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Appl Microbiol Biotechnol (1999) 51: 545552

Springer-Verlag 1999


J. G. Zeikus M. K. Jain P. Elankovan

Biotechnology of succinic acid production and markets

for derived industrial products

Received: 27 October 1998 / Received revision: 22 January 1999 / Accepted: 22 January 1999

Abstract Succinic acid, derived from fermentation of

agricultural carbohydrates, has a specialty chemical
market in industries producing food and pharmaceutical
products, surfactants and detergents, green solvents and
biodegradable plastics, and ingredients to stimulate animal and plant growth. As a carbon-intermediate
chemical, fermentation-derived succinate has the potential to supply over 2.7 108 kg industrial products/
year including: 1,4-butanediol, tetrahydrofuran, c-butyrolactone, adipic acid, n-methylpyrrolidone and linear
aliphatic esters. Succinate yields as high as 110 g/l have
been achieved from glucose by the newly discovered
rumen organism Actinobacillus succinogenes. Succinate
fermentation is a novel process because the greenhouse
gas CO2 is xed into succinate during glucose fermentation. New developments in end-product recovery
technology, including water-splitting electrodialysis and
liquid/liquid extraction have lowered the cost of succinic
acid production to U.S. $ 0.55/kg at the 75 000 tonne/
year level and to $ 2.20/kg at the 5000 tonne/year level.
Research directions aimed at further improving the
succinate fermentation economics are discussed.

Succinic acid is a common metabolite formed by plants,
animals and microogranisms. Many anaerobic microbes
produce succinate as the major end-product of their

J. G. Zeikus (&) M. K. Jain P. Elankovan

MBI International, 3900 Collins Road, Lansing, MI 48910, USA
Tel.: +1-517-337-3181
Fax: +1-517-337-2122
J. G. Zeikus
Michigan State University, Departments of Biochemistry and
Microbiology, 410 Biochemistry Building, East Lansing, MI, USA
Tel.: +1-517-353-4674
Fax: +1-517-353-9334

energy metabolism. Nonetheless, only recently has interest focused on the development of succinic acid as an
important industrial fermentation process. This review
will explain why succinic acid fermentations may, in the
future, involve larger production volumes than citric
acid, and could, perhaps, approach that of ethanol.
Green technology is becoming more of a driving force
in the chemical industry because of the current need to
decrease pollution caused by petrochemical processing
and the future need to replace the dwindling hydrocarbon economy with a renewable, environmentally sound,
carbohydrate economy.
Succinic acid as a fermentation product and process
has distinct advantages over ethanol fermentation. For
example, 2 mol waste CO2 is formed/mol glucose during
ethanol fermentation, whereas the succinate fermentation consumes CO2. Being a CO2-xing fermentation
makes succinate a very green technology. Furthermore,
integrating the succinate and ethanol fermentations with
recovery would decrease carbon lost as waste CO2 and
produce three commercial products: succinic acid, ethanol and diethyl succinate.
The industrial potential for succinic acid fermentations was recognized as early as 1980 (Zeikus 1980).
Recently, interest in higher-value organic acid fermentations, including succinic acid, has intensied especially
in the question of how it relates to utilization of agricultural carbohydrates such as corn, cassava and sugar
cane (Jain et al. 1989; Zeikus et al. 1995). The purpose
of the present review is to provide an updated view on
the potential for succinate acid fermentation and the
current achievements and limitations of the process.

Uses and markets for succinic acid

Currently, more than 15 000 tonnes of industrial succinic acid is sold, and it is, by and large, produced petrochemically from butane through maleic anhydride. It
is sold at a spot price of about U.S. $ 5.908.80/kg depending on its purity. At present, only natural succinic


acid sold in the food market is produced by fermentation. New fermentation technology developed at MBI
International, however, can produce succinic acid for
sale at about $ 2.20/kg at the level of 5000 tonnes/year
and the price drops to below $ 0.55/kg at above 75 000
tonnes/year. These economics promise to open new
specialty and commodity chemical markets for succinic
There are four major existing markets for succinic
acid. The rst and largest is as a surfactant/detergent
extender/foaming agent. The second market is as an ion
chelator, where it is used in electroplating to prevent
corrosion and pitting of metals. The third is the food
market, where it is used as an acidulant/pH modier, as
a avoring agent, and as an anti-microbial agent. The
fourth market is the production of health-related agents,
including pharmaceuticals, antibiotics, amino acids, and
vitamins. The total market size for these four existing
uses of succinic acid is more than $ 400 000 000/year.
Fermentation-derived succinic acid has the potential
to become a large-volume commodity chemical that
would form the basis for supplying many important
intermediate and specialty chemicals for the consumer
products industries. As a commodity chemical, succinic
acid could replace many commodities based on benzene
and intermediate petrochemicals, resulting in a large
reduction in pollution from the manufacturer and in the
consumption of over 250 benzene-derived chemicals
(Ahmed and Morris 1994). Figure 1 illustrates a potential map of the routes leading to succinic-acid-based
intermediate and specialty chemicals. The production of
succinic acid and most of its derivatives is currently at

