Khd

ci C^H2OH

ß-d-Glucose (4C i chair) Figure 4. Structures of D-glucose.

ß-d-Glucose (4C i chair) Figure 4. Structures of D-glucose.

temperatures and pressures. Dextrose content of the starch hydrolysate was limited to about 86% dry basis (db) as a result of the formation of degradation products (11). Dextrose content was increased by partial or complete replacement of acid with one or more enzymes. These processes are referred to as acid-enzyme (A-E) or enzymeenzyme (E-E), depending on whether initial starch hydrolysis (the starch thinning or liquefaction step) is conducted with acid or a bacterial a-amylase. In either case, subsequent conversion to dextrose (saccharification) is achieved with glucoamylase, a fungal enzyme that releases dextrose from the nonreducing end of starch polymers by successive hydrolysis of a-1,4 and 1,6 glucosidic linkages.

By 1960, the A-E process was in general use, and hydrolysate dextrose content was increased to 92 to 94% db (12). A typical process involved thinning a 30 to 35 wt % starch slurry with acid (HC1 or H2S04) at a temperature and pressure necessary to achieve a DE level of 10 to 20. DE is a measure of the reducing-sugar content of a starch hydrolysate calculated as dextrose and expressed as a percentage of the total dry substance. Formation of acid-

reversion products during thinning limited the dextrose yield attained during saccharification. Higher dextrose content was achieved by the E-E process that was developed in the 1960s. This process used a heat-resistant bacterial a-amylase for starch thinning. Lower temperature and nearly neutral liquefaction conditions limited side reactions and resulted in hydrolysate dextrose contents of 95 to 97% db (11,13). Initial processes were based on a-amylase derived from the bacterium Bacillus subtilis. In a typical process, 30 to 40 wt % starch is thinned at 85°C (185°F) for 1 h followed by a short heat treatment at 120 to 140°C (248 to 284°F). Then, a second enzyme addition is made, and reaction at 85°C (185°F) is continued to complete the liquefaction step (Fig. 5). The resulting 10 to 15 DE hydrolysate is then saccharified with glucoamylase. The high temperature heat treatment is required to solu-bilize insoluble starch particles that are believed to be amylose-fatty acid complexes formed during initial liquefaction (14). Since the high temperature inactivates the enzyme, a second enzyme addition is necessary to continue hydrolysis before the complex reforms.

Elimination of the heat treatment step was made possible by the commercialization of a-amylases that are extremely thermostable and capable of efficient starch hydrolysis at a temperature above 100°C (212°F). Enzymes derived from Bacillus stearothermophilus (15) and Bacillus licheniformis (16) are used for this purpose (Fig. 6). Starch slurry is thinned continuously with a steam-injection heater at 30 to 40 wt % solids, pH 6 to 6.5, and 103 to 107°C (217 to 225°F) for 6 to 10 min. A 1 to 2 h hold at about 95°C (203°F) completes the thinning and yields a 10 to 15 DE

10-15 DE hydrolysate

Figure 5. Simplified process flowsheet. Starch liquefaction using Bacillus subtilis a-amylase.

10-15 DE hydrolysate

Figure 5. Simplified process flowsheet. Starch liquefaction using Bacillus subtilis a-amylase.

10-15 DE hydrolysate

Figure 6. Simplified process flowsheet. Starch liquefaction using Bacillus stearothermophilus or Bacillus licheniformis a-amylase.

10-15 DE hydrolysate

Figure 6. Simplified process flowsheet. Starch liquefaction using Bacillus stearothermophilus or Bacillus licheniformis a-amylase.

hydrolysate for saccharification. The high initial temperature is sufficient to disrupt the amylose-lipid complex. Concurrently, the thermostable enzyme hydrolyzes the starch to a point where reassociation of the complex cannot occur. The thermostable a-amylases are able to survive the initial liquefaction with little loss in activity since the halflife of a-amylases derived from B. stearothermophilus and B. licheniformis is about 2 h (15) and 1 h (16), respectively, at 105°C (221°F).

