3
0.10.3
0
0
0.20.4
0
0
0
30
6.0

kg m3
kg m3
kg m3
kg m3
kg m3
mg m3
mg m3
mg m3
mg m3

813

adjusted to 66.5 with sodium hydroxide, and a 25% v/v

inoculum generally is used. For inoculum development, conidia are recovered from stock agar slants and are inoculated into
vegetative seedculture media (106 conidia cm3); pellet-like
mycelia is obtained after incubation at 30
C for 1524 h and is
used to inoculate seed fermenters at a density of 2050
pellets cm3.
The fermentation is carried out under continuous automatic
control of sterile air sparging (1.01.5 vvm), temperature
(33
C), pressure on the tank top (23 bar), pH (5.56.5 by
addition of 3050% NaOH solution to neutralize the gluconic
acid formed), and foam level. It is completed within w30 h
with yield factors of 0.971 kg of gluconic acid per kilogram of
glucose consumed (against a theoretical yield of 1.09 kg kg1)
and gluconate productivities of 913 kg m3 h1.
In the fed-batch operation, the mycelium may be reused up
to ve times without any loss in gluconate productivity
provided that the levels of glucose oxidase activity and other
microelements (i.e., iron and manganese) are kept under
control. Stepwise addition of glucose may be used to increase
gluconate concentration to 580 kg m3.
At the end of fermentation, the mycelium is removed using
aseptic centrifugation, under vacuum-belt ltration or crossow microltration and may be used as a source of glucose
oxidase or may be disposed off via incineration. The claried
broth, generally containing w300 kg m3 of sodium gluconate,
is ltered, decolorized using a granular activatedcarbon
column, concentrated under vacuum to 4550% total solids,
neutralized to pH 7.5 with NaOH, and then spray or drum
dried. If 50% gluconic acid is required, the concentrated liquor
may be passed through a cation exchanger to remove Na ions.
Further crystallization at 3070
C or at more than 70
C
allows crystals of the d-lactone or g-lactone to be precipitated,
respectively.

Lactic Acid
Lactic acid (2-hydroxypropionic acid: C3H6O3) may be
produced by chemical synthesis or fermentation. Of the two
enantiomers, L-() and D-() lactic acid, only the L-() isomer
is used by human metabolism and, because of the slight
toxicity of the D-() isomer, it is preferred for food uses.
Because the chemical route yields a mixture of L-lactic acid and
D-lactic acid and relies on costly raw materials, all lactic acid
manufacturing industries have switched to fermentation-based
technologies (Table 1). The free acid is used as an acidulant/
preservative in several food products (cheese, meat, jellies,
beer) (see Preservatives: Traditional Preservatives Organic
Acids); sodium lactate is used for carcass decontamination;
ammonium lactate is used as a source of nonprotein nitrogen
in feeds; sodium and calcium stearoyl lactylates are used as
emulsiers and dough conditioners. The large increase in lactic
acid production is due to its use in the synthesis of polylactic
acid (PLA), a polyester used for biodegradable plastics for food
packaging, compost, and garbage bags, and disposable tableware, as well as several medical applications, such as reabsorbable sutures, orthopedic implants, and controlled drug
release. The current economically viable industrial process for
PLA production is via the dehydrated cyclic dilactate ester

814

FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)

(lactide) formation. In brief, lactic acid is rst polymerized into

oligomers of PLA consisting of 3070 lactyl units (CHCH3
COO). These oligomers are then depolymerized by
increasing the polycondensation temperature and lowering the
pressure in the presence of transition metalbased catalysts
(i.e., stannous octoate at 0.05%) to distil the lactide. Finally, by
opening its ring, it is possible to obtain high molar mass
polymers (100300 kDa) with appropriate optical and
mechanical properties (i.e., tensile strength >50 MPa). Other
comonomers, such as caprolactone, hydroxybenzoic acid, and
others, can be incorporated to provide environmentally safe
materials. PLA appears to be a sustainable alternative to
petroleum-based plastics, because lactide is produced from the
fermentation of renewable resources, such as corn starch (as in
the industrial plant of Nature Works LLC, Blair, Nebraska, USA:
140 000 metric tons per year). Nevertheless, to minimize
competition for land and food, research studies have started to
develop second-generation PLA products from lignocellulosic
hydrolysates (e.g., crushed corncobs).