Fig. 1 Map of routes to succinic acid-based products

the advanced research and development stage. Because

of its structure as a linear saturated dicarboxylic acid,
succinic acid can be used as an intermediate chemical
and be converted to 1,4-butanediol, tetrahydrofuran,
c-butyrolactone, and other four-carbon chemicals that
have a world-wide market in excess of 275 106 kg/
year. Major advances have been made in the chemical
catalysis technology needed to convert succinic acid into
these intermediate chemicals. For example, succinic acid
can be readily hydrogenated to 1,4-butanediol, which
can then be further carbonylated to adipic acid (Sado
and Tajima 1980; Dake et al. 1987; Jain et al. 1989). The
following are only a few important succinic-acid-based
intermediate chemicals that are currently in the research
and development or pilot demonstration stage:
1. N-Methylpyrrolidone. N-Methyl pyrrolidone has been
recommended as a replacement for methylene chloride
as a solvent. It is much less volatile than methylene
chloride so it can be captured and recycled without release of toxic emissions to the atmosphere. The market
for methylene chloride is in the order of millions of
tonnes per year.
2. 1,4-Butanediol. 1,4-Butanediol is an important industrial solvent and raw material for polybutylene
terephthalate resins and automotive and electrical parts.
The markets for engineered, ``stronger-than-steel,''
plastics has grown rapidly.
3. c-Butyrolactone. This is a chemical intermediate and
ingredient of paint removers and textile products. It is
the raw material for pyrrolidone derivatives.

Commodity Chemicals
Adipic Acid

Specialty Chemicals



4,4 Esters

Plant Growth


Food Ingredients

Feed Additives


Green Solvents


Butanoic Acid

Detergents and

Health Agents
Maleic Anhydride


Chelators and
Corrosion Inhibitors

Maleic Acid

Malic Acid

Fumaric Acid

Itaconic Acid
Aspartic Acid


4. Adipic acid. The precursor to nylon 6,6, adipic acid is

a raw-material chemical used in the manufacture of lubricants, foams and food products.
5. Tetrahydrofuran. Tetrahydrofuran is a solvent and
key ingredient of adhesives, printing inks, and magnetic
6. Linear aliphatic esters. These are important compounds used in making resins, plastics, and other industrial and consumer end-products.
Succinic salts may also be used as additives to animal
feed for ruminants and monogastrics animals such as
pigs. Succinate has been shown to increase propionate
production in the rumen and act as a glycogenic material
and a precursor for protein synthesis. Furthermore,
succinate salts can act as buers and energy sources for
young pigs and other monogastric animals. Hence, the
crude succinate salt produced from carbohydrates by
fermentation could nd new major markets as a specialty, functional product for animal nutrition to reduce
the usage of antibiotics (such as monensin and lasalocid
in certain animal feeds (Bergen and Bates 1984). Succinic
is also a growth promoter for stressed plants (Kinnersley
et al. 1997).
New uses for succinic acid in specialty chemicals and
materials are rapidly growing. Sodium succinate is a
avor enhancer that can replace monosodium glutamate, and dilysine succinate is a salty avor enhancer for
low-sodium foods (Turk 1993). Succinic acid is also an
intermediate for green chemicals and materials. For example, diethyl succinate is a useful solvent for cleaning
metal surfaces or for paint stripping, and ethylenediaminedisuccinate is a replacement for EDTA (Zwicker
et al. 1997). The new biodegradable plastic Bionelle is an
ester comprised of succinic acid and 1,4-butanediol.
Other potential uses of succinic acid include (a)
succinylation of lysine residues to improve the physical
and functional attributes of soy proteins in foods, (b)
producing modied succinimides used as fuel constituents (Wollenberg and Frank 1988) and in water purication, (c) in the succinylation of cellulose to improve
water absorbitivity and (d) the use of succinylated starch
as a thickening agent (Diamantoglon and Meyer 1988).

