Characterization and functional properties of digestive proteases in two sparids; gilthead seabream (Sparus aurata) and common dentex (Dentex dentex)

Digestive proteases present in two sparids, seabream (Sparus aurata) and common dentex (Dentex dentex), have been characterized using both biochemical and electrophoretic techniques. Although optimum pH and temperature for maximum activity of both acid and alkaline proteases were similar in the two species, important differences in total activity, as well as in thermal and pH stability were found. Specific inhibitors and SDS-PAGE zymograms were used to clarify such differences. Evidences support the existence of a more active and complex protease set in dentex. Results are discussed from the perspective of their application to the formulation of feeds for each species.     

In vitro evaluation of the effect of a high plant protein diet and nucleotide supplementation on intestinal integrity in meagre (Argyrosomus regius)

In the present study, the protective effect of nucleotides over damages induced by the consumption of a diet containing a high amount of vegetable ingredients (560 g kg^?1) in the intestinal epithelia of the meagre (Argyrosomus regius) was assessed by assays performed with an Ussing-type chamber. Two experimental feeds were prepared including or not a commercial mixture of nucleotides (1 g kg^?1). Nucleotides significantly enhanced fish growth during the experiment. On the other hand, differences in the integrity and functionality of intestinal epithelia were evidenced by a change in the polarity of intestinal trans-epithelial potential. Samples of fish fed on the control diet showed a preferentially secretory short-circuit current, while those of fish receiving the nucleotide-supplemented diet showed a significantly lower and preferentially absorptive negative current. It is concluded that alterations of intestinal physiology juvenile meagre resulting from the intake of high amounts of plant ingredients could be minimized by nucleotide supplementation.     

Development of digestive enzymes in larvae of Mayan cichlid Cichlasoma urophthalmus

The development of digestive enzymes during the early ontogeny of the Mayan cichlid (Cichlasoma urophthalmus) was studied using biochemical and electrophoretic techniques. From yolk absorption (6?days after hatching: dah), larvae were fed Artemia nauplii until 15?dah, afterward they were fed with commercial microparticulated trout food (45% protein and 16% lipids) from 16 to 60?dah. Several samples were collected including yolk-sac larvae (considered as day 1 after hatching) and specimens up to 60?dah. Most digestive enzymes were present from yolk absorption (5?C6?dah), except for the specific acid proteases activity (pepsin-like), which increase rapidly from 8?dah up to 20?dah. Three alkaline proteases isoforms (24.0, 24.8, 84.5?kDa) were detected at 8?dah using SDS?CPAGE zymogram, corresponding to trypsin, chymotrypsin and probably leucine aminopeptidase enzymes, and only one isoform was detected (relative electromobility, Rf?=?0.54) for acid proteases (pepsin-like) from 3?dah onwards using PAGE zymogram. We concluded that C. urophthamus is a precocious fish with a great capacity to digest all kinds of food items, including artificial diets provided from 13?dah.     

Differences in the digestive biochemistry between the intertidal clam, Ruditapes decussatus, and the subtidal clam, Venerupis pullastra

The present study was aimed to evaluate differences in physicochemical characteristics (pH and temperature optima) of the main digestive enzymes (amylase, cellulase, laminarinase, and protease) in two commercially important bivalves, the intertidal clam, Ruditapes decussatus, and the subtidal clam, Venerupis pullastra, as well to assess individual variability of such activities. The assays were performed on extracts obtained from the digestive glands and crystalline styles of individuals collected in the wild and laboratory-conditioned during 1 week. In both species, maximum activity for amylase and cellulase was measured at pH 6 and that of laminarinase at pH 5.5, while pH profiles of protease activity showed important differences between them. Temperature profiles also showed interspecific variations, mainly in laminarinase. Carbohydrase activities were comparatively much higher in R. decussatus and proteolytic activities in V. pullastra. The observed differences both in physicochemical characteristics and relative importance of digestive enzymes are discussed in relation to the amount and type of food available in their respective habitats.     

Multi-atom resonant photoemission: A method for determining near-neighbor atomic identities and bonding

Measurements and theoretical calculations are reported for an interatomic multi-atom resonant photoemission (MARPE) effect that permits direct determination of near-neighbor atomic identities (atomic numbers). MARPE occurs when the photon energy is tuned to a core-level absorption edge of an atom neighboring the emitting atom, with the emitting level having a lower binding energy than the resonant level. Large peak-intensity enhancements of 33 to 105 percent and energy-integrated effects of 11 to 29 percent were seen in three metal oxides. MARPE should also be sensitive to bond distance, bonding type, and magnetic order, and be observable via the secondary processes of x-ray fluorescence and Auger decay.     

Improvement of the desorption of the pesticide 2,4‐D via complexation with HP‐β‐cyclodextrin

Inclusion complex formation of 2,4‐dichlorophenoxyacetic acid (2,4‐D) with hydroxypropyl‐β‐cyclodextrin (HP‐β‐CD) has been proposed as a way of modifying the behaviour of the pesticide in the soil environment. The present study assesses the effect of complex formation on 2,4‐D physicochemical properties (aqueous solubility, crystallinity and dissolution rate) and its behaviour on soils. The solid complexes were prepared using different methods (spray drying, kneading and heating in a sealed container). To confirm the complex formation in the solid state differential scanning calorimetry, hot stage microscopy and x‐ray diffraction techniques were employed. Complex formation in solution was studied by phase solubility. The presence of HP‐β‐CD increased the 2,4‐D solubility nine times approximately. The apparent stability constant was determined as 98.6 M ⁻¹. The dissolution rates of the 2,4‐D/HP‐β‐CD complexes were examined and compared with that of the pure pesticide. The results indicated that the complex may have great utility as a rapid way of dissolving the pesticide. Batch experiments were performed to study the adsorption–desorption of 2,4‐D on soils and the influence of the HP‐β‐CD over these processes. The results showed that HP‐β‐CD could increase the desorption of 2,4‐D previously adsorbed on soils. © 2000 Society of Chemical Industry     

Two Unusual Cases of Canine Prostatitis: Prostatitis in a Castrated Dog and Preputial Oedema in an Intact Male

In this study, two unusual presentations of canine prostatitis are described; in the first case a 10‐years‐old neutered Boxer dog was presented to the Veterinary Teaching Hospital of the University of Extremadura with a complaint of anorexia, apathy and preputial discharge. In the second case, a local veterinarian referred an 8‐years‐old male Labrador to the Veterinary Teaching Hospital of the University of Extremadura. The dog had a history of pain in the caudal abdomen and preputial oedema. The final diagnosis in both cases was acute prostatitis. It is concluded that although canine prostatitis is a common disease, sometimes can have presentations that may differ from those classically described in the literature.     

Dimethylformamide Improves the In vitro Characteristics of Thawed Stallion Spermatozoa Reducing Sublethal Damage

A total of 42 ejaculates were used in the experiment; six ejaculates per stallion, obtained from seven Pure Spanish stallions (PRE), were split and frozen in freezing media with different concentrations and combinations of cryoprotectant (CPA): (i) Cáceres (skim milk based extender) containing 2.5% glycerol (2.5GL), (ii) Cáceres containing 1.5% glycerol and 1.5% dimethylformamide (1.5%GL–1.5%DMFA), (iii) Cáceres extender supplemented with 1.5% glycerol and 2.5% dimethylformamide (1.5%GL–2.5%DMFA) and (iv) Cáceres extender supplemented with 4% dimethylformamide (4%DMFA). After at least 4 weeks of storage in liquid nitrogen (LN), straws were thawed and semen analysed by computer‐assisted sperm analysis and flow cytometry (membrane lipid architecture (Merocyanine 540), integrity and sublethal damage (YoPro‐1) and mitochondrial membrane potential (JC‐1)). After thawing, better results were observed in samples frozen in 4%DMFA or in combinations of 1.5%GL–2.5%DMFA, in fact total motility increased by 16% in the 4%DMFA group compared to 2.5%GL (P < 0.05). Also, there was an increment in the percentage of progressive motile sperm in the 1.5%GL–2.5%DMFA group (9.8% 2.5GL vs 19% in the 1.5%GL–2.5%DMFA group p < 0.05); also, samples frozen in the 4%DMFA group had more intact (YoPro‐1 negative) sperm post‐thawing, 29.3% in 2.5%GL vs 36.7% in 4%DMFA group (p < 0.05). Membrane lipid architecture was not affected by any of the cryoprotectants tested, while samples frozen in 4%DFMA had a lower percentage of mitochondria with lower membrane potential. It is concluded that DMFA improves the outcome of cryopreservation of stallion spermatozoa mainly reducing sublethal cryodamage.