Biochemical Conversion of Uridine Diphosphate Glucose to Uridine Diphosphate 3-Ketoglucose by Washing Cells of Agrobacterium tumefaciens

The relation between the rate of increase in nonprotein nitrogenous compounds (NPN) of rabbit muscle and muscle pH ranging from 5.9 to 7.2 was examined during the post-mortem storage. Muscle of a high ultimate pH was prepared by the injection of ICH₂COOH into the vein. The more the muscle pH kept away from 6.3, the more NPN increased. Therefore, it has been suggested that the post-mortem proteolysis is mainly attributed to the acid proteolytic system comprising cathepsins in muscles at a pH lower than 6.3 and to the neutral proteolytic system in muscles at a pH higher than 6.3.

The ratio of the increment of ninhydrin positive materials to that of Cu-Folin phenol reagent positive materials among NPN was relatively large in muscles at a high pH. This result has suggested that the neutral proteolytic system was more abound in exopeptidase activity than acid proteolytic system.     

Studies on the Browning of Kori-tofu

Kori-tofu was stored under RH 95% at 60°C and the changes of its nitrogenous compounds in the course of browning were examined. The water-soluble and 10% TCA-soluble fractions prepared from the ether extraction residue of Kori-tofu were analyzed. The nitrogen content of water-soluble fraction temporarily decreased and then increased gradually in the course of browning, while that of 10% TCA-soluble fraction increased gradually during the browning process. Amino acid analyses of 10% TCA-soluble fraction and its hydrolyzate revealed that no free amino acid was detected in unbrowned Kori-tofu, whereas in browned Kori-tofu occurred peptides, ammonia and free amino acids such as aspartic acid, serine, glutamic acid, glycine and alanine.

Free amino acids similar to those detected in browned Kori-tofu occurred in the model system consisting of lysozyme and methyl linoleate, crotonaldehyde, glyoxal, methyl-glyoxal and also the brown substance prepared from glycine and methyl linoleate during storage. The formation mechanism of free amino acids from proteins was discussed.     

Proteolytic activity of Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus in frozen-stored Kashkaval cheese

Proteolytic activity of Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus in Kashkaval cheeses of varying aging times, stored at −10 to −12°C for 12 months, was studied. It was established that the proteolysis of Kashkaval cheese induced by the starter culture was significantly delayed by freezing. The noncasein nitrogen (NCN/TN) and nonprotein nitrogen (NPN/TN) as a percentage of total nitrogen increased slightly during frozen storage of Kashkaval. It was found that NCN/TN and NPN/TN values increased to a larger extent in frozen-stored Kashkaval samples with shorter aging time. Enhanced proteolysis was observed during ripening of thawed Kashkaval cheese. There was greater accumulation of noncasein nitrogen in thawed Kashkaval samples compared to the control samples. The enhanced proteolysis during ripening of thawed Kashkaval cheese resulted in larger amounts of high and medium molecular weight peptides and lower amounts of low molecular weight peptides and free amino acids as compared to controls.     

Changes in Total Nitrogen and Free Amino Acids in Stem Cuttings of Mulberry (Morus alba L.)

Changes in total nitrogen and free amino acid contents in stemcuttings of Morus alba have been studied. The fresh and dryweights and total nitrogen amounts of the parent stems of cuttingsdecreased initially after cutting. Their increase follows theformation of main roots in cuttings, suggesting that, like carbohydrates,sugars and starch, stored nitrogenous substances are used forsprouting and rooting of cuttings. Amino acids found in stems,roots and shoots are those common in other higher plants withthe exception of pipecolic acid and 5-hydroxypipecolic acid.Significant changes in the levels of asparagine, proline, arginine,-aminobutyric acid and alanine in roots, bark and wood of parentstems were observed during cutting growth, whereas those ofother amino acids remained comparatively constant; the mostpredominant amino acid in the starting materials was proline.while that in the cuttings during growth was asparagine. Theresults suggest that, among free amino acids, asparagine, prolineand arginine play the major part in storage of nitrogen in mulberry.The importance of glut-amine and asparagine in nitrogen metabolismin mulberry has been discussed.     

Some evidence for the possible presence of peptides in two chondrites by use of a sequencing procedure

In hot water extracts of samples from interior of two ordinary chondrites (Goumoshnik and Pavel) after acid hydrolysis using diphenylindonyl-isothiocyanate (DIITC) and fluorescent detection (FD) of the obtained diphenylindonyl thiohydantoins (ITH) 10 to 13 protein and nonprotein amino acids were found. In the same manner the presence of compounds containing amino groups but not of free amino acids was established in nonhydrolysed extracts. Sequencing was performed on the amino compounds using DIITC and FD of the cleaved ITH up to 9 steps with extracts of Goumoshnik and up to 14 with those of Pavel. At each step 1 to 6 different protein and nonprotein amino acids were liberated. The presence of nonprotein amino acids supports the view that the peptides in meteorites are indigenous.     

The effect of nitrogen status on the regulation of plant-mediated proteolysis in ingested forage; an assessment using non-nodulating white clover

Production of nitrogenous waste by livestock agriculture is a significant environmental concern in terms of pollution of land and water. In the rumens of cattle and sheep, the excessive proteolysis which contributes to inefficiency of nutrient use involves both the rumen microbial population and the intrinsic plant proteases that can mediate protein degradation in ingested fresh forage on exposure to the environmental stresses of the rumen. Here, white clover (Trifolium repens) plants that do not form root nodules, and so are dependent on nitrate supplied to the roots, have been used to determine how nitrogen status of the plant affects the rate of plant‐mediated proteolysis in forage under conditions that simulate ingestion by grazing ruminants. Plants were grown from seed and supplied with nutrient solution containing 2.5, 5.0, 7.5 or 10 mM nitrate. Protein, free amino acid and protease activity were determined in leaves which had been placed in an in vitro system designed to simulate conditions experienced in the rumen (anaerobic phosphate buffer maintained at 39°C in the dark). Foliar protein content increased with increasing nitrate supply, while in vitro incubation of leaves resulted in time‐dependent decreases in protein concentration and increases in amino acid concentration. Regardless of nitrate supply, 50% of the protein was degraded in 6 h and 80% after 24 h. As the extent of protein decrease was determined by initial protein content, more protein degradation occurred in those plants grown with the highest nitrate supply: after 6 h, 130.7 mg g^?1 dry matter (DM) was degraded in leaves grown at 10 mM nitrate but only 52.3 mg g^?1 DM in leaves grown at 2.5 mM nitrate. Hence, although the percentage of proteolysis is independent of foliar protein concentration, the latter is critical to the quantity of protein degraded. Heat‐stable serine and cysteine proteases were active throughout the term of the in vitro incubation. Although proteolysis in ingested forage can continue for many hours, mediated by heat‐stable proteases, maximum amino acid accumulation accounted for less than 40% of initial protein. Therefore, it is proposed that continued and extensive proteolysis occurs following leaf excision and exposure to rumen conditions because amino acid accumulation is insufficient to initiate those feedback systems which sense cytoplasmic amino acid concentration and prevent excessive proteolysis during normal source–sink relations.     

Studies on Changes in Stored Shell Eggs

Being solubilized by treatment with 0.01 m mercaptoethanol, the ovomucin gel(B) was found in free boundary electrophoresis to contain subunits which were consisted of two components. Changes in the physicochemical properties of all the insoluble ovomucin gel(B) and sol(B) obtained from stored egg white were studied after this treatment.

The fast moving component of the ovomucin gel(B) in free boundary electrophoresis decreased during storage and disappeared completely after 30 days. On the other hand, the fast moving component of the ovomucin sol(B) increased during storage.

The acid mucoprotein concentration of the ovomucin gel(B) in acrylamide gel electrophoresis decreased and that of the ovomucin sol(B) increased during storage, although the protein pattern did not show significant changes.

The interaction of the ovomucin gel(B) with lysozyme decreased whereas that of the ovomucin sol(B) increased during storage.

By summarizing these results, a model of ovomucin gel structure and a mechanism of egg white thinning were proposed.     

Manganese deficiency leads to elevated amino acid pools in citric acid accumulating Aspergillus niger

Free amino acid pools have been investigated in a citric acid accumulating strain of Aspergillus niger during batch growth under manganese sufficient and deficient conditions by means of an improved chromatographic method. Studies on the mycelial content of several nitrogenous compounds under manganese sufficient and deficient conditions showed that manganese deficiency resulted in lower amino acid pool sizes during trophophase and considerable accumulation during idiophase, and in a reduction of the protein and nucleic acid contents. Addition of cycloheximide to mycelia grown with sufficient manganese also caused an elevation of free amino acid pool sizes, thus indicating that impairment of protein synthesis by manganese deficiency is responsible for the observed rise in amino acid concentration. Furthermore it was observed that the manganese deficient mycelia excreted high amounts of all amino acids suggesting that manganese deficiency may also affect membrane permeability.     

Changes in growth,proteins and free amino acids of developing seed and pod of fenugreek

The growth and development under field conditions of the pods and seeds of two cvs of Trigonella foenum graecum are described. Samples were harvested at different stages of ripeness for the determination of dry matter, protein and free amino acid content. During maturation, reserves of solutes are established in the pod wall before the seeds begin their exponential phase of growth. Later, these reserves disappear, providing about 20% of the seed's requirements for nitrogen. SDS-electrophoresis was used to follow the formation of proteins and it was shown that the synthesis of storage proteins takes place prior to dehydration of the seed. Production soluble nitrogenous compounds precedes protein accumulation. Free amino acids follow the same pattern. 4-Hydroxyisoleucine represents nearly 80% of free amino acid of dry seeds. The concentration does not decrease in the later stages of maturation of the seed but this unusual amino acid is absent in the storage proteins of the seeds.     

Studies on the Nitrogen Metabolism in Tobacco Plants. A.

The nitrogenous compounds of tobacco saps have been studied both qualitatively and quantitatively and the following results were obtained.

(1) Nitrate nitrogen accounts for 40 to 70% of the total nitrogen and the rest is composed mostly of amino and alkaloid nitrogen.

(2) Amides and basic amino acids compose a large part of the amino and amide nitrogen. Among the amino acids and amides of the tobacco saps glutamine is the highest in the content and asparagine, lysine, leucine and serine follow glutamine.

(3) Topping procedure increased remarkably the alkaloid contents in the sap but decreased the amino acid nitrogen as compared with those of the untopped plant sap.     

Nitrogenous compounds stimulate glucose‐derived acid production by oral Streptococcus and Actinomyces

Both Streptococcus and Actinomyces can produce acids from dietary sugars and are frequently found in caries lesions. In the oral cavity, nitrogenous compounds, such as peptides and amino acids, are provided continuously by saliva and crevicular gingival fluid. Given that these bacteria can also utilize nitrogen compounds for their growth, it was hypothesized that nitrogenous compounds may influence their acid production; however, no previous studies have examined this topic. Therefore, the present study aimed to assess the effects of nitrogenous compounds (tryptone and glutamate) on glucose‐derived acid production by Streptococcus and Actinomyces. Acid production was evaluated using a pH‐stat method under anaerobic conditions, whereas the amounts of metabolic end‐products were quantified using high performance liquid chromatography. Tryptone enhanced glucose‐derived acid production by up to 2.68‐fold, whereas glutamate enhanced Streptococcus species only. However, neither tryptone nor glutamate altered the end‐product profiles, indicating that the nitrogenous compounds stimulate the whole metabolic pathways involving in acid production from glucose, but are not actively metabolized, nor do they alter metabolic pathways. These results suggest that nitrogenous compounds in the oral cavity promote acid production by Streptococcus and Actinomyces in vivo.     

Variations in the content of free and bound amino acids during dormancy and the first cell cycle in Helianthus tuberosus tuber explants

Abstract

Qualitative and quantitative analysis of free and bound amino acids and amides during dormancy and the most important phases of the first cell cycle was carried out in tubers of Helianthus tuberosus.

In the dormant tuber arginine was confirmed to be the most abundant amino acid. A high amount of asparagine was also present; on the contrary glutamine was found in very low concentrations. During the progression of dormancy, all the free amino acids and amides declined while aspartic and glutamic acid increased.

During the G₁ phase of the first cell cycle induced by 2,4-D, all the free amino acids and amides decreased with the exception of glutamic acid.

At 18, 20, 24 h of activation with 2,4-D, corresponding to the S phase and the beginning of mitosis, bound amino acids were also determined. In these phases of the cell cycle they increased reaching a maximum at 20 h; on the other hand the free amino acid and amide content, especially aspartic acid, asparagine and arginine, decreased with the exception of glutamic acid, alanine and phenylalanine.     

Studies on Roasting Changes of Proteins

Changes of casein and lysozyme during roasting at various times and temperatures 100 ~ 300°C), especially those in amino acid composition and formation of some nonvolatile degradation products, were investigated.

Tryptophan, methionine, basic amino acids and β-hydroxy amino acids were easily decomposed as compared with acidic amino acids and other neutral amino acids in casein and Iysozyme. In hot water extract of roasted casein were detected some free amino acids, peptides, organic acids such as α-ketoglutaric, tartaric and malic acids, and indole. It is considered that free amino acids are produced mainly through ionic cleavage of peptide bond with the water bound within casein.     

The role of alanine synthesis and nitrate-induced nitric oxide production during hypoxia stress in Cucurbita pepo nectaries

Floral nectar is a sugary solution produced by nectaries to attract and reward pollinators. Nectar metabolites, such as sugars, are synthesized within the nectary during secretion from both pre-stored and direct phloem-derived precursors. In addition to sugars, nectars contain nitrogenous compounds such as amino acids; however, little is known about the role(s) of nitrogen (N) compounds in nectary function. In this study, we investigated N metabolism in Cucurbita pepo (squash) floral nectaries in order to understand how various N-containing compounds are produced and determine the role of N metabolism in nectar secretion. The expression and activity of key enzymes involved in primary N assimilation, including nitrate reductase (NR) and alanine aminotransferase (AlaAT), were induced during secretion in C. pepo nectaries. Alanine (Ala) accumulated to about 35% of total amino acids in nectaries and nectar during peak secretion; however, alteration of vascular nitrate supply had no impact on Ala accumulation during secretion, suggesting that nectar(y) amino acids are produced by precursors other than nitrate. In addition, nitric oxide (NO) is produced from nitrate and nitrite, at least partially by NR, in nectaries and nectar. Hypoxia-related processes are induced in nectaries during secretion, including lactic acid and ethanolic fermentation. Finally, treatments that alter nitrate supply affect levels of hypoxic metabolites, nectar volume and nectar sugar composition. The induction of N metabolism in C. pepo nectaries thus plays an important role in the synthesis and secretion of nectar sugar.     

Effect of Gamma Irradiation on the Amino Acid of Potatoes

The free amino acids of potatoes irradiated with the doses of 7,000, 15,000 and 30,000 rad were determined by ion-exchange chromatography.

After 15 days storage following irradiation, it was shown that the concentration of asparatic acid, proline and aliphatic amino acids increased with increasing irradiation doses, while that of basic amino acids and glutamic acid especially decreased. However, after 105 days of storage, the similarity of the free amino acid content of irradiated potatoes to that of non-irradiated and non-stored potatoes was observed.

On the concentration of protein-bound amino acids, there were no significant differences between non-irradiated and 15,000 rad irradiated potatoes.     

Effect of the parasitoid,Cardiochiles nigriceps,on the nitrogen excretion of its host,Heliothis virescens

Excretion of nitrogenous products by larvae of the tobacco budworm, Heliothis virescens, and the effect of parasitism by the parasitoid, Cardiochiles nigriceps, on the host's excretory products were investigated. The dry weight of faeces produced after parasitism declined although the per cent of total nitrogen in the faeces did not differ significantly between non-parasitized and parasitized larvae. No free ammonia and only small amounts of ammonium nitrogen were found in the faeces. Uric acid was the only purine found and parasitism significantly decreased its excretion. Parasitism apparently had no qualitative effect on the presence of amino acids in the faeces.Amino nitrogen was the major contributor of faecal nitrogen and was reduced in parasitized larvae. Parasitism significantly increased the excretion of proteins in H. virescens larvae. One pteridine was identified in H. virescens and parasitism had no qualitative effect on the pteridines found in host faeces.The major nitrogenous compounds in the faecal material from the parasitoid were uric acid, amino nitrogen, and protein. In addition several pteridines were identified.     

Permeability Characteristics and Amino Acid Incorporation during Senescence (Ripening) of Banana Tissue

Changes in free space of banana tissue during ripening were measured using radioisotopes. Free space increased significantly about 44 hours before the onset of, and rose exponentially during the respiratory climacteric. The increase in free space indicates a progressive increase in the proportion of cells which becomes completely permeable to solutes in the ambient solution by simple diffusion. At the respiratory peak the tissue was essentially 100% free space to mannitol, sucrose, fructose and chloride.

The capacity for active uptake of solutes declined about one day before the onset of the respiratory rise and fell to a very low level by the respiratory peak.

There was no change in the level of protein or amino acids during ripening. Assays of tissue sections before and after washing indicated an increased rate of leakage of amino acids during ripening.

Studies of incorporation of 3 concentrations of ¹⁴C-labeled leucine and phenylalanine indicated marked changes in the size and specific activity of the amino acid pool at the site of protein synthesis just prior to and during the climacteric rise, due to a diffusive mixing of the labeled substrate with the previously sequestered endogenous, unlabeled pool of substrate. The use of a high concentration of exogenous substrate (above saturation for uptake) resulted in an apparent constant specific activity of the metabolic pool through the ripening period. Data from these studies indicated a decline in amino acid incorporation during the climacteric.

It was concluded that the initiation of permeability changes marks the onset of senescence in banana. The causative relations between alterations in permeability, the respiratory rise and other chemical changes attending fruit ripening are discussed.

     

Muramidase Catalyzed Hydrolysis of p-Nitrophenylacetate

It was found that muramidase can catalyze the hydrolysis of p-nitrophenylacetate (NPA) producing p-nitrophenol and acetic acid. The activity of muramidase for NPA, however, simply increased on raising a temperature and with an increase in alkalinity of reaction mixture. The mechanism of muramidase catalyzed hydrolysis of NPA differs from that of chymotrypsin which can catalyze burstly the hydrolysis of NPA by its histsdine residue.

The amount of reducing power produced owing to the hydrolysis of glycol chitin by muramidase was not affected by the presence of NPA, and inversely, the hydrolysis of NPA was not affected by glycol chitin. Obviously, there was no competitive inhibition between NPA and glycol chitin. The responses of modified muramidase to glycol chitin and to NPA did not correspond at all. The catalytic site of muramidase for glycol chitin may be different from that of muramidase for NPA. The hydrolysis of NPA is catalyzed by some free amino groups in the muramidase molecule, while the catalytic site for glycol chitin is not known.     

14C Photoassimilate Partitioning in Developing Sunflower Seeds

Changes with age of protein and oil content in field-grown Helianthusannuus seeds were followed during the grain filling period.Seeds were sampled from different zones on the flower head:peripheral, intermediate, and central. Regardless of seed position,at maturity protein and oil content accounted for approximately18% and 50% of the dry weight respectively. In an attempt todetermine the importance of the role of photosynthesis in grain-filling,¹⁴CO₂ was incorporated into sunflower leaves and translocatedradioactive photoassimilates in the seed were studied. ¹⁴C-labelledproducts including carbohydrates, nitrogenous compounds andlipids were determined as a function of seed position, lengthof chase period, and seed age. Within 8 h, ¹⁴C-labelled photosynthateis detected in the seed, up to 80% of seed-incorporated radioactivitybeing in the form of free sugars and organic acids. The conversionrate from free sugars to storage compounds (lipids, proteinsand starch) varies according to seed position and age. Lipidsconstitute the major photosynthetic sink, reaching levels ofgreater than 80% of the total seed-incorporated radioactivity.The effects of abscisic acid on uptake and partitioning of ¹⁴Csucrose into immature excised cotyledons were also studied. Key words: Sunflower, photosynthate, abscisic acid     

Volatile and Non-volatile Forms of Aroma Compounds in Tea Leaves and Their Changes due to Injury

Fresh tea leaves were homogenized in a chloroform-methanol mixture (1:1, v/v), and separated into chloroform-soluble and methanol-water-soluble fractions after addition of water. From the chloroform-soluble fraction, the volatile forms of the aroma compounds were obtained. The non-volatile forms of the aroma compounds were associated with the methanol-water-soluble fraction, and were converted to volatile forms by hydrolysis with dilute acid.

The amount of the aroma compounds in the free form, such as cis-3-pentenol, hexanol, cis-3-hexenol, trans-2-hexenol, linalool, linalool oxide (cis, 5-membered), linalool oxide (trans, 5-membered), linalool oxide (trans, 6-rnembered), linalool oxide (cis, 6-membered), nerol, geraniol, phenylmethanol, and 2-phenylethanol, markedly increased during black tea manufacture. However, those in the bound form, showed a slight decrease during the manufacture. The increases in the former were also brought about by maceration, or treatment of the tea leaves with monoiodoacetate or malonate.