Organisms and diversity

Succinic acid is a common intermediate in the metabolic
pathway of several anaerobic and facultative microorganisms. For example, succinate is formed from sugars
or amino acids by propionate-producing bacteria such
as Propionibacterium species, typical gastrointestinal
bacteria such as Escherichia coli, Pectinatus sp., Bacteroides sp., rumen bacteria such as Ruminococcus
avefaciens, Actinobacillus succinogenes, Bacteroides
amylophilus, Prevotella ruminicola, Succcinimonas amylolytica, Succinivibrio dextrinisolvens, Wolinella succinogenes, and Cytophaga succinicans (Bryant and Small

1956; Bryant et al. 1958; Scheinger and Wolin 1973;

Davis et al. 1976; Van der Werf et al. 1997; Guettler
et al. 1999). A number of Lactobacillus strains have also
been reported to produce succinic acid in de ManRogosa-Sharpe broth to various extents (Kaneuchi et al.
Most of the succinate-producing microorganisms
have been isolated from the rumen because, in this
ecosystem, succinate serves as an important precursor
for propionate, which is absorbed through the rumen
wall for subsequent oxidation to provide energy and
biosynthetic precursors for the animal. Animals receive
both forage (such as hay) and grains (such as corn) as
part of their diet, which allows for a great diversity of
rumen microorganisms. Ruminococcus avefaciens and
Fibrobacter succinogenes are major cellulose-digesting
anaerobes and major producers of acetic and succinic
acids in the rumen (Weimer 1993). Succinivibrio dextrinisolvens thrives in the rumen when the diet of the
animal is high in starch and it forms succinate, acetate,
formate, and lactate as end-products of glucose fermentation (O'Herrin and Kenealy 1993). Actinobacillus
succinogenes 130Z is a ruminal, osmotolerant, facultative anaerobic bacterium that has the ability to utilize a
wide range of substrates including L-arabinose, cellobiose, fructose, galactose, glucose, lactose, maltose, mannitol, mannose, sucrose, D-xylose, and salicin (Guettler
et al. 1999). This rumen bacterium produces succinate in
very high concentrations, along with acetate, pyruvate,
formate, or ethanol. Succinate is also produced by
microorganisms isolated from the digestive system of
other animals. For example, Anaerobiospirillum succiniciproducens isolated from the mouth of the beagle dog is
a gram-negative obligately anaerobic bacterium that
produces succinate, acetate, formate, ethanol, and lactate, from glucose and lactose (Davis et al. 1976).

Physiology, biochemistry and genetics

All succinic-acid-producing bacteria form mixed-acid
fermentations, producing varying amounts of succinate
as well as other products, including, ethanol, lactic,
acetic, and formic acid. Escherichia coli, for example, produces succinate as a minor fermentation product, typically 12 mol/100 mol glucose (Wood 1961). On
the other hand, A. succiniciproducens forms succinate up
to 120 mol/100 mol glucose (Samuelov et al. 1991;
Nghiem et al. 1997).
The physiology of succinate-producing bacteria varies signicantly. Table 1 illustrates the major physiological dierences between those strains that display
very high succinate yields during fermentation of glucose
in complex media. A. succiniciproducens diers from
E. coli and A. succinogenes strains in being an obligate
anaerobe. A. succiniciproducens and A. succinogenes use
the phosphoenolpyruvate (PEP) carboxykinase pathway
exclusively, whereas E. coli utilizes multiple pathways to
form succinic acid (Van der Werf et al. 1997). Most

Table 1 Physiological dierences between high-yielding succinate-producing bacterial strains. PEP phosphoenolpyruvate

