Toshiyasu Yamaguchi

Antihypertensive Effects of Hydrolysates of Wakame (Undaria pinnatifida) and Their Angiotensin-I-Converting Enzyme Inhibitory Activity

Aim: The angiotensin-I-converting enzyme (ACE) inhibitory and antihypertensive activities of wakame hydrolysates have been investigated in several studies. Methods: Wakame (Undaria pinnatifida) was hydrolyzed using 17 kinds of proteases and the inhibitory activity of the hydrolysates for ACE was measured. Of these hydrolysates 4 with potent ACE inhibitory activity were administered singly and orally to spontaneously hypertensive rats (SHR). Results: The systolic blood pressure of SHR decreased significantly after single oral administration of protease S ‘Amano’ and proleather FG-F hydrolysates (10 mg protein/kg body weight). In a long-term feeding experiment, 7-week-old SHR were fed standard chow supplemented with protease S ‘Amano’-derived wakame hydrolysates for 10 weeks. In SHR fed the 1 and 0.1% wakame hydrolysates, elevation of systolic blood pressure was still significantly suppressed for 7 weeks. Conclusions: The hydrolysates derived from wakame by protease S ‘Amano’ have a powerful ACE-inhibitory activity (IC50 = 86 µg protein/ml) and were effective in spite of their slight bitterness as ‘physiologically functional food’ with antihypertensive activity.

Effect of severe environmental thermal stress on redox state in salmon

Fish are exposed to many kinds of environmental stressors and the chances of succumbing to infectious diseases may be increased a result. For example, an acute increase in temperature can induce numerous physiological changes in the body. In the present study, we examined the redox state in response to a severe acute stress resulting from heat shock in teleost coho salmon (Oncorhynchus kisutch). The plasma lipid peroxides levels in fish gradually increased after heat shock treatment. By 2.5 h post-heat stress, plasma glutathione (GSH) levels had decreased, but they had returned to basal levels by 17.5 h post-stress. Plasma superoxide dismutase activities in stressed fish were significantly increased compared with those in control fish at 17.5 h post-stress, but had returned to basal levels by 48 h post-stress. Expression levels of hepatic GSH and heat shock protein 70 gradually increased after heat shock treatment. These results concerning the changing patterns of multiple important redox-related biomarkers suggest that severe thermal stressors can affect the redox state and induce oxidative stress in ectothermal animals, such as fish, in vivo. Hence, manipulation of appropriate thermal treatment may possibly be useful to control fish fitness.

Complementary DNA Cloning and Molecular Evolution of Opine Dehydrogenases in Some Marine Invertebrates

The complete complementary DNA sequences of genes presumably coding for opine dehydrogenases from Arabella iricolor (sandworm), Haliotis discus hannai (abalone), and Patinopecten yessoensis (scallop) were determined, and partial cDNA sequences were derived for Meretrix lusoria (Japanese hard clam) and Spisula sachalinensis (Sakhalin surf clam). The primers ODH-9F and ODH-11R proved useful for amplifying the sequences for opine dehydrogenases from the 4 mollusk species investigated in this study. The sequence of the sandworm was obtained using primers constructed from the amino acid sequence of tauropine dehydrogenase, the main opine dehydrogenase in A. iricolor. The complete cDNA sequence of A. iricolor, H. discus hannai, and P. yessoensis encode 397, 400, and 405 amino acids, respectively. All sequences were aligned and compared with published databank sequences of Loligo opalescens, Loligo vulgaris (squid), Sepia officinalis (cuttlefish), and Pecten maximus (scallop). As expected, a high level of homology was observed for the cDNA from closely related species, such as for cephalopods or scallops, whereas cDNA from the other species showed lower-level homologies. A similar trend was observed when the deduced amino acid sequences were compared. Furthermore, alignment of these sequences revealed some structural motifs that are possibly related to the binding sites of the substrates. The phylogenetic trees derived from the nucleotide and amino acid sequences were consistent with the classification of species resulting from classical taxonomic analyses.

A simple and rapid method for the analysis of fish histamine by paper electrophoresis

The described analytical method for histamine determination in fish and seafood consists of sample extraction, adsorption onto a paper disc, application of the paper disc onto electrophoresis paper, electrophoresis for only 10 min, drying, and color developing by Pauly’s reagent. Histamine can be satisfactorily detected and completely separated from histidine, carnosine and other Pauly reagent-positive compounds. This method does not require expensive instrumentation and any tedious pretreatment to eliminate potential interference by other imidazole compounds, such as histidine or carnosine. This method can be used to detect histamine in multiple fish and seafood samples simultaneously that contain as little as 15 p.p.m. histamine (1.5 mg/100 g).

Inhibitory Effects of Hot Water Extract of the Stevia Stem on the Contractile Response of the Smooth Muscle of the Guinea Pig Ileum

The effects of a hot water extract of the stem of Stevia rebaudiana on the smooth muscle of isolated guinea pig ileum were investigated. The butyl alcohol layer of the extract antagonized the contractions of the isolated guinea pig ileum induced by histamine (1×10−5 M) and acetylcholine (1×10−5 M) in a concentration-dependent manner. The butyl alcohol layer of the extract also showed inhibition of CaCl2 (1×10−3–3.8×10−1 M)-induced contractions. The antagonism of the extract was considered to be non-specific, but this action might be related to an influx of extracellular Ca2+. With column chromatography preparation, the active component was assumed to be as stevioside. The antagonistic effects exerted by the stem extract of Stevia rebaudiana contributed to the gastroprotective activity of the extract in animals fed dietary histamine.

