LANTS SODIUM GLUCONATE PRODUCTION Fermentation with Aspergillus niger R. H. BLOW, V. F. PFEIFER, A. J. MOYER, D. H. TRAUFLER, AND H. F. CONWAY Northern Regional Research Laboratory, Peoria 5, 111.
C. K. CROCKER, R. E. FARISON, AND D. V. HANNIBAL The Diversey Corp., Chicago, I l l .
C
AUSTIC soda solutions in natural waters have extensive use in industry. Most waters contain significant amounts of calcium and magnesium salts which cause the formation of precipitates in the caustic soda solutions. It has been discovered recently ( 1 ) that sodium gluconate has the property of preventing the formation of the insoluble hard water precipitates in natural waters t o which caustic soda is added. Such an effect has been observed to occur over a wide range of caustic soda concentrations. The quantity of sodium gluconate required to prevent this precipitation naturally varies with the degree of hardness of the water, but sodium gluconate amounting to 5 or 10% of the weight of the caustic soda appears t o be sufficient for waters usually encountered commercially. The possibility that this observation would have commercial significance is immediately apparent since, if hard water salts are prevented from precipitating in caustic soda solutions, it might be possible t o avoid scale deposition in automatic equipment such as is commonly used in bottle washing machines. These machines are used by the dairy, carbonated beverage, and brewing industries. T h e formation of scale almost invariably occurs and lessens the life of the machines, as a result of increased load and wear. The cost of operations is increased because of the loss of caustic soda in the porous scale that is removed periodically from the equipment. Other applications are apparent in the textile industry where the use of sodium gluconate might prevent deposition of hard water salts on the fabric, thus obviating the need for their subsequent removal. Since these commercial applications might well become extensive, especially if sodium gluconate were available a t a low (#ofit,this investigation was undertaken t o determine the feasibility of the direct production of sodium gluconate and t o obtain eufic-ient.data for estimation of investment and operating costs. Gluconic wid may be formed as the main product of fermentation when glucose solutions, appropriately supplemented with other nutrients, are subjected to fermentation by various molds and bacteria. The gluconic acid fermentation has been studied extensively by May, TIcrrick, Moyer, Hellbach, Wells, Stubbs, and others (2-10). It was shown by Wells, Moyer, Stubbs, Herrick, and May ( I O ) that the acid may be produced successfully from glucose by fermentation with Aspergillus niger, a mold possessing many desirable characteristics for this purpose. With this organism it is necessary t o neutralize the fermenting medium 1
continuously if the fermentation is to consume the glucose efficiently, since high acidity is inhibitory t o the fermentation (6). Calcium carbonate has been generally chosen for continuous neutralization of the acid as it is formed, aince it may be incorporated into the original medium with no interference t o the fermentation. T h e product of the fermentation studies mentioned is calcium gluconate which has uses in the p h s b c e u t i c a l and medical fields. Sodium gluconate may bt. prepared from gluconic acid by neutralization with a sodium base, or it may be prepared from calcium gluconate by acidification with sulfuric acid, filtration to remove calcium sulfate, and subsequent neutralization with a sodium base. Since these procedures are necessarily expensive and time consuming, it is debiwble t o neutralize continuously the gluconic acid formed during fermentation of glucose with a sodium base so that the main product of fermentation is sodium gluconate. This paper describes pilot plant experiments in which sodium gluconate was produced directly by continuous neutralization of gluconic acid formed during submerged culture fermentation of glucose with A . niger NRRL-3. The acid, as produced, was neutralized with sodium hydroxide a t a controlled pH. The engineering data and informatioa obtained were used as a basis for approximate cost calculations. EQUIPMENT AND PROCEDURES
Equipment. Pilot plant fermentations were conducted in a stainless steel fermentor of 300-gallon capacity, which was provided with a 40-gallon stainless steel caustic storage tank and an 80-gallon copper seed tank. The stainless steel fermeritor, shown in Figure 1, is 13 feet tall with an internal diameter of 24 inches and is e uipped with a half jacket. Agitation is provided by a prope8er-type blade attached t o a horizontal shaft mounted 15 inches above the bottom of the fermentor. The speed of the agitator can be varied between 100 and 250 r.p.m. The temperature of the medium can be maintained within 0.5' F. of the desired point by circulation of water through the jacket. The temperature of the cooling water is automatically controlled. ?vIoqt fermentations in this tank were conducted a t a volume of 150 gallons, corresponding to a liquid depth of approximately 7 feet. Sterile air for the fermentation was introduced through a perforated pipe-cross sparger. Air was sterilized by passing it through a 10-inch column containing 8 feet of 10- to 24mesh activated carbon. Vegetative type inoculum was grown in the 80-gallon copper seed tank. This tank is 4 feet high and 2 feet in diameter, with
Present address, The Shattrick Chemical Co.. Denver. Colo.
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INDUSTRIAL AND ENGINEERING CHEMISTRY
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Tu prepere ii npore ausptmion to be used 88 iimulum for 8 150&Ion pilot plant fermentation, ton t o twenty I-liter Erlenmeyev Naaks, each containing 150 ml. of liquid sporulation medium, were inoculated with apores of A . niyeier and inoubated a t 30" C. for ripproriniai.ely 7 days. Liquid sporulation medium had the folloxing cornposit.ion:
ictiitrate, , , ,g I'otnto axtrrtet, mi. Beer, 1n1.
0.01
:to.()
im.0 1.50 I Ooo
*gnr, grams Tap water, to makr, nil.
Use 150 nil. iii 1-liter Erlenrrieyrr flasks Stwilizntion: :10 minuins !It, 121" C. (15 lb.jxi. inch
gage)
.At the end of the ineu1,ation period, the spore-liuring myeeli:i from the tlseks and suspended in 10 liters of sterile saier containing 0.01% 01 it v-ctting agoat, such :LS sodium lauryl sulfate. This qmre suspension >viis introduced i n t o ti,(. ferb$t:ro removed
inentor toservoas inoculum.
To prepare vegetative growth inoculuin for a 150-gsllon piial plant fermentation, two Hssks of liquid sporulation medium were inoculated, incubated a t 30" C. for spproximetely 7 days, the spores suspended i n 2 liters of sterile water as above, and the suspcuuion blown into 45 g d l a ~of germinnt,ion medium prQpriced RS follows:
Figure 1.
Stainless Steel 300-GalIoii Fermentor for Sodium Glueonate Production
diahed top and conical bottom, and is equip& with b perforated pipe spsrgor for distribution of sterile air. The temperature of the inoculum was controlled by the circulation of water through internal coils. The seed tank is not provided tiith sn agitat.or. Sodium hydroxide for continuously neutralizing the gluconic acid &s formed was mixed in tho 40-gallon stainless steel supply tank. This tank is Drovided with a n nnitator and ia calibrated 80 that the volume of the caustic may be keasumd. The pH of the fermenting medium war recorded and automnt,idIv Medium from bottom ~.,contmlled. . . ~ ~ ~ . %-=. circulated ~ ~ ~ ~the~ ~ of the fermentor through an enameled-iron pH Row chamber and electrode assembly and back into the top of the fermentor. The rhnmber snd cloctrodes were suitable foro ration a t presaurea up tu 30 pounds per square inch ga c * E l i solution from the ..upply tank m s added automatic& to control the pH a t a predetermined constnnt value. The controilar incorporated throttling range and droop correction adjustments so that. effective pH control was sttaiaable. Foaming of thc medium during fermentation was id, 32, 107 (1940). Wells, P. A., Lynch, D. F. J., Herrick, H. T., :+nd S l a y , 0. E., Chem. and M e t . Eng., 44, 188 (1937).
Wells, P. A, hloycr, A. J., Stubbs, ,J, .J,, IIerrirk, H . T., and M a y , 0. E., IND.ENG.CHEM.,2 9 , G53 (1937). RECEIVEDSeptember 4 , 1951. The work dencribed 111 this paper was conducted under a memorandum of understanding between t h e Northern Kegional Research Laboratory a n d The Dlversey Corp.