O2 relation

Succinate pathway

Escherichia coli mutants

succinogenes mutants

Obligate anaerobe

PEP carboxykinase



PEP carboxykinase


importantly, A. succinogenes, unlike E. coli or A. succiniciproducens, is a moderate osmophile and has high
tolerance to succinate salts, which is crucial to process
requirements for product recovery (see next section).
A. succinogenes variant strains can yield 110 g/l succinic
acid (Guettler et al. 1996). It should be pointed out that
industrial strains of Corynebacterium can make monosodium glutamate at 150 g/l. Thus 15% succinate is a
potential target product yield for future genetic strain
A. succinogenes forms succinate via the PEP carboxykinase pathway (see Fig. 2), using four key enzymes, i.e., PEP carboxykinase, malate dehydrogenase,
fumarase, and fumarate dehydrogenase, whereas, in
E. coli, six pathways have been recognized in the formation of succinic acid, and the PEP carboxykinase
pathway plays a minor role (Clark 1989; Van der Werf
Fig. 2 Proposed catabolic
pathway for glucose fermentation in A. succiniciproducens
and A. succinogenes. Steps: 1
phosphoenolpyruvate carboxykinase, 2 malate dehydrogenase, 3 fumarate reductase, 4
pyruvate kinase, 5 pyruvate
ferredoxin oxidoreductase, 6
acetate kinase, 7 alcohol dehydrogenase, 8 lactate dehydrogenase (Samuelov et al. 1991)

Succinate yield (g/l)

Tolerance to succinate salts:

Na 96 g/l: Mg 130 g/l

et al. 1997). Recently E. coli (Millard et al. 1996) was

found to increase succinate fermentation by overproducing PEP carboxylase, but not PEP carboxykinase.
The existence of dierent enzymes indicates dierent
pathways for succinate formation in E. coli and the
studies of Alam and Clark (1989) also conrm multiple
routes for succinate formation. They found that a
fumarate-reductase-negative mutant of E. coli continued
to produce succinate at concentrations of 15%40% as
compared to the concentration of the wild-type.
The PEP carboxykinase pathway used for succinic
acid production by A. succiniciproducens and A. succinogenes is regulated by CO2 levels (Samuelov
et al. 1991; Van der Werf et al. 1997). In these bacteria,
PEP carboxylase (PEP carboxykinase) functions catabolically to x CO2 and synthesize oxaloacetate from
PEP. At low CO2 levels (10 mol CO2/100 mol glucose),


ADP Phospho enol pyruvate



High CO2





CO2 (5)
Acetyl CoA




Acetyl phosphate


Low CO2


A. succiniciproducens produces lactic acid as a major

reduced end-product, whereas A. succinogenes produces
ethanol as a major-end product. Under high CO2 levels
(100 mol CO2/100 mol glucose), succinate is the major
reduced product and only traces of lactic acid or ethanol
are produced by these organisms.
The CO2 concentration has been shown to regulate
the levels of key enzymes of the PEP carboxykinase
pathway in A. succinogenes (see Fig. 2) at high CO2
levels (Samuelov et al. 1991). PEP carboxykinase levels
rise, whereas alcohol dehydrogenase and lactate dehydrogenase are not detectable. Consequently, CO2 functions as an electron acceptor and alters the ux of PEP,
which metabolises to pyruvate and lactate/ethanol at
low CO2 levels but makes succinate at high CO2 levels.
Hydrogen was shown to increase the succinate/acetate product ratio during glucose fermentation by
A. succinogenes in minimal medium from 1 to 1.5 (Van
der Werf et al. 1997). Low-redox-potential electrons
generated by pyruvate oxidation are used to reduce fumarate to succinate (see Fig. 2). Consequently, during
glucose fermentation the molar ratio of succinate/acetate is one. In the presence of glucose and hydrogen, A.
succinogenes can generate low-redox-potential electrons
by its hydrogenase, therefore, more PEP ows to
succinate than to pyruvate.
Since the discovery of PEP carboxykinase (Utter
and Kurahashi 1953) only a few studies have been
conducted on the purication of the prokaryotic enzyme (Goldie and Sanwal 1980; Salvarrey et al. 1989;
Podkovyrov and Zeikus 1993; Schocke and Weimer
1997). PEP carboxykinase from anaerobic non-ruminal
A. succiniciproducens has been puried and characterized (Podkovyrov and Zeikus 1993). The enzyme was
oxygen-stable and had a pH optimum of 6.57.1, being
stable from pH 5.0 to 9.0. The enzyme displayed Michaelis-Menten kinetics for the substrate PEP and the
cosubstrates bicarbonate and ADP with a Km of
0.54 mM, 17 mM and 0.42 mM respectively. The enzyme required Mn2+ or Co2+ in addition to Mg2+ to
exhibit maximum activity (Podkovyrov and Zeikus
1993). This enzyme has also been puried and characterized from the anaerobic ruminal bacterium Ruminococcus avefaciens (Schocke and Weimer 1997). In
comparison, in the nonruminal A. succiniciproducens,
the apparent Km values are two- to sixfold lower for
the substrates in the direction of PEP carboxylation
than for those in the direction of oxaloacetate decarboxylation.
The A. succiniciproducens pckA gene has been cloned,
sequenced, and expressed in E. coli (Laivenieks et al.
1997). The sequence of the A. succiniciproducens was
similar to those of all known ATP/ADP-dependent PEP
carboxykinases. In particular, the A. succiniciproducens
enzyme was 67.3% identical and 79.2% similar to the
E. coli enzyme. The recombinant enzyme was overexpressed in E. coli but this did not lead to increased
succinate production. The puried enzyme was indistinguishable from the native enzyme by sodium dodecyl

