fermenation

1. • 2,3-Butanediol fermentation is anaerobic fermentation of glucose with 2,3-butanediol as one of the end
products. The overall stoichiometry of the reaction is
• 2 pyruvate + NADH --> 2CO2 + 2,3-butanediol.
• Butanediol fermentation is typical for the facultative anaerobes Klebsiella and Enterobacter and is tested for
using the Voges–Proskauer (VP) test.
• The metabolic function of 2,3-butanediol is not known, although some have speculated that it was an
evolutionary advantage for these microorganisms to produce a neutral product that's less inhibitory than other
partial oxidation products and doesn't reduce the pH as much as mixed acids.
• There are many important industrial applications that butanediol can be used for, including antifreeze, food
additives, antiseptic, and pharmaceuticals. It also is produced naturally in various places of the environment.
2,3-Butanediol fermentation

2. Comparison with mixed acid fermentation
• 2,3-butanediol fermentation produces smaller amounts of acid than mixed acid fermentation, and
butanediol, ethanol, CO2 and H2 are the end products. While equal amounts of CO2 and H2 are created during
mixed acid fermentation, butanediol fermentation produces more than twice the amount of CO2 because the
gases are not produced only by formate hydrogen lyase like they are in the mixed acid fermentation
• 2,3 Butanediol is produced at varying levels in aerated fermentations as long as the dissolved oxygen level is
limiting (i.e., the culture is trying to consume more oxygen than is available). The degree of oxygen limitation
dictates the ratios of 2,3-butanediol to by-products produced.
• Under anaerobic conditions, 2,3-butanediol-producing facultative anaerobes produce acidic products, lowering
the external and intracellular pH.
• α-acetolactate synthase, which catalyzes the first reaction to produce 2,3-butanediol, has an optimum at pH
6.0.
• When the intracellular pH drops, this enzyme becomes active to divert carbon flux from acid production to the
neutral solvent.
• An enzyme catalyzing the same reaction catalyzes the first reaction of valine synthesis from pyruvate.
• This enzyme is referred to as the pH 8.0 enzyme while the enzyme involved in 2,3-butanediol synthesis
is referred to as the pH 6.0 enzyme.
• Klebsiella pneumoniae, Klebsiella oxytoca and Enterobacter aerogenes ferment glycerol to various products
including 2,3-butanediol.

3. Butanediol fermentation by some Gramnegative facultative anaerobic bacteria. Facultative anaerobes
belong to the genera Erwinia, Klebsiella and Serratia and produce 2,3-butanediol in addition to lactate and
ethanol. 1, EMP pathway; 2, lactate dehydrogenase; 3, pyruvate:formate lyase; 4, formate:hydrogen lyase;
5, acetaldehyde dehydrogenase; 6, alcohol dehydrogenase; 7, α-acetolactate synthase; 8, α-acetolactate
decarboxylase; 9, 2,3-butanediol dehydrogenase.

4. • pyruvate also undegoes reduction or oxidation generating various organic acids and alcohols including lactate,
formate, acetate, succinate, ethanol and acetoin. In order to minimize such unwanted byproducts, several
approaches including mutant screening, genetic engineering and fermentation technology were suggested for
enhancing 2,3-BD production .
• In addition, pyruvate is transformed into acetolactate by two distinct enzymes. The first enzyme is catabolic α-
acetolactate synthase, which has optimum pH value of 5.8 in acetate, and is part of the butanediol pathway. The
second enzyme in the butanediol biosynthesis pathway is acetolactate decarboxylase, with an optimum pH value
of 6.3, and it catalyzes the formation of acetoin via the decarboxylation of acetolactate. There is also a third
enzyme named diacetyl (acetoin) reductase, which catalyzes a reversible reduction of acetoin to 2,3-BD and an
irreversible reduction of diacetyl to acetoin. Diacetyl (acetoin) reductase is a tetrameric enzyme which uses
NADH as a coenzyme

5. Industrial applications
• 2,3-butanediol has a variety of industrial applications and products it can produce.
• The levo isomer of butanediol has a low freezing point of -60 °C, which allows it to work as an antifreeze
agent. Through catalytic dehydrogenation, butanediol can form diacetyl. Diacetyl is a food additive that can
be used to add flavor.
• 0.1% butanediol will kill most pathogenic bacteria due to its antiseptic properties. Through esterification,
forms of precursors of polyurethane foams are produced. These can be used in various applications,
including in pharmaceuticals, cosmetics, lotions, ointments, and antiperspirants. Butanediol itself even has
applications in the pharmaceutical industry as a drug carrier

6. Fig. The metabolic pathways for 2,3-butanediol synthesis in bacteria (modified, based on Magee & Kosaric,
1987). 1, glycolysis and pentose phosphate pathway enzymes; 2, pyruvate kinase; 3, α-acetolactate synthase;
4,α-acetolactate decarboxylase; 5, acetoin reductase (2,3-butanediol dehydrogenase); 6, pyruvate–formate
lyase; 7, phospho-transacetylase; 8, acetate kinase; 9, acetaldehyde dehydrogenase; 10, ethanol
dehydrogenase; 11, actate dehydrogenase; 12, phosphoenolpyruvate decarboxylase ; 13, malate
dehydrogenase; 14, fumarase; 15, succinate dehydrogenase.