Interaction of fatigue and hypercapnia in the canine diaphragm

We studied 10 open-chest dogs and measured the pressure across the diaphragm (Pdi) in each period of the protocol during stimulation at frequencies of 1, 20, 50, and 80 Hz. Three ranges of arterial PCO2 (PaCO2) were examined: less than or equal to 26, 36-50, and greater than or equal to 89 Torr. The diaphragm was fatigued with repetitive phrenic stimulation (30 Hz). During the fatiguing activity, five of the animals were subjected to hypercapnia and the other five to hypocapnia. A frequency-Pdi curve was generated for each period in the protocol. The data show that 1) fatiguing to 50% of the initial Pdi value during hypercapnia was significantly more rapid than during hypocapnia; 2) both the prefatigue and postfatigue mean Pdi values over all interactions of frequency, fatigue, and PaCO2 were unaffected by the fatiguing environment (hypercapnia vs. hypocapnia); 3) the percent reduction of Pdi by hypercapnia was the same at all four frequencies; 4) hypocapnia did not alter either the pre- or postfatigue frequency-Pdi curve; and 5) one-half relaxation time, unaffected by PaCO2, was prolonged by fatigue. We conclude that the hypercapnic diaphragm has less endurance than the hypocapnic diaphragm and that although both fatigue and hypercapnia decrease Pdi, they appear to be separate entities working through different mechanisms.     

Production of pectinases by Aspergillus niger in solid state fermentation at high initial glucose concentrations

Exopectinase (exo-p) and endopectinase (endo-p) production by Aspergillus niger CH4 in solid state culture was studied at initial glucose concentrations of 100, 250, 350 and 450 g/l. The highest activity of exo-p (35 U/g) was produced at 72 and 120 h in the medium containing 100 and 250 g glucose/l, respectively. The maximum endo-p activity (9 U/g) was produced at 72 h in the medium with 250 g glucose/l. The reduction in pectinase production at 350 and 450 g/l initial glucose concentration was due neither to repression of the synthesis of the enzyme nor to the glucose consumption rate of the strain but due to a drastic drop in pH of the medium.S. Solis-Pereyra, E. Favela-Torres, M. Gutiérrez-Rojas, G. Saucedo-Castañeda and G. Viniegra-González are with the Departamento de Biotecnologia, Universidad Autónoma Metropolitana, A.P. 55-535, C.P. 09340, México D.F., México; S. Roussos is with the Laboratoire de Biotechnologie, ORSTOM, B.P. 5045, 34032, Montpellier Cedex, France, and P. Gunasekaran is with the Department of Microbial Technology, School of Biological Sciences, Madurai Kamaraj University, Madurai, 625-021, India.     

Strategies for large scale inoculum development for solid state fermentation system: Conidiospores of Trichoderma harzianum

Summary The suitability of disc fermenter for efficient production of conidiospores by Trichoderma harzianum is limited to a working capacity of 0.61 agar medium due to large decrease in spore production per cm² of the culture surface area with further increase in the capacity. In contrast, Zymotis, a large scale solid state fermenter designed at ORSTOM, France and use of inert solid support for absorbing nutrients offer many advantages for production of conidiospores in quantity sufficient enough to inoculate pilot and larger fermenters. Five times higher production of conidiospores in Zymotis, as compared to the agar medium in flask, constitutes a success in the development of large scale inoculum.     

Effects of diaphragmatic ischemia on the inspiratory motor drive.

To assess the effect of diaphragmatic ischemia on the inspiratory motor drive, we studied the in situ isolated and innervated left diaphragm in anesthetized, vagotomized, and mechanically ventilated dogs. The arterial and venous vessels of the left diaphragm were catheterized and isolated from the systemic circulation. Inspiratory muscle activation was assessed by recording the integrated electromyographic (EMG) activity of the left and right costal diaphragms and parasternal intercostal and alae nasi muscles. Tension generated by the left diaphragm during spontaneous breathing attempts was also measured. In eight animals, left diaphragmatic ischemia was induced by occluding the phrenic artery for 20 min, followed by 10 min of reperfusion. This elicited a progressive increase in EMG activity of the left and right diaphragms and parasternal and alae nasi muscles to 170, 157, 152, and 128% of baseline values, respectively, an increase in the frequency of breathing efforts, and no change in left diaphragmatic spontaneous tension. Thus the ratio of left diaphragmatic EMG to tension rose progressively during ischemia. During reperfusion, only the frequency of breathing efforts and alae nasi EMG recovered completely. In four additional animals, left diaphragmatic ischemia was induced after the left phrenic nerve was sectioned. Neither EMG activity of inspiratory muscles nor respiratory timing changed significantly during ischemia. In conclusion, diaphragmatic ischemia increases inspiratory motor drive through activation of phrenic afferents. The changes in alae nasi activity and respiratory timing indicate that this influence is achieved through supraspinal pathways.     

Effect of phrenic afferent stimulation on pattern of respiratory muscle activation.

Ventilation and electromyogram (EMG) activities of the right hemidiaphragm, parasternal intercostal, triangularis sterni, transversus abdominis, genioglossus, and alae nasi muscles were measured before and during central stimulation of the left thoracic phrenic nerve in 10 alpha-chloralose anesthetized vagotomized dogs. Pressure in the carotid sinuses was fixed to maintain baroreflex activity constant. The nerve was stimulated for 1 min with a frequency of 40 Hz and stimulus duration of 1 ms at voltages of 5, 10, 20, and 30 times twitch threshold (TT). At five times TT, no change in ventilation or EMG activity occurred. At 10 times TT, neither tidal volume nor breathing frequency increased sufficiently to reach statistical significance, although the change in their product (minute ventilation) was significant (P less than 0.05). At 20 and 30 times TT, increases in both breathing frequency and tidal volume were significant. At these stimulus intensities, the increases in ventilation were accompanied by approximately equal increases in the activity of the diaphragm, parasternal, and alae nasi muscles. The increase in genioglossus activity was much greater than that of the other inspiratory muscles. Phrenic nerve stimulation also elicited inhomogeneous activation of the expiratory muscles. The transversus abdominis activity increased significantly at intensities from 10 to 30 times TT, whereas the activity of the triangularis sterni remained unchanged. The high stimulation intensities required suggest that the activation of afferent fiber groups III and IV is involved in the response. We conclude that thin-fiber phrenic afferent activation exerts a nonuniform effect on the upper airway, rib cage, and abdominal muscles and may play a role in the control of respiratory muscle recruitment.     

In vitro propagation of olive (Olea europaea L.) cv. Koroneiki

Single node explants of 'Koroneiki' olive trees werecultured for one month on a modified Driver-Kuniyuki for Walnut medium, lackinggrowth regulators. The explants were subcultured once a month on a mediumsupplemented with zeatin riboside, 6-(--dimethylallylamino)purine,6-benzyladenine or thidiazuron. Zeatin riboside proved to be superior to othercytokinins in inducing shoot proliferation. The combination of olive knotextract at 25 or 50 mg l^–1 with cytokininssuppressed shoot proliferation. After two months at the proliferation stage,theexplants were cultured for one week in the dark in 1 ml liquidWoody Plant Medium supplemented with IBA, -NAA or IBA+-NAA. Theexplants were then transferred to the same solid medium lacking growthregulators, with a small layer of perlite on the surface. The combination ofthetwo auxins at 1+1 mg l^–1 resulted in almost 76%rooting. The combination of olive knot extract at 50 mgl^–1 with auxins increased the rooting percentage up toalmost 87%. Artificial infection of explants with the bacteriumPseudomonas savastanoi pv. savastanoiinhibited rhizogenesis, even in the presence of auxins. Rooted explants weresuccessfully acclimatised under a mist system, with the survival rate reachingalmost 75%.     

Challenges and opportunities of the bio-pesticides production by solid-state fermentation: filamentous fungi as a model

In recent years, production and use of bio-pesticides have increasing and replacing some synthetic chemical pesticides applied to food commodities. In this review, biological control is focused as an alternative, to some synthetic chemical treatments that cause environmental, human health, and food quality risks. In addition, several phytopathogenic microorganisms have developed resistance to some of these synthetic chemicals and become more difficult to control. Worldwide, the bio-pesticides market is growing annually at a rate of 44% in North America, 20% in Europe and Oceania, 10% in Latin and South American countries and 6% in Asia. Use of agro-industrial wastes and solid-state fermentation (SSF) technology offers an alternative to bio-pesticide production with advantages versus conventional submerged fermentations, as reduced cost and energy consumption, low production of residual water and high stability products. In this review, recent data about state of art regarding bio-pesticides production under SSF on agroindustrial wastes will be discussed. SSF can be defined as a microbial process that generally occurs on solid material in the absence of free water. This material has the ability to absorb water with or without soluble nutrients, since the substrate must have water to support the microorganism’s growth and metabolism. Changes in water content are analyzed in order to select the conditions for a future process, where water stress can be combined with the best spore production conditions, obtaining in this way an inexpensive biotechnological option for modern agriculture in developing countries.     

Enzymatic synthesis of perillyl alcohol derivatives and investigation of their antiproliferative activity

The monoterpene perillyl alcohol (POH), an intermediate in the plant terpenoid biosynthetic pathway, has well-established tumor chemopreventive and chemotherapeutic potential. We have previously shown that the primary hydroxyl group of POH is essential for its antitumor and anti-angiogenic activities. In the current study we present the enzymatic synthesis of two POH derivatives with different polar and hydrophobic characteristics, namely perillyl glucoside and perillyl glucoside fatty ester, through a two-step modification. Initial glucosylation of POH on its active hydroxyl group with D-(+)-glucose and subsequent esterification of the perillyl glucoside product with vinyl laurate were carried out using almond β-glucosidase and lipase B from Candida antarctica, respectively, in a low-water system. Optimization of enzymatic reactions was performed to achieve the highest possible conversion yields. The antitumor cell proliferation activity against mouse Lewis lung carcinoma cells was retained in both derivatives, although the perillyl glucoside ester showed greater inhibition than perillyl glucoside. Our results underline the feasibility of enzymatically producing novel bioactive analogs of phytochemicals displaying useful physicochemical properties.     

Dioxygenase from Aspergillus fumigatus MC8: molecular modelling and in silico studies on enzyme–substrate interactions

Flavoenzymes have been extensively studied for their structural and mechanistic properties because they find potential application as industrial biocatalysts. They are attractive for biocatalysis because of the selectivity, controllability and efficiency of their reactions. Some of these enzymes catalyse the oxidative modification of protein substrates. Among them oxygenases (monoxoygenases and dioxygenases) are of special interest because they are highly entantio as well as regio-selective and can be used for oxyfunctionalisation. Dioxygenase enzymes catalyse oxygenation reactions in which both di-oxygen atoms are incorporated into the product. A dioxygenase enzyme purified from Aspergillus fumigatus MC8 was subjected to protein digestion followed by peptide sequencing. The sequence analysis of the peptide fragments resulted in identifying its match with that of an extracellular dioxygenase sequence from the same species of fungus existing in the protein database. The sequence was submitted to protein homology/analogy recognition engine online server for homology modelling and the 3D structure was predicted. Subsequently, the in silico studies of the enzyme–substrate (protein–ligand) interaction were carried out by using the method of molecular docking simulations wherein the modelled dioxygenase enzyme (protein) was docked with the substrates (ligands), catechin and epicatechin.