sulfate/polyacrylamide gel electrophoresis and had the

same activity as the native enzyme (Laivenieks et al.
A. succiniciproducens produces succinate and acetate
in a ratio of about 4:1 in glucose-containing complete
medium, thus a considerable amount of carbon is lost as
acetate. A uoroacetate-resistant variant of the nonruminal A. succiniciproducens produced succinic acid with
very minute amounts, if any, of acetic acid (Guettler and
Jain 1996). Similarly, uoroacetate-resistant variants
have also been derived from the ruminal facultative
bacterium A. succinogenes and have produced succinic
acid at a concentration of about 110 g/l (Guettler et al.
1996). Genetically engineered strains of E. coli have been
developed that yield up to 45 g/l succinate (Millard et al.
1996; Stols and Donnelly 1997).

Fermentation and succinic acid recovery systems

Fermentation as a process technology for producing
industrial organic chemicals in general, and organic acids in particular, has been known for more than a century. Anaerobic fermentations for production of
succinic acid operate optimally at pH values where the
succinate salt rather than the free acid is produced
(Datta et al. 1992). However, the free acid is required for
the manufacture of commodity and specialty chemicals.
Moreover, contaminating proteins and cell by-products
need to be removed from the nal product. Thus, to be
cost eective, the separation process requires removal of
cells and protein-like impurities, concentration of the
salt, conversion of the succinic salt into free acid, and
polishing of the free acid to its required purity (Datta
1992; Datta et al. 1992). The cost of ineciencies associated with product recovery, concentration, acidication and purication have been very high in the past,
and have made fermentation-based chemical technology
impractical. Normally the downstream purication costs
account for 60%70% of the product cost (Baniel and
Eyal 1995). However, improvement in existing technologies along with the recent development of new technologies are enabling the recovery and purication of
fermentation-based succinic acid from its dilute broth
and making the production of succinic-acid-based
chemicals from carbohydrates via fermentation practical
and economically attractive.
One of the classical methods to recover organic acids
from fermentation broth is liquid/liquid extraction. This
is a well and extensively used operation in the chemical
industry (Hanson 1971) and there have been several
eorts in the past two decades to develop process
technologies for fermentation-based products. A number of patents and review articles have been published in
this area (King and Paul 1987; King and Starr 1992;
Baniel and Eyal 1995; King and Poole 1995; Baniel
1998a, b; Baniel et al. 1998). With one major exception,
none of these has been carried through to major pro-


duction. The exception is a citric acid extraction process

using a tertiary amine extractant and back tertiary extraction into water at high temperature (Baniel and Eyal
Another traditional product recovery method is
based on simultaneous fermentation and crystallization
of the calcium salt upon addition of calcium hydroxide
to the aqueous fermentation broth (Datta 1992; Datta
et al. 1992; Berglund et al. 1991). The crystals can be
recovered by simple ltration and the cells and proteins
can be removed by washing the crystals. The washed
crystals can be converted into soluble succinic acid solution and insoluble calcium sulfate solution by adding
concentrated sulfuric acid solution. The free succinic
acid could be ltered, polished with activated carbon or
ion-exchange resins and crystallized to produce pure
succinic acid crystals. The disadvantage of this process is
handling large amounts of solids and slurry and producing equal amounts of calcium sulfate waste product,
which needs to be treated and disposed of.
Another emerging technology, which holds higher
potential for commercialization and environmentally
friendly processes for succinic acid production, is
simultaneous electrodialysis acidication and crystallization technology, and simultaneous acidication by
ion-exchange resins and crystallization process (Zeikus
et al. 1995). Figure 3 shows the simultaneous acidication by bipolar membrane electrodialysis and purication by crystallization process. Carbohydrate is
Fig. 3 Succinic acid production by simultaneous acidication and
purication by electrodialysis and crystallization; CSL corn steep
liquor, ED electrodialyzed; CIP cleaning-in-place