Organisms and Metabolic Pathways Involved

Lactic acid can be produced using homofermentative lactic
acid bacteria (LAB), facultatively anaerobic Bacillus species
(B. coagulans), and molds (Rhizopus microsporus, Rhizopus
oryzae). Recently, lactic acid producing genetically modied
strains of Escherichia coli and Saccharomyces cerevisiae have
been developed. The choice of the species depends on several
considerations, including the ability to use the type of sugars
available in the substrate, growth temperature, nutritional
needs, acid tolerance, and type of lactic acid isomer
produced.
Thermophilic lactobacilli (Lactobacillus delbrueckii subsp.
delbrueckii, L. delbrueckii subsp. bulgaricus, Lactobacillus helveticus)
tolerate higher concentrations of lactate and higher temperatures
(4852
C), thus involving higher productivity and yields
and reduced contamination risks. They produce D-() or DL-lactic
acid (some industrial strains that have been claimed to be
L. delbrueckii produce L-() lactic acid) and may be less suitable
for food, feed, or biomedical applications. Lactococci, mesophilic
lactobacilli (Lactobacillus casei subsp. casei, Lactobacillus amylophilus), and thermophilic streptococci (Streptococcus thermophilus)
have lower temperature optima or reduced acid tolerances, but
they may be desirable for other reasons (e.g., production of pure
L-() lactic acid, hydrolysis of starch). Recently, genetic engineering has been used to produce L. helveticus and Lactobacillus
plantarum strains, which produce optically pure L-() or D-()
lactic acid (see Lactobacillus: Introduction; Lactococcus: Introduction; and Streptococcus thermophilus).
Homofermentative LAB ferment hexoses via the glycolytic
pathway (see Metabolic Pathways: Release of Energy (Anaerobic)). Pyruvate is reduced to lactate by stereospecic lactate
dehydrogenase(s) (L-LDH or D-LDH). LDH is allosteric (activators: fructose-1,6-bisphosphate and Mn2) in lactococci and
nonallosteric in homofermentative lactobacilli. Undissociated
lactic acid acts as a noncompetitive inhibitor for growth and
lactic acid production by diffusing through the membrane and
decreasing intracellular pH: pH control during fermentation
reduces the inhibition, but the maximum lactic acid concentration achievable is usually lower than 150 kg m3.

Concomitant substrate and product inhibition has been

reported for several species.
LAB are fastidious microorganisms and require supplementation of fermentation media with peptides and growth
factors, usually in the form of yeast extract. Because this may
account for 3035% of substrate costs, they may be replaced by
less demanding B. coagulans and R. oryzae, both being
L()-lactate producers, even if smaller yields (as low as 70%
for R. oryzae because of concomitant fumaric acid and ethanol
production) and acid tolerance may offset the advantage of
using lower amounts of supplements.

Substrate Production and Recovery

Lactic acid can be produced from a variety of raw substrates
(whey and whey permeate, beet and cane molasses, starch and
corn starch hydrolysates, wood hydrolysates; see Fermentation
(Industrial): Media for Industrial Fermentations). Some species
(Lb. amylophilus) can hydrolyze starch, but the pseudoplastic
behavior of starchy substrates makes the pH control difcult.
When whey permeate is used, supplementation with milk
protein hydrolysates (510 kg m3) and yeast extract (up to
20 kg m3) is required. Lactic acid production is usually
a growth-associated production process, but nongrowthassociated production becomes signicant when growth is
limited by a lack of nutrients or high undissociated acid
concentration. The pH is controlled at 56.5 by the automatic
addition of NaOH, Na2CO3, or NH4OH or by the addition of
CaCO3. Fermentation is carried out under anaerobic or microaerophilic conditions and lactic acid yield is usually between 85
and 98% with isomer purity as high as 99%. Batch fermentations result in high product concentration (120150 kg m3)
but in low productivity (2 kg m3 h1). Conversely, continuous
fermentations with cell-recycle or immobilized cells give rise to
higher productivities (2080 kg m3 h1) and lower lactate
concentrations (<50 kg m3). End-product inhibition may be
circumvented by using integrated fermentation processes, in
which lactic acid is removed from the culture broth by several
techniques, including electrodialysis, ion-exchange resins, or
nanoltration.
Recovery of lactate is made complicated by the high solubility of its salts. The traditional process involves precipitation
of calcium lactate and regeneration of lactic acid by the addition of sulfuric acid followed by further purication steps (ion
exchange and decolorization). Alternative processes include the
extraction by liquid membranes, electrodialysis, and ion
exchange. In particular, the recent industrial use of electrodialysis with bipolar membranes in France resulted in the virtual
elimination of gypsum waste production. Conventional
recovery by the precipitation method seems to be the most
economical route.

