Gluconic Acid Production

2.2. Gluconic Acid Production

Gluconic acid possesses enormous commercial applications in a variety of product and product utilities in pharmaceuticals as well, namely :

manufacture of metal, leather and food. Calcium gluconate to provide and supplement Ca 2+ to pregnant mothers.

Na + and Ca 2+ salts in alkaline serve as effective metal sequestering agents for Fe 3+ , Al 3+ , Cu 2+ . Ferrous gluconate caters for Fe for the treatment of anemia in humans. Sodium gluconate finds its use as a sequestering agent in detergents. δδδδδ -gluconolactone functions as a baking powder additive.

In the domain of ‘industrial microbiology’ gluconic acid enjoys a creditable long historical evidence. Alsberg (1911) first and foremost reported the production of gluconic acid commencing with the Pseudomonas. In 1928, the first ever commercial surface process by making use of a ‘fungus’, Penicillium leuteum-purpurogenum, saw the light of the day gracefully which remarkably gave a yield ranging between 80-87% of its theoretical values.

Interestingly, as to date the submerged processes have gained a world-wide acceptance and recog- nition by employing either the fungus—Aspergillus niger or the bacterium—Acetobactor suboxydans, which ultimately gave rise to a range of value-added products, such as : gluconic acid ; glucose oxidase ; and sodium — and calcium-gluconates.

Note : There are several organisms that have been duly optimized via research to yield ‘gluconic acid ’, but unfortunately have not been exploited commercially, prominently include the fol- lowing : fungi : Endomycopsis, Gonatobotrys, Penicillinum, Pullularia, Scopulariopsis ; bacteria : Vibrio, Pseudomonas.

Fermentation Production. The production of gluconic acid from glucose is predominantly carried out by a flavoprotein which is termed as glucose-oxidase*, or more precisely glucoseaerode- hydrogenase that essentially serves as an enzyme mediating this oxidation. The overall reaction takes place in two steps as illustrated below :

C COOH

CHOH 2 STEP 2

HO 2 2 O 2 CHOH

FAD = Flavin Adenine Dinucleotide FADH 2 = Flavin Adenine Dinucleotide Dihydrate

* It has also been identified as a ‘antibiotic’ in fermentation broths by virtue of its antimicrobial activity ; and,

PHARMACEUTICAL BIOTECHNOLOGY

Step-1 : The dehydrogenation of a mole of D-glucose in the presence of FAD gives rise to the formation of δ -D-gluconolactone. Besides, the transferance of 2 H-atoms from FADH 2 to oxygen (O 2 ) yields a mole of hydrogen peroxide (H 2 O 2 ) that gets instantly split up into a mole of water (H 2 O) by the help of the enzyme catalase. Step-2 : The resulting product obtained from Step-1 i.e., δ -D-gluconolactone takes up the water from the previous step and legitimately yields a mole of gluconic acid.

Methodology : The various sequential steps involved in the production of gluconic acid are as stated under :

(1) The Aspergillus niger (fungus) — gluconic acid fermentation may be preferably accom- plished by submerged culture process usually at pH ranging between 4.5 to 6.5, predomi- nantly needs a growth culture medium wherein both P and N are limiting.

(2) The A. niger mycelium, once formed in an initial gluconic acid-growth fermentation ad- equately, is judiciously reutilized in the ensuing successive ‘replacement culture’ fermentations, of course, as long as the prevailing glucose-oxidase activity of the mycelium remains highly virile and active.

(3) Soonafter the initial fermentation process yields the mycelium, the consequent successive fermentations are nothing but exclusively enzymatic transformations specifically brought about by the ensuing glucose-oxidase of the mycelium. In usual practice, the entire fermentative run lasts for 20 hours at 28-30°C having a high aeration rate ranging between 1-15 vvm.

(4) Recycling of the mycelium evidently gives rise to lowering of the requirement for inoculum built-up. However, as and when the system is in dire need of inoculum, it is introduced as spores or pregerminated spores directly to the production fermentor.

(5) Growth culture medium for gluconic acid production essentially comprises of glucose (25%) together with various salts, CaCO 3 , a boron compound, and sometimes even cornsteep liq- uor is added. The overall fermentation is invariably carried out at 28 ± 2°C supported by adequate agitation, aeration, and control of heat generation either using water-jacketed fermentors or employing cooling-coils.

(6) Once the fermentation is completed, the gluconic acid is recovered by neutralization of the fermented broth with calculated amount of Ca(OH) 2 in order to permit the crystallization of

calcium gluconate. Finally, gluconic acid is recovered from the calcium gluconate (salt) by the addition of a measured and calculated quantity of concentrated H 2 SO 4 .

Important Highlights : Following are some of the important highlights of gluconic acid pro- duction :

(1) Simply by enhancing the pressure in the system, the solubility of O 2 , and hence the produc- tion of gluconic acid may be accelerated commercially upto 90-95%.

(2) The replacement culture fermentation is performed, and the ensuing medium is devoid of N- containing substances to prevent further growth of the mycelium. Under these highly spe- cific and stringent production parameters the conversion of glucose into gluconic acid gets elevated upto 95%.

(3) These exists a healthy and legitimate competition between the microbiological process and

the chemical methods that ultimately give rise to high yields of gluconic acid.

MICROBIAL TRANSFORMATIONS