enzymatically hydrolyzed, blended with other nutrients,

and fed to a continuous seed fermentor. The fermentor is
nished in a sequencing batch fermentor. Carbon dioxide (CO2) is required by the organism and sparged into
the fermentor. The fermentation is neutralized by a sodium hydroxide (NaOH) solution, forming soluble sodium succinate during the fermentation. The whole
broth is ltered with a microlteration unit to separate
the cells and large insoluble particles from the succinate
broth. The ltered sodium succinate is fed to a batch
desalting electrodialysis unit, where the ionic species are
separated from the non-ionic species (sugars) and molecules with large molecular masses (proteins and polysaccharides) under the inuence of a direct electric
current. The desalting electrodialysis membranes, containing ion-exchange groups, have a xed charge.
Membranes with positively xed charged groups (anion
membranes) selectively allow the passage of succinate
ions and repel sodium ions, and the negatively charged
(cation) membranes selectively pass sodium ions and
repel succinate ions. By this mechanism, sodium
succinate and other ionic species are transported across
the ion-exchange membranes and separated from concentrated sugars, proteins and amino acids. This concentrated sodium succinate solution is passed through a
set of chelating ion-exchange columns to replace the
divalent cation with sodium ions. The softening step is
necessary to prevent fouling of the bipolar membranes.
The sodium succinate solution is then fed to a batch
bipolar membrane electrodialysis unit where the ionic
species are converted to their equivalent acid and base
forms and separated. A bipolar membrane is a new class
of membranes, it can produce protons (H+) and hy-







Spent Cells

Cooling Water



Base Recycle




ED Concentrate Succinate
& CIP Fluids

Acid Crystals

Spent Broth
50% Succinic
Acid Solution
Crystallizer Mother Liquor Recycle


Chelating Ion
Exchange Columns

Spent Broth

ED Electrolytes
& CIP Fluids



droxyl (OH ) ions from water in aqueous solution and

cause the ions to migrate toward the electrode of opposite charge. Sodium succinate is converted to succinic
acid, which is enriched as the process proceeds. Sodium
ions are transported across the cation membrane and
associate with the hydroxyl ions to form sodium hydroxide, which is reused for fermentor neutralization.
Finally, the succinic acid solution is passed through
an evaporative crystallizer to produce very pure succinic
acid crystals (Glassner and Datta 1992). The production
of succinic acid by this process is cost-eective and can
easily be scaled-up for commercial use. The major limitation of this process is that the membranes can not
handle divalent cations; therefore, fermentations neutralized with magnesium or calcium hydroxides can not
be acidied and puried by this process.

Conclusions and future directions

Fermentation-derived succinic acid is an economic process for supplying the existing succinic acid specialty
chemical markets. However, the price per kilogram in
large volumes must drop from $ 0.55 to below $ 0.45 to
open up the commodity markets for succinic-acidderived industrial products. Production of succinic acid
by fermentation can generate signicant new markets for
agricultural carbohydrates. Figure 4 illustrates the potential impact of succinic acid fermentations on the
utilization of U.S. corn for the production of four-carbon intermediate chemicals. In the current decade, fermentation-produced lactic acid has become a biobased
commodity chemical, joining ethanol and citric acid as
green intermediate chemicals. In the next decade, improvements in the yields of succinic-acid-producing
strains and lower-cost product recovery technologies
should enable commodity-scale production. It should be

noted that the commodity price for glucose is approximately $ 0.18/kg and, because more than 1 kg succinic
acid is produced/kg glucose, a future target production
price of $ 0.35/kg succinic acid seems reasonable.
The theoretical chemical yield for succinic acid is:
Theoretical chemical yield improvements can also be
made with strains that consume H2 or HCOOH as follows:
C6 H12 O6 2CO2 2 H2
C6 H12 O6 2 HCOOH
These theoretical succinate yields have not yet been
achieved in practice. We are currently demonstrating, in
electrochemical bioreactor systems using electron mediators, that H2 can be replaced by electricity as an electron donor for enhanced succinate production (Park
DH, Laivenieks M, Guettler MV, Jain MK, Zeikus JG,
unpublished data). During fermentation by Actinobacillus on glucose plus electricity, electricity was found to
enhance glucose consumption, growth and succinate
production by approximately 25%, while it decreased
acetate production by approximately 50%. Genetic
engineering techniques now need to be used on
A. succinogenes strains to achieve the theoretical
succinate yield of two moles of succinate from one glucose and two CO2.
Similarly, there is still the considerable task of lowering the costs of succinic acid recovery technology.
Decreasing the number of unit operations and developing lower-cost liquid-extraction procedures appear to
be promising ways of improving the overall process

