In vitro digestibility and amino acid composition of pearl millet (Pennisetum typhoides) and other cereals.

The purpose of this study was to compare in vitro digestibility, protein distribution patterns, and amino acid composition of pearl millet with other major cereals. Digestibility values for the pearl millet varieties were higher than that of sorghum and comparable to that of maize. In contrast to sorghum, digestibility of pearl millet and maize did not decrease significantly upon cooking. Protein distribution patterns of uncooked pearl millet and shifts in the different fractions as a result of cooking also resembled that of maize and not sorghum. The amino acid profile of pearl millet is more favorable than that of normal sorghum and normal maize and is comparable to the small grains, wheat, barley, and rice. On the basis of these findings, it appears that pearl millet is a nutritious and well-digested source of calories and proteins for humans.

Improving the in vitro protein digestibility of sorghum with reducing agents.

We have shown in previous reports that cooked sorghum protein is less digestible than other cooked cereal proteins. The pepsin-indigestible proteins in sorghum were found to be mainly prolamin proteins. Cooking sorghum in the presence of 2-mercaptoethanol increased protein digestibility (in vitro with pepsin or trypsin/chymotrypsin) to a level comparable with other cereals. At a concentration of 100 mM, other reducing agents (dithiothreitol, sodium bisulfite, and L-cysteine) were equally effective in improving sorghum digestibility. When maize was cooked in the presence of 2-mercaptoethanol, protein digestibility increased 5% compared to 25% for sorghum. Cooking barley, rice, and wheat with 2-mercaptoethanol had no significant effect on protein digestibility. The addition of reducing agents appears to prevent the formation of protein polymers linked by disulfide bonds.

Digestibility and utilization of protein and energy from Nasha, a traditional Sudanese fermented sorghum weaning food.

Whole grain sorghum flour was fermented into Nasha, a traditional Sudanese food, and freeze-dried or drum-dried. It was cooked and fed to convalescent malnourished infants and small children as 61% of total diet calories and all of 6.4% protein calories, with (Lys+) and without lysine supplementation to 3% of protein. Apparent absorptions of nitrogen were 73 +/- 5 and 74 +/- 6% of intake, significantly (P less than 0.01) less than those from preceding (Cas-1, 86 +/- 3%) and following (Cas-2, 85 +/- 3%) isonitrogenous casein diets. Apparent retentions of nitrogen from Nasha (26 +/- 10%) were significantly lower than those from Lys + (34 +/- 9%, P less than 0.05), Cas-1 (35 +/- 11%, P less than 0.01) or Cas-2 (49 +/- 9%, P less than 0.01). Retentions from Cas-2 were higher than those from Cas-1 or Lys + (P less than 0.01). Fecal wet and dry weights were higher (P less than 0.02) during both Nasha diets and Cas-2 than during Cas-1. Fecal energy and carbohydrate were significantly (P less than 0.01) higher from either Nasha diet than from either casein diet; fecal fat was not different. Two children received drum-dried Nasha without further cooking; digestibilities were not different from those of the cooked product but biological value was much lower. When properly cooked and consumed along with small amounts of a good source of lysine, Nasha is a satisfactory weaning food.

Pepsin digestibility of proteins in sorghum and other major cereals.

We have shown previously that sorghum is highly digestible in the rat. However, other workers have shown that sorghum is much less digestible than wheat, maize, and rice in young children. Because the rat does not show these digestibility differences, we developed an empirical pepsin digestion method, first reported in 1981, which simulates the digestion values found in children. In this report the method has been improved and used to analyze wheat, maize, rice, millet, and sorghum and certain processed samples of millet and sorghum. The pepsin digestion values parallel those found in children for wheat, maize, rice, and sorghum. In addition, a processed sorghum product that gave a high digestion value in children also gave a high value with the in vitro pepsin method.

Digestibility of sorghum proteins.

Published information indicates that rice, maize, and wheat proteins are much more digestible in children than sorghum proteins are (66-81% compared with 46%). However, this digestibility difference cannot be demonstrated with the weanling rat, which gave digestibility values of 80% for cooked and 85% for uncooked sorghum gruels. Therefore, a search was made for a laboratory system sensitive to the digestibility differences between sorghum and other cereals. We found that porcine pepsin in vitro shows these digestibility differences. Using pepsin, we have found that uncooked sorghum proteins have a high digestibility (78-100%), which drops to a range of 45-55% after cooking. Two fermented sheet-baked sorghum products (kisra and abrey) from Sudan gave pepsin digestibility values of 65-86%. In contrast, unfermented cooked gruels made in our laboratory from the same flours used for the kisra and abrey gave pepsin values of only 44-56%. Therefore, fermentation improves pepsin digestibility of sorghum. The digestibility values of other sorghum-based foods prepared in the semiarid tropics need surveying. Those with high pepsin digestibility values hopefully should be more digestible (in children) than are the cooked sorghum gruels studied to date by human nutritionists.

Plasmin-mediated proteolysis of casein in bovine milk.

Plasminogen was found to be present in bovine milk by crossreactivity between rabbit antiserum to plasminogen and casein prepared from milk by acid precipitation. This result was further supported by recovery of intact 125I-labeled plasminogen from rabbit milk after its intravenous injection. Freshly isolated whole bovine casein was observed to undergo slow autoproteolysis at 37 degrees C. Polyacrylamide gel electrophoresis revealed gradual disappearance of major caseins accompanied by appearance and increase in intensity of numerous electrophoretic bands. This autoproteolysis was inhibited by low concentrations of epsilon-aminocaproic acid (0.1 mM) and diisopropyl fluorophosphate (1 mM); catalytic amounts of urokinase accelerated the process. Autoproteolysis of isolated bovine beta-casein was shown by both urea and sodium dodecyl sulfate gel electrophoresis to result in formation of gamma 1- and gamma 2-caseins. Similar electrophoretic bands were formed when beta-casein was degraded by plasmin prepared from bovine blood serum. These results support the hypothesis that bovine plasmin occurs in milk and is identical to alkaline milk protease.

Methods for improving cereal protein quality.

Three methods for improving cereal protein quality are discussed. Two older methods are supplementation with limiting essential amino acids and with protein concentrates high in those amino acids. The most recent method (since 1964) is the replacement of the normal cereal grain with its high lysine mutant counterpart. Three high lysine cereals are now available, corn, barley, and sorghum. In animal feeding, least cost formulas will determine which of the three improvement methods will be used. In human nutrition, cost, availability, palatability and acceptance are all equally important factors. In animals, pounds of gain per pound of feed will be the final measure of cereal protein quality. In humans, especially preschool children, the most important criterion will be the ability of the improved cereal protein to build a strong immune defense system. Animal studies show that protein quality is more important than calories when calories are restricted to less than ad libitum consumption. It is therefore essential that children restricted in their total energy intake have the best cereal protein quality possible to protect their immune system.

Kinetic evidence for an intermediate in the deacetylation of monoacetyl-chymotrypsin.

Mono[14C]acetyl-chymotrypsin was prepared by treating alpha-chymotrypsin with a 10-fold molar excess of p-nitrophenyl[14C]acetate at pH 5, and the acetylated enzyme was isolated free of excess reagents by gel filtration. Deacetylation at pH 6.0 was followed by observing the decrease in acid-precipitable radioactivity and provided a first-order rate constant of 0.02 +/- 0.008 min-1. Reactivation of the acetylated protein was followed by continuously monitoring the appearance of esterolytic activity towards alpha-N-acetyltyrosine ethyl ester. Reactivation at pH 6.0 occurred exponentially with a first-order rate constant of 0.2 +/- 0.015 min-1, the reactivated enzyme exhibiting an apparent catalytic contant (k' cat) of 1200 +/- 60 min-1, which decreased to a value of 945 +/- 15 min-1 by an apparent first-order process with a rate constant of 0.025 +/- 0.006 min-1. These results are interpreted in terms of a two-step deacetylation of monoacetyl-chymotrypsin involving an acetylated intermediate with esterase activity.

Protein quality of two varieties of high-lysine maize fed alone and with black beans or milk to normal and depleted weanling rats.

Opaque-2 corn, modified high-lysine corn with vitreous endosperm, and common corn were compared in rat growth studies. Corn alone supplied 8.6% protein; and at both 8.6 and 11.4% protein, corn furnished 60% of the protein and either milk or black beans supplied 40%. The modified corn, which also was high in both lysine and tryptophan, did not differ significantly from Opaque-2 corn in its ability to stimulate growth and nitrogen deposition in both well-nourished and partially protein-depleted rats; and both were significantly superior to common corn when fed alone. Milk was a more effective supplement for corn than black beans, especially when high-lysine varieties were fed, whereas beans exerted a similar effect on all varieties of corn. The depleted rats deposited more nitrogen as a result of eating most diets than did well nourished rats. Implications of these findings for preschool children are discussed.

Endosperm protein synthesis in maize mutants with increased lysine content.

The endosperm proteins of the maize mutants, opaque-2, opaque-7, floury-2, brittle-2, and the double mutant of opaque-2 and brittle-2, were separated into five soluble fractions by the Landry-Moureaux method. As compared to their isogenic normal counterparts, the mutant endosperms had higher concentrations of albumins, globulins, and glutelin-3, and lower concentrations of prolamines. The combination of the opaque-2 and brittle-2 genes enhanced this difference. Although the four mutant genes are located on three different chromosomes, they exert a similar effect on endosperm protein composition. Five other starchmodifying mutants with high lysine content resemble the brittle-2 mutant in endosperm protein composition, when the gene is present either singly or combined with opaque-2. Therefore, the pattern of protein synthesis in all maize mutants with high lysine concentrations may be either identical or very similar. Because no synergistic effect on lysine concentration is obtained when floury-2 is combined with opaque-2, different pathways leading to reduced zein synthesis may exist in the floury and starch-modifying mutants with high lysine concentrations.