Separation and Purification of Molecular Species of Galactolipids by High Performance Liquid Chromatography

The NaCl concentration of the growth medium affected hydrogen production by Lyngbya sp. (No. 108) strain. Cells grown in medium containing 3% NaCl produced the most hydrogen. The carbohydrate content of this strain also increased with increasing NaCl concentration of the growth medium up to 720 fig/mg cells at 5 % NaCl. In the presence of 20 finlol/ml MFA (monofluoroacetic acid), inhibition of hydrogen production was observed. We extracted the glycogen from this nonheterocystous filamentous cyanobacterium, Lyngbya sp. (No. 108), and observed that glycogen and carbohydrate consumption of this strain is coincident with hydrogen production.

These results led us to the conclusion that the reserve glycogen or other carbohydrate were used as sources of electron donors for hydrogen production, and that the NaCl concentration of the medium affected the hydrogen production by this strain.     

Kinetic Properties of a Dipeptidase from Streptococcus cremoris

Some kinetic properties of a dipeptidase purified from a cell-free extract of Streptococcus cremoris H 61 were investigated. The Km values of this enzyme for various dipeptides were divided into 3 groups. Group 1 comprised mainly of neutral dipeptides, such as Leu-Gly, Leu-Leu and Leu-Ala, which had relatively low Km values (in the range 4.0-6.6 mm). Group 2 consisted of dipeptides with aromatic large amino acids either at the N- or C-terminal positions, like Leu-Phe, Phe-Ala and Leu-Tyr, which had very low Km values (in the range 1.0-2.4 mm). Group 3 was made up by dipeptides with acidic or basic amino acids at the N-terminals; His-Ala and Glu-Val were typical of this group. These had very high Km values (in the range 10–20 mm). Substantial substrate competition was found to exist in the presence of His-Ala. Bestatin inhibited the enzyme competitively with Leu-Gly and was found to have an apparent Ki value of 3.0 × 10^?8 m for the enzyme. Further, the enzyme was completely inhibited by EDTA at a concentration of 2.0 × 10^?5 m. On the other hand, once the activity was inhibited by EDTA, it could be restored by Co²⁺ and Zn²⁺ in the acidic pH side, and by Ca²⁺ and Mn²⁺ in the alkaline pH side.     

Effects of Lipids on the Rheological Properties of Aqueous Soybean Protein Dispersions

The flow properties of soybean protein–lipid–water suspension systems and coagulated gels were related to a protein–lipid interaction. For powdered soybean lecithin-added soybean protein suspension systems and their heat-induced gels, the yield stress (σ_y) and the consistency index (K) increased with increasing amounts of added lipid, but the flow behavior index (n) and the thixotropy index (TI) decreased. On the other hand, there were only small changes in the magnitudes of the thixotropic parameters and the viscometric parameters (σ_y, K and n) after adding soybean oil at various concentrations to soybean protein dispersions. These facts suggested that the formation of a protein–phospholipid complex increased the effective particle size, and that the intermolecular entanglements and linkages among the protein molecules or among the protein-phospholipid complexes were weakened by the addition of polar lipids. The thixotropy index defined in this study is available for characterizing the stress decay that occurs within soybean protein dispersions and heat-induced gels as they are sheared.     

Monohydroperoxides Formed by Autoxidation and Photosensitized Oxidation of Methyl Eicosapentaenoate

Methyl eicosapentaenoate (methyl 5,8,11,14,17-eicosapentaenoate) was subjected to autoxidation and methylene blue sensitized photooxidation. Methyl eicosapentaenoate monohydroperoxides, the primary products of the autoxidation and photosensitized oxidation, were isolated by silica gel column chromatography, and characterized by ultraviolet, infrared and nuclear magnetic resonance spectra. The isomeric composition of the monohydroperoxides were determined by gas chromatography-mass spectrometry as follows: the 5-, 8-, 9-, 11-, 12-, 14-, 15- and 18-isomers (autoxidation), and the 5-, 6-, 8-, 9-, 11-, 12-, 14-, 15-, 17- and 18-isomers (photosensitized oxidation). Methyl eicosapentaenoate was readily oxidized both by autoxidation and by photosensitized oxidation.     

Variations of Human Milk Protein Preparations from Individual Milk Samples

The acid coagulability of casein from 54 individual human milk samples and the variation of coagulability of their casein preparations by rennin were examined. The casein and whey protein preparations from individual human milk samples were also compared by polyacrylamide gel electrophoresis (PAE). Casein coagulated distinctly from 22 human milk samples when the pH was adjusted to 4.6 with acid, but it did not from other 32 samples. Twenty-two samples of casein preparations coagulated distinctly by rennin in the presence of calcium ions but 19 samples just became turbid. When the classification of human casein based on PAE pattern of major six bands was applied in our preparations, type A appeared most often and type C did least. Any regular relationship was not found between variation of the PAE pattern of casein preparations from individual human milk samples and that of acid coagulability or rennin coagulability.     

Reconstitution of Human Casein Micelle and Its Properties

Human casein micelles were reconstituted from isolated κ- and β-caseins and calcium ions. Micelle formation was recognized in the presence of calcium chloride even at the low concentration of 5mM. At pH levels ranging from 5.5 to 8.0, the re-formed micelles were quite stable so that precipitation of β-casein was not observed. The large micelles were constituted by a higher ratio of β-casein to κ-casein (16:1 by weight) than the small micelles (3: 1). The κ-casein in the small micelles contained carbohydrates to about 43% (w/w) in the molecule, whereas that in the large micelles was only about 25%. When the casein micelles were re-formed from κ-easein and fractionated β-casein components, the extent of phosphorylation of the β-casein component was found to influence the micelle formation; i.e., the β-casein component with no phosphate (the 0-P form) was disadvantageous to form micelles, but the component with 5 phosphates (the 5-P form) formed micelles most easily.     

Purification and Some Properties of α-Galactosidase from Tubers of Stachys affinis

An α-galactosidase from tubers of S. affinis was purified about 130 fold by ammonium sulfate fractionation, chromatography on DEAE-cellulose and gel filtration on Sephadex G-75. The purified enzyme showed a single protein band on disc gel electrophoresis. The molecular weight of the enzyme was determined to be approximately 42,000 by gel filtration and 44,000 by SDS disc gel electrophoresis. The optimum reaction pH was 5.2. The enzyme hydrolyzed raffinose more rapidly than planteose. The activation energy of raffinose and planteose by the enzyme was estimated to be 7.89 and 11.4 kcal/mol, respectively. The enzyme activity was inhibited by various galactosides and structural analogs of d-galactose. Besides hydrolytic activity, the enzyme also catalyzed the transfer reaction of d-galactosyl residue from raffinose to methanol.     

Studies on Molecular Properties of αs1-κ-Casein Complex by the Hydrodynamical Methods

Some molecular properties of α_s1-κ-casein complex, α_s1- and κ-casein polymers were examined by gel filtration, ultracentrifugation, and viscometry at pH 7.1. The Stoke’s radii of α_s1-κ-casein complex, α_s1- and κ-casein polymers were 99, 44 and 108 Å, respectively. The molecular weight of the above proteins were approximately 45 × 10⁴, 10 × 10⁴ and 80 × 10⁴, respectively. The stokes radius of α_s1-κ casein complex reduced compared with that of κ-casein polymer and the molecular weight of the complex was about half that of κ-casein polymer. These results suggest that κ-casein polymer dissociates into 4 smaller particles when α_s1-κ-casein complex is formed. The frictional coefficient and Scheraga-Mandelkern constants for each protein suggest that the molecular shape of α_s1-casein polymer is globular and that of α_s1-κ-casein complex and κ-casein polymer is rod-like.     

Similarity between γ-Casein and a Product of β-Casein Hydrolysis by Milk Protease

A γ-casein-like fraction (FIV) was isolated from the β-casein A hydrolyzate by milk protease and compared with γ-casein. The mobility in polyacrylamide gel electrophoresis, sedimentation coefficient, molecular weight, amino acid composition and terminal amino acids of FIV were almost coincided with those of γ-casein. It is suggested that γ-casein is possibly a product of β-casein hydrolysis by milk protease.     

Properties of Glycomacropeptide and Para-K-casein Derived from Human K-Casein and Comparison of Human and Bovine K-Caseins as to Susceptibility to Chymosin and Pepsin

The nutritional efficiency of quinolinic acid as a substitute for nicotinic acid was investigated using weanling rats Of the Sprague Dawley strain (3-weeks old) fed a nicotinic acid-free, tryptophan-limited diet containing various amounts of nicotinic acid or quinolinic acid. Judging from the growth response, food efficiency ratio, levels of NAD activity in the blood, liver, brain and upper small intestine, and urinary excretion of niacin we have concluded that exogenous quinolinic acid is approximately 1/9 as active as nicotinic acid. As many foods contain quinolinic acid, dietary quinolinic acid cannot be ignored from the standpoint of tryptophan and nicotinic acid replacement.