127 Million Bushel

4.0 Billion Lbs.



3.6 Billion Lbs.

2 Succinic Acid

2.4 Billion Lbs.

Adipic Acid, 1,4-Butanediol,

Tetrahydrofuran, Gamma Butyrolactone
USES: Plants, Clothing, Carpets, Nylon
Polyesters, Manufacturing Plastics

Fig. 4 Opportunity for the industrial use of corn to make succinic

acid derivatives in the U.S. market

Ahmed I, Morris D (1994) Replacing petrochemical with biochemicals. Institute for Local Self Reliance, Washington, DC
Alam KY, Clark DP (1989) Anaerobic fermentation balance of
Escherichia coli as observed by in vivo nuclear magnetic resonance spectroscopy. J Bacteriol 171: 62136217
Baniel AM (1998a) Recovery of carboxylic acid from organic solutions that contains an amine and an extraction enhancer. US
patent 5,780,276
Baniel AM (1998b) Recovery of carboxylic acid from organic solution that contains an amine and an extraction enhancer. US
patent 5,773,653
Baniel AM, Eyal AM (1995) Citric acid extraction. US patent
Baniel AM, Eyal AM, Mizrahi J, Hazan B, Fisher RR, Kolstad JJ,
Stewart BF (1998) Lactic acid production, separation and/or
recovery process. US patent 5,780,678; US patent 5,510,526
Bergen WG, Bates DB (1984) Ionophores: their eect on production eciency and mode of action. J Anim Sci 58: 14651483
Berglund KA, Elankovan P, Glassner DA (1991) Carboxylic acid
purication and crystallization process. US patent 5,034,105

Bryant MP, Small N (1956) Characteristics of two new genera of
anaerobic curved rods isolated from the rumen of cattle.
J Bacteriol 72: 2226
Bryant MP, Bouma C, Chu H (1958) Bacteroides ruminicola n. sp.
and the new species Succinomonas amylolytica. Species of succinic acid producing anaerobic bacteria by the bovine rumen.
J Bacteriol 76: 1523
Clark DP (1989) The fermentation pathways of Escherichia coli.
FEMS Microbiol Rev 63: 223234
Dake S, Gholap RV, Chaudhuri RV (1987) Carbonylation of 1,4butanediol diacetate using rhodminum complex catalyst: a kinetic study. Ind Eng Chem Res 26: 15131518
Datta R (1992) Process for the production of succinic acid by anaerobic fermentation. US patent 5,143,833
Datta R, Glassner DA, Jain MK, Vick Roy JR (1992) Fermentation and purication process for succinic acid. US patent
Davis CP, Cleven D, Brown J, Balish E (1976) Anaerobiospirillum,
a new genus of spiral-shaped bacteria. Int J Syst Bacteriol 26:
Diamantoglon M, Meyer G (1988) Process for the production of
water-insoluble bers of cellulose monoesters of maleic acid,
succinic acid and phthalic acid, having an extremely high absorbability for water and physiological liquids. US patent
Glassner DA, Datta R (1992) Process for the production and purication of succinic acid. US patent 5,143,834
Goldie AH, Sanwal BD (1980) Allosteric control by calcium and
mechanism of desensitization of phosphoenol pyruvate carboxykinase of Escherichia coli. J Biol Chem 255: 13991405
Guettler MV, Jain MK (1996) Method for making succinic acid,
Anaerobiospirillum succiniciproducens variants for use in process
and methods for obtaining variants. US patent 5,521,075
Guettler MV, Jain MK, Rumler D (1996) Method for making
succinic acid, bacterial variants for use in the process, and
methods for obtaining variants. US patent 5,573,931
Guettler MV, Rumler D, Jain MK (1999) Actinobacillus succinogenes sp. nov., a novel succinic acid producing strain from the
bovine rumen. Int J Syst Bacteriol 49: 207216
Hanson J (1971) Recent advances in liquid-liquid extraction.
Pergmon, Oxford
Jain MK, Datta R, Zeikus JG (1989) High-value organic acids
fermentation emerging processes and products. In: Ghosh TK
(ed) Bioprocess engineering: the rst generation. Ellis Harwood,
Chichester, pp 366398
Kaneuchi C, Seki M, Komagata K (1988) Production of succinic
acid from citric acid and related acids by Lactobacillus strains.
Appl Environ Microbiol 54: 30533056
King JC, Paul A (1987) Process for sorption solute recovery. US
patent 4,670,155
King JC, Poole LJ (1995) Carboxylic acid absorption regeneration
process. US patent 5,412,126
King JC, Starr J (1992) Recovery of carboxylic acids from water by
precipitation from organic solutions. US patent 5,104,492
Kinnersley A, Coleman R, Tolbert E (1997) Method for stimulating plant growth using GABA and succinic acid. US patent
Laivenieks M, Vieille C, Zeikus JG (1997) Cloning, sequencing,
and overexpression of the Anaerobiospirillum succiniciproducens
phosphoenol pyruvate carboxykinase (pckA) gene. Appl Environ Microbiol 63: 22732280

Millard CS, Chao Y-P, Liao JC, Donnelly MI (1996) Enhanced

production of succinic acid by overexpression of phosphoenol
pyruvate carboxylase in Escherichia coli. Appl Environ Microbiol 62: 18081810
Nghiem NP, Davison BH, Suttle BE, Richardson GR (1997)
Production of succinic acid by Anaerobiospirillum succiniciproducens. Appl Biochem Biotechnol 6365: 565576
O'Herrin SM, Kenealy WR (1993) Glucose and carbon dioxide
metabolism by Succinivibrio dextrinosolvens. Appl Environ
Microbiol 59: 748755
Podkovyrov S, Zeikus JG (1993) Purication and characterization
of phosphoenol pyruvate carboxykinase, a catabolic CO2-xing
enzyme, from Anaerobiospirillum succiniciproducens. J Gen
Microbiol 139: 223228
Sado T, Tajima M, Noguchi Research Foundation (1980) Carbonylation of 1,4-butanediol by homogeneous catalyst. Japan
Kakai Tokkyo Koho 8,051,037
Salvarrey MS, Cannata JJB, Cazzulo JJ (1989) Phosphoenol
pyruvate carboxykinase from the moderate halophile Vibrio
costicola. Biochem J 260: 221230
Samuelov NS, Lamed R, Lowe S, Zeikus JG (1991) Inuence of
3 levels and pH on growth, succinate production,
and enzyme activities of Anaerobiospirillum succiniciproducens.
Appl Environ Microbiol 57: 30133019
Scheinger CC, Wolin MJ (1973) Propionate formation from cellulose and soluble sugars by combined cultures of Bacteroides
succinogenes and Selenomonas ruminantium. Appl Microbiol 26:
Schocke L, Weimer PJ (1997) Purication and characterization of
phosphoenol pyruvate carboxykinase from the anaerobic
ruminal bacterium Ruminococcus avefaciens. Arch Microbiol
167: 289294
Stols L, Donnelly MI (1997) Production of succinic acid through
overexpression of NAD+-dependent malic enzyme in an
Escherichia coli mutant. Appl Environ Microbiol 63: 2695
Turk R (1993) Metal free and low metal salt substitutes containing
lysine. US patent 5,229,161
Utter MF, Kurahashi K (1953) Mechanism of action of oxalacetic
carboxylase from liver. J Am Chem Soc 75: 758
Van der Werf MJ, Guettler MV, Jain MK, Zeikus JG (1997) Environmental and physiological factors aecting the succinate
product ratio during carbohydrate fermentation by Actinobacillus sp. 130Z. Arch Microbiol 167: 332342
Weimer PJ (1993) Eects of dilution rate and pH on the ruminal
cellulolytic bacterium Fibrobacter succinogenes S85 in cellulosefed continuous culture. Arch Microbiol 160: 288294
Wollenberg RH, Frank P (1988) Modied succinimides in fuel
composition. US patent 4,767,850
Wood WA (1961) Fermentation of carbohydrates and related
compounds. In: Gunsalus IC, Stanier RY (eds) The bacteria,
vol 2. Academic Press, New York London, pp 59149
Zeikus JG (1980) Chemical and fuel production by anaerobic
bacteria. Annu Rev Microbiol 34: 423464
Zeikus JG, Elankovan P, Grethlein A (1995) Utilizing fermentation
as a processing alternative: succinic acid from renewable resources. Chem Proc 7173
Zwicker N, Theobald U, Zahner H, Fiedler HP (1997) Optimization of fermentation conditions for the production of
ethylene-diamine-disuccinic acid by Amycolatopsis orientalis.
J Ind Microbiol Biotechnol 19: 280285