The origin of lysine-containing proteins in opaque-2 maize endosperm.

The reduction of zein synthesis in the maize (Zea mays L.) opaque-2 mutant is associated with an increased percentage of lysine in the endosperm protein. When expressed on an endosperm basis, we found that W64A opaque-2 contains 490 micrograms of lysine compared with 350 micrograms in W64A normal. SDS-PAGE analysis of endosperm proteins indicated that several non-zein proteins are more abundant in the mutant than in normal genotype. To determine the subcellular origin of these proteins, we separated an endosperm homogenate from developing kernels by sucrose density gradient centrifugation and used marker enzyme assays and immunoblot analyses to identify cellular components. Amino acid analysis of proteins in the gradient fractions showed that the majority of the lysine occurs in soluble proteins at the top of the gradient. To identify these proteins, we prepared a complex antiserum against the entire soluble protein fraction and used it to immunoscreen an endosperm cDNA expression library. Sequence analysis of clones identified mRNAs involved in carbohydrate metabolism, amino acid biosynthesis, and protein synthesis. RNA dot blot hybridization analysis with these clones revealed significant variation in the levels of transcripts between normal and opaque-2 endosperm, but we identified several mRNAs that are elevated in opaque-2 and that may encode proteins responsible for the enhanced lysine content.

Isolation and characterization of microorganisms associated with the traditional sorghum fermentation for production of sudanese kisra.

Sorghum flour obtained from Sudan was mixed with water in a 1:2 (wt/vol) ratio and fermented at 30 degrees C for 24 h. The bacterial populations increased with fermentation time and reached a plateau at approximately 18 h. At the end of 24 h, sorghum batter pH had dropped from 5.95 to 3.95 and the batter had a lactic acid content of 0.80%. The microbial population during the 24 h of fermentation consisted of bacteria (Pediococcus pentosaceus, Lactobacillus confusus, Lactobacillus brevis, Lactobacillus sp., Erwinia ananas, Klebsiella pneumoniae, and Enterobacter cloacae), yeasts (Candida intermedia and Debaryomyces hansenii), and molds (Aspergillus sp., Penicillium sp., Fusarium sp., and Rhizopus sp.). P. pentosaceus was the dominant microorganism at the end of the 24-h fermentation. When three consecutive fermentations using an inoculum from the previous fermentation were carried out, the bacterial population increase plateaued at 9 h. The microbial populations in these fermentations were dominated by P. pentosaceus.

Carbohydrate composition and nutritional quality for rats of sorghum tô prepared from decorticated white and whole grain red flour.

The carbohydrate composition and nutritional quality of acid, neutral and alkali tô, prepared from a decorticated white (DW) flour containing no polyphenols or whole grain red (WGR) flour high in polyphenols, were studied. The diets were characterized with regard to non-starch polysaccharides, Klason lignin, resistant starch (RS) and amino acid content. The nutritional properties were studied in balance trials with rats. Digestible energy of DW flour was higher than that of WGR flour because of a lower dietary fiber (DF) content and a higher digestibility of DF. Recovery of cellulose in feces of rats fed diets containing DW flour was 45-59% and recovery of noncellulosic polysaccharides (NCP) was 17-31%. In rats fed diets derived from WGR flour, recoveries were 76-83 and 54-67% for cellulose and NCP, respectively. Cooking resulted in formation of appreciable amounts of RS. Twenty-three to fifty-six percent of the RS in DW tô and 59-74% of RS from WGR tô were recovered in feces. Endosperm protein kafirins formed complexes during cooking. The result was a lower true protein digestibility and higher biological value in tô than in flour. Amino acid data revealed that the unavailable kafirins serve as a nitrogen source for the hindgut microflora. A high affinity of dietary polyphenols for proline and glycine residues can be postulated from digestible amino acid data. The net effect was a change in the excretory route of nitrogen from urine to feces.

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.