Other Organic Acids Produced by Fermentation

Propionic Acid
Propionic acid (C3H6O2) and its salts are used as mold inhibitors in bakery products, although other nonfood uses are
important (see Permitted Preservatives Propionic Acid). It may
be produced by fermentation by members of the genera

FERMENTATION (INDUSTRIAL) j Production of Some Organic Acids (Citric, Gluconic, Lactic, and Propionic)
Propionibacterium (P. freudenrichii, thoenii, acidipropionici), Veillonella, Clostridium, and Selenomonas, but currently it is produced
by chemical synthesis because of the shortcomings of the
fermentation route (low productivities, <1 kg m3 h1 in batch
processes; low product concentrations, <50 kg m3; difculty in
product separation from acetic acid, which invariably is
produced from sugars and lactate). The high microbial
productivities (214 kg m3 h1) obtained in continuous
fermentations using immobilized cells or membrane-recycle
reactors, as well the possibility of obtaining pure propionic acid
from alternative low-cost substrates, like crude glycerol from the
biodiesel industry, might refocus industrial manufacturers
toward the fermentation route. For instance, use of a metabolically engineered strain of P. acidipropionici (ACK-Tet) resulted in
a propionic acid concentration of 106 kg m3 with a product
yield of 0.540.71 g per g of glycerol consumed and a propionic
acid-to-acetic acid ratio of 22.4.

See also: Arthrobacter; Aspergillus; Bacillus: Introduction; Bread:

Bread from Wheat Flour; Yarrowia lipolytica (Candida Lipolytica);
Escherichia coli: Escherichia coli; Fermentation (Industrial):
Basic Considerations; Fermentation (Industrial): Media for
Industrial Fermentations; Fermentation (Industrial): Control of
Fermentation Conditions; Fermentation (Industrial): Recovery of
Metabolites; Fermented Foods: Fermentations of East and
Southeast Asia; Fungi: The Fungal Hypha; Fungi: Classication
of the Hemiascomycetes; Fungi: Classication of the
Deuteromycetes; Genetic Engineering; Gluconobacter;
Lactobacillus: Introduction; Metabolic Pathways: Release of
Energy (Aerobic); Metabolic Pathways: Release of Energy
(Anaerobic); Preservatives: Traditional Preservatives Organic
Acids; Permitted Preservatives Propionic Acid;
Propionibacterium; Streptococcus thermophilus; Vinegar; Yeasts:
Production and Commercial Uses.

815

Further Reading
Anastassiadis, S., Aivasidis, A., Wandrey, C., 2003. Continuous gluconic acid
production by isolated yeast-like mould strains of Aureobasidium pullulans. Applied
Microbiology and Biotechnology 61, 110117.
Berovic, M., Legisa, M., 2007. Citric acid production. Biotechnology Annual Review 13,
303343.
Hofvendahl, K., HahnHgerda, B., 2000. Factors affecting the fermentative lactic acid
production from renewable resources. Enzyme and Microbial Technology 26,
87107.
Joglekar, H.G., Rahman, I., Babu, S., Kulkarni, B.D., Joshi, A., 2006. Comparative
assessment of downstream processing options for lactic acid. Separation and
Purication Technology 52, 117.
John, R.P., Nampoothiri, K.M., Pandey, A., 2007. Fermentative production of lactic
acid from biomass: an overview on process developments and future perspectives.
Applied and Microbiology and Biotechnology 74, 524534.
Legisa, M., Mattey, M., 2007. Changes in primary metabolism leading to citric acid
overow in Aspergillus niger. Biotechnology Letters 29, 181190.
Magnuson, J.K., Lasure, L.L., 2004. Organic acid production by lamentous fungi.
Chapter 12. In: Tkacz, J.S., Lange, L. (Eds.), Advances in Fungal Biotechnology for
Industry, Agriculture, and Medicine. Kluwer Academic/Plenum Publishers, New
York, pp. 307340.
Okano, K., Tanaka, T., Ogino, C., Fukuda, H., Kondo, A., 2010. Biotechnological production
of enantiomeric pure lactic acid from renewable resources: recent achievements,
perspectives, and limits. Applied Microbiology and Biotechnology 85, 413423.
Papagianni, M., 2007. Advances in citric acid fermentation by Aspergillus niger: biochemical
aspects, membrane transport and modeling. Biotechnology Advances 25, 244263.
Sauer, M., Porro, D., Mattanovich, D., Branduardi, P., 2007. Microbial production of
organic acids: expanding the markets. Trends in Biotechnology 26 (2), 100108.
Singh, O.V., Kumar, R., 2007. Biotechnological production of gluconic acid: future
implications. Applied Microbiology and Biotechnology 75, 713722.
Yoo, D.-K., Kim, D., 2009. Production of optically pure poly(lactic acid) from lactic acid.
Polymer Bulletin 63, 637651.
Zhang, A., Yang, S.-T., 2009. Propionic acid production from glycerol by metabolically engineered Propionibacterium acidipropionici. Process Biochemistry 44,
13461351.