Regardless of the type of thinning utilized, dextrose is produced by the action of glucoamylase during saccharification (17). Glucoamylase is produced from strains of Aspergillus niger in submerged fermentation. A broth is obtained containing two or more glucoamylase isozymes, a-amylase, transglucosidase, and other enzymes, eg, protease and cellulase. The a-amylase assists in saccharification (11); however, transglucosidase catalyzes the formation of isomaltose and, therefore, must be removed before use. Removal of the enzyme is accomplished by adsorption on a clay mineral or by other techniques. Alternatively, selection for an Aspergillus mutant that does not produce transglucosidase eliminates the need for a removal step (18). Glucoamylase is also produced froma/i/ii-zopus organism in Japan by surface fermentation; however, enzyme properties are somewhat inferior to those of Aspergillus.

Saccharification of liquefied starch hydrolysate is conducted by batch or continuous processes in large agitated reactors that are often several million liters in size. The reaction is conducted at 58 to 61°C (136 to 142°F), pH 4 to 4.5 with a glucoamylase dosage that is sufficient to produce the maximum dextrose yield in 1 to 4 days. At the normal solids level of 30 to 35 wt %, maximum hydrolysate dextrose content is about 95 to 96% db. During saccharification, isomaltose and maltose are produced by the reverse reaction (reversion) of glucoamylase, in which two dextrose molecules are combined. Consequently, if the reaction is extended beyond the time needed to achieve maximum dextrose content, the dextrose level decreases as a result of excessive formation of these reversion products.

Dextrose content can be increased by conducting saccharification at lower solids. As solids level is reduced, the forward reaction is favored and a higher dextrose content is achieved. At a solids level of 10 to 12 wt %, a dextrose content of 98 to 99% db can be attained (19). However, operation at low solids is not commercially viable because of increased evaporation cost, the need for larger saccharification tanks, and the risk of microbial infection.

Dextrose level can also be increased by using enzymes that enhance the action of glucoamylase during saccharification. Enzymes available for this purpose are pullulan-ase and Bacillus megaterium amylase. Pullulanase is specific for the hydrolysis of a-1,6 glucosidic linkages and, when used in combination with glucoamylase, increases the rate and extent of dextrose production (20). A B. megaterium enzyme has been commercialized that combines hydrolysis and transferase activity to assist glucoamylase in increasing dextrose content (21). Either enzyme is effective in increasing hydrolysate dextrose content by 0.5 to 1.0% db.

After saccharification, the high dextrose content hydrolysate is clarified by centrifugation or filtration (usually either precoat or membrane) to remove insolubles. Concentration (usually by evaporation) to lower the water content and refining to reduce color and ionic contaminants are the next steps. Refining can be done with activated carbon (either powdered or granular), or with ion exchange resins (decolorizing and/or demineralizing resins), either singly or in combination. A simplified flowsheet for the manufacture of high dextrose content products is shown in Figure 7.

Commercial dextrose products include crystalline monohydrate or anhydrous dextrose and liquid dextrose. In addition, the hydrolysate can be refined and evaporated to a high DE corn syrup or dried to a solid product. Compositions are listed in Table 4.

Crystalline dextrose products can be manufactured using either batch or continuous crystallization technology. Sites for crystal growth can be provided either by the addition of seed crystals or by spontaneous nucleation. Subsequent crystal growth is encouraged by maintaining a controlled degree of supersaturation. Control of the entire process is critical to ensure that the resulting crystals are of a size and shape that will readily separate from the remaining mother liquor in perforate bowl centrifuges to provide a dry product of high purity.

The predominant technique for the production of monohydrate dextrose uses batch cooling-type crystallizers, although continuous crystallization using either evaporation (23) or cooling (24) to maintain supersaturation have been described. A process employing a continuous precrystal-lizer supplying batch crystallizers has been used to increase productivity (25).

In the dominant process, clarified, refined, and concentrated liquor having a dextrose content above about 80% db is mixed with a substantial bed of seed crystals (typically 20 to 30 wt %) left from the previous batch in the

Starch hydrolysis {liquefaction and saccharification)

Clarification, concentration, and refining

Chromatographic separation

Crystallization, centrifugal separation, and drying

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