Isolation of Zein Using 100% Ethanol

Cereal Chem. 83(5):565-568 Traditionally, zein is isolated and recovered from corn gluten meal (GCM) using aqueous alcohol as the solvent. Recovery of zein from this solvent is inconvenient and costly. Zein is insoluble in 100% ethanol at room temperature, but it is soluble at 120°C in ethanol. Absolute ethanol effectively extracted zein from CGM, distillers dried grains (DDG), and ground corn. Zein was extracted from CGM with absolute ethanol in a high-pressure reactor at 130°C. After extracting at 130°C for 45 mm, the solution was pumped out of the extractor and allowed to cool. Upon cooling, the zein precipitated from solution. The precipitate was removed from the solution and air-dried, resulting in 14% recovery of the starting material. The recovered precipitate had an average protein content of >90% on a dry basis, accounting for 20% of the CGM protein and recovered 35% of its zein. No differences were seen in the amount of zein extracted from CGM samples that were hand-collected off the dewatering screen and gently dried, versus commercial CGM samples. The commercial CGM did produce a greater amount of solubles. The extraction procedure also worked at temperatures as low as 90°C. The lower temperature did produce lower yields of extracted zein. The zein extracted at the lower temperatures was less brown, but zein extracted at either temperature was almost fully soluble in traditional zein solvents. Zein was first extracted by Gorham (1821) from whole corn using aqueous ethanol. Osborne (1891) was granted the first patent for a commercial method for zein extraction. Commercial production of zein was not achieved until 1938 when the Corn Products Refining Company built a small pilot plant (Anon. 1939). A much larger plant was built by Corn Products Refining Company in 1943 (Anon. 1944). These plants recovered zein by spraying the aqueous alcohol zein solution into cold moving water (Horesi et a! 1941). Today, zein is recovered from the aqueous alcohol solvent by chilling the extract to -10 to -20°C (Lawton 2002). Zein precipitates from aqueous alcohols at these low temperatures. Spray drying or vacuum drying have been suggested as methods to recover zein from aqueous alcohol solutions (Shukla and Cheryan 2001; McInnis and Tang 2003). The problem with using aqueous ethanol is that zein remains in solution at room temperature. To recover the zein, it either has to be precipitated from the solvent, or large amounts of solvent have to be driven from zein in the drying step. Potential problems are encountered using a spray dryer to recover zein from an aqueous alcoholic solution. Alcohols are explosive under heated closed systems such as a spray dryer. Zein is a film former and has a tendency to form films as it exits the sprayer of the dryer. It also has to be totally dry by the time it hits the wall of the spray dryer. If not, it will either form films on the wall or stick to it. Either way, the zein remains in the heated dryer too long. To save time and expense, it would be desirable to find a single component solvent for zein where the solvent could easily be removed or precipitated. Zein is soluble in 100% ethanol at 120°C (Manley and Evans 1942). Once the solution is cooled to <120°C, zein precipitates and can be recovered. The objective of this work was to see if 100% ethanol at elevated temperatures would be an effective extractant for zein. MATERIALS AND METHODS Corn gluten meal (CGM) was obtained from Aventine Renewable Energy (Pekin, IL). CGM was obtained either dry (commercial United States Department of Agriculture, Agricultural Research Service, Plant Polymer Research Unit, National Center for Agricultural Utilization Research, 1815 N. University Street, Peoria, IL 61604. Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by the USDA implies no approval of the product to the exclusion of others that may also he suitable. Phone 309-681-6419. E-mail: [email protected] DOl: 10.1094/CC-83-0565 This article is in the public domain and not copyrightable. It may be freely reprinted with customary crediting of the source. AACC International, Inc., 2006. product) or wet (from their dewatering screen). Wet CGM was air-dried and ground through a pin mill. Corn grits were donated by Bunge Milling (St. Louis, MO) and distillers' dried grains were donated by Big River (Burlington IA). Ethanol and petroleum ether were reagent-grade and purchased from Sigma-Aldrich (St. Louis, MO). Samples were defatted with petroleum ether using a soxhlet extractor. Extractions were run for 55 hr. Nitrogen analysis was performed using a PerkinElmer 2400 Series II Dumas-type elemental analyzer. Approximately 2-3 mg of material was used for each measurement and each sample was run in triplicate. The instrument was calibrated using an acetanilide standard. Protein content was calculated from the nitrogen analysis using a 6.25 factor. Samples that were vacuum-dried before extraction were dried overnight at 35°C and 70 mm of Hg to <1% moisture. Moisture percent of samples was determined using the modified Approved Method 44-I5A (AACC International 2000). Because of the amount of material extracted, some samples were dried using 1 g or 0.5 g. Crude zein recovered from the ethanol extraction was redissolved in either 70% aqueous ethanol (v/v), 88% aqueous isopropanol (vlv), dimethyl foramide, 2-ethoxy ethanol, or ethyl lactate. Crude zein (1 g) was mixed with 10 mL of solvent and stirred in a closed beaker for 16 hr (overnight). After stirring overnight, the zein-solvent mixture was transferred to a tared centrifuge tube and centrifuged for 20 min at 2,000 rpm. The liquid was discarded and the solids in the tube were air-dried in the hood for 48 hr, followed by drying at 100°C for 2 hr in a forced-air oven. The solubles percent was calculated by subtracting the starting weight from the remaining weight and was an average of three solublization runs. Extraction Method Samples (37.5 g) were either vacuum-dried or used as received and were extracted using 300 mL of 200 proof ethanol in a pressure reactor (model 4842, Parr Instrument Co., Moline, IL). The extraction was performed at 130°C unless otherwise stated and stirred at 150 rpm for almost 1 hr. Approximately 10-15 min was needed for the reaction vessel to reach temperature. After reaching the set temperature, the substrate was extracted for an additional 30 mm. After 30 mm, the stirrer was turned off and the sample was allowed to settle to the bottom of the reactor for 15 mm. The reactor was equipped with a valve and a tube that extended about halfway down the reactor's vessel. This tube was fitted with a fine mesh screen to keep the extracted sample separate from the solution. Upon opening the valve, the difference in the pressure inside the vessel (70 psi) and the outside pressure allowed the liquid to he easily pumped into a beaker. The zein Vol. 83, No. 5, 2006 565