The vegetable rennet of Cynara cardunculus L. contains two proteinases with chymosin and pepsin-like specificities

The flowers of cardoon (genus Cynara) are traditionally used in Portugal for cheese making. In this work the vegetable rennet of the species Cynara cardunculus L. was characterized in terms of enzymic composition and proteolytic specificity of its proteinases (cardosin A and cardosin B). Cardosin A was found to cleave insulin B chain at the bonds Leu15-Tyr16, Leu17-Val18 and Phe25-Tyr26. In addition to the bonds mentioned cardosin B cleaves also Glu13-Ala14, Ala14-Leu15 and Phe24-Phe25 indicating that it has a broader specificity. The kinetic parameters for the hydrolysis of the synthetic peptide Leu-Ser-Phe(NO₂)-Nle-Ala-Leu-oMe were also determined and compared to those of chymosin and pepsin. The results obtained indicate that in terms of specificity and kinetic parameters cardosin A is similar to chymosin whereas cardosin B is similar to pepsin. It appears therefore that the enzyme composition of cardoon rennet closely resembles that of calf rennet.     

Dihydrochalcones, coumarins and alkaloids from Metrodorea nigra

Two novel dihydrochalcones, 2′,3,4′,6′-tetrahydroxy-4-methoxy-3′,5-di-(3,3-dimethylallyl)-dihydrochalcone and 2′,.3,6′-trihydroxy-4-methoxy-5-(3,3-dimethylallyl)-3′,4′-(2″,2″-dimethyldihydropyran)-dihydrochalcone, have been isolated from fresh fruits of Metrodorea nigra. Stems and leaves showed a similar composition and we have isolated common steroids, simple coumarins, several furocoumarins, furoquinoline alkaloids and a furofuran lignan. From stems, we have also isolated the pentacyclic 6-C-monoterpenyl-5,7-dioxycoumarin, deoxybruceol. Structures of the isolated compounds were elucidated on the basis of spectral data.     

Factors controlling somatic embryogenesis

Histological and ultrastructural, molecular and elemental distribution changes were investigated during the induction of direct somatic embryogenesis using theCamellia japonica leaf culture system. In this culture system, direct somatic embryogenesis is induced in a controlled way in a specific leaf region (leaf blade) within a leaf. Embryogenic and non-embryogenic leaf regions have characteristic energy-dispersive X-ray spectra already before induction. According to these results electron probe X-ray microanalysis (EPMA) can be a tool for early diagnosis of embryogenic competence. Histological studies showed that severe fluctuations in the number of calcium oxalate crystals and in starch accumulation occur after induction but only in induced tissues. Changes in the cell wall composition of competent cells occur shortly after the induction treatment. The induction of morphogenesis is linked to the appearance of callose covering the surface cells of induced leaves and calluses. A 2^nd deposition of material (cutin) is necessary for normal somatic embryogenesis to occur. The involvement of lipid transfer proteins in the appearance of cutin in the embryogenic regions of the explant is suggested.     

Influence of external factors on secondary embryogenesis and germination in somatic embryos from leaves of Quercus suber

Somatic embryogenesis was obtained in cultures of leaves from young seedlings of Quercus suber L. A two-stage process, in which benzyladenine and naphthaleneacetic acid were added first at high and then at low concentrations, was required to initiate the process. Somatic embryos arose when the explants were subsequently placed on medium lacking plant growth regulators. The embryogenic lines remained productive, by means of secondary embryogenesis, on medium without growth regulators. However, this repetitive induction was influenced by the macronutrient composition of the culture medium. Both low total nitrogen content and high reduced nitrogen concentration decreased the percentage of somatic embryos that showed secondary embryogenesis. Our results suggest that alternate culture on medium that increases embryo proliferation and a low salt medium prohibiting embryo formation will partially synchronize embryo development. Chilling slightly reduced secondary embryogenesis but gave a modest increase in germination. Maturation under light followed by storage at 4 °C for at least 30 days gave the best results in switching embryos from an embryogenic pathway to a germinative one. Under these conditions 15% of embryos showed coordinated root and shoot growth and 35% formed either shoots or mostly roots. These percentages were higher than those of embryos matured in darkness. This result indicates that a specific treatment is required after maturation and before chilling to activate the switch from secondary embryo formation to germination.Abbreviations BA benzyladenine - NAA naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - BA indolebutyric acid - MS Murashige & Skoog (1962) medium - SH Schenk & Hildebrandt (1972) medium - G Gamborg (1966, PRL-4-C) medium (macronutrients in mg l^–1: NaH₂PO₄·H₂O, 90; Na₂HPO₄, 30; KCl, 300; (NH₄)₂SO₄, 200; MgSO₄·7H₂O, 250; KNO₃, 1000, CaCl₂·2H₂O, 150) - PGR plant growth regulator     

Analysis of vertical ozone and nitrogen oxides profiles in aPrunus cerasifera canopy

An automatic system was installed for continuous analyses of ozone, sulphur dioxide, nitrogen monoxide and nitrogen dioxide in an experimental orchard with a canopy ofPrunus cerasifera plants in summer 1993. Air samples from three elevations (0.8 m, 1.6 m and 3 m above ground) were sequentially analyzed. Ozone concentrations above the canopy were usually higher than within the canopy; their relationships with stomatal resistance have been investigated. Sulphur dioxide levels were negligible. Nitrogen oxides showed a complex profile, with no particular trend, likely due to a reciprocal exchange between the atmosphere and the ground surface.     

Synthesis and biological evaluation of novel NK-1 tachykinin receptor antagonists: The use of cycloalkyl amino acids as a template

In the course of a program aimed at synthesizing novel, potent NK-1 tachykinin receptor antagonists, we developed upon a bioactive model by comparing the low energy structures of a series of peptide and nonpeptide Substance P antagonists. The comparison was based on the super imposition of the aromatic rings, assuming that the rest of the molecule behaves predominantly as a template to arrange the key aromatic groups in the right spatial position. A series of 2-aminocyclohexane carboxylic acid analogues were then selected as the best templates for reproducing the postulated bioactive structure, leading to