Alanine racemase activity in the microalga thalassiosira sp.

In this paper, the authors report the detection of alanine racemase activity in the marine diatom Thalassiosira sp. Since the Thalassiosira sp. was cultured under germ-free conditions, it appeared that D-alanine was not derived from bacteria but was produced through catalysis by algal alanine racemase. The rate of conversion of L-alanine to D-alanine was approximately the same as that for the reverse reaction, and the enzyme catalyzed the equilibration of the D- and L-forms. The crude enzyme preparation obtained from the cells at the stationary phase of the growth cycle had an optimal pH of approximately 9.5. The Lineweaver—Burk analysis showed that the Km for D- and L-alanine was 16.5 mM and 29.4 mM, respectively. It appears that the enzyme is highly specific for D- or L-alanine because it does not catalyze the racemization of other amino acids. In addition, after gel filtration, the enzyme did not require exogenous pyridoxal 5′-phosphate (PLP) for its activity, however, the effects of several chemicals suggest that the enzyme may be PLP-dependent. The enzyme is more similar to that found in invertebrates when compared with that found in bacteria. This is the first report on the occurrence of alanine racemase activity in the microalga Thalassiosira sp.

Metabolism of exogenous histamine in rainbow trout (Oncorhynchus mykiss)

Information on the metabolism of exogenous histamine in fish is of much concern regarding the effect of dietary histamine on fish. Histamine catabolic enzymes, diamine oxidase and histamine N-methyl transferase were measured in the tissues of rainbow trout. Diamine oxidase was detected in the stomach, pylorus caeca and intestine. Histamine N-methyl transferase was detected only in the liver.A change in the contents of histamine and its metabolites was observed in the tissues of rainbow trout after oral administration of histamine. A large amount of imidazole acetic acid was observed in the serum, kidney, liver and muscle. On the other hand, 1-methyl histamine was detected only in the liver. Histamine and its metabolites, imidazole acetic acid and 1-methyl histamine were metabolized and diminished within 48 hr in all tissues.These results showed that histamine was metabolized by two metabolic routes in rainbow trout. One is the main pathway producing imidazole acetic acid by intestinal diamine oxidase and the other is the complementary pathway producing 1-methyl histamine by liver N-methyl-transferase.

Antioxidant activities of aqueous extract from Stevia rebaudiana stem waste to inhibit fish oil oxidation and identification of its phenolic compounds

We investigated the potential for exploiting Stevia rebaudiana stem (SRS) waste as a source of edible plant-based antioxidants finding for the first time that the hot water extract of SRS had significantly higher antioxidant activity against fish oil oxidation than that of the leaf, despite SRS extract having lower total phenolic content, DPPH radical scavenging activity and ORAC values. To locate the major antioxidant ingredients, SRS extract was fractionated using liquid chromatography. Five phenolic compounds (primary antioxidant components in activity-containing fractions) were identified by NMR and HR-ESI-MS: vanillic acid 4-O-β-d-glucopyranoside (1), protocatechuic acid (2), caffeic acid (3), chlorogenic acid (4) and cryptochlorogenic acid (5). Further analysis showed that, among compounds 2-5, protocatechuic acid had the highest capacity to inhibit peroxides formation, but exhibited the lowest antioxidant activities in DPPH and ORAC assays. These results indicate that SRS waste can be used as strong natural antioxidant materials in the food industry.

Detection of plasmalogen from plasma low density lipoprotein and high density lipoprotein in carp, Cyprinus carpio, and rainbow trout, Oncorhynchus mykiss.

The study revealed the presence of plasmalogens in the low density lipoprotein (LDL) and high density lipoprotein (HDL) of the fish. The composition of the plasmalogen in the carp plasma LDL phospholipids was 0.94 and 0.23% in the HDL; the LDL phospholipids in the rainbow trout were 0.44% and the HDL was 0.18%. Aldehydes from the plasmalogen were derivatized with dansylhydrazides and separated by high performance liquid chromatography (HPLC). Their presence was detected using a fluorescence detector. Hexadecanal (C16: 0), octadecanal (C18: 0) and octadecenal (C18: 1) were determined to be the major components in the carp and rainbow trout.

Effects of Thermal Stressors on Growth-Related Gene Expressions in Cultured Fish

Growth in fish is regulated by the growth hormone (GH)-growth hormone receptor (GHR)-insulin-like growth factor-1 (IGF-1) axis. However, the effect of severe acute stressors on the GH-IGF-1 axis in fish is not well understood. The present study determined the changes in mRNA expression of growth-related genes gh, ghr, and igf and the redox state in coho salmon (Oncorhynchus kisutch), in response to severe acute stress. Severe stress consisted of exposure to heat shock (adequate rearing temperature +11 °C for 2 h). The plasma expression patterns of redox state-related biomarkers, such as glutathione, lipid peroxides, and superoxide dismutase, in response to heat shock suggest that heat shock might induce oxidative stress in fish. After exposure to heat shock, ghr mRNA levels in the pituitary glands and liver increased, whereas levels decreased 48 h post-stress. Hepatic igf1 mRNA expression levels gradually decreased in response to the stressor. On the other hand, the pituitary gh mRNA expression did not change in response to the stressor. These findings showed that a heat shock-induced oxidative stress could affect the redox state and the expression of several growth-related genes in coho salmon. The results of this study also suggest that the expression of several growth-related genes in fish may be affected differently by the types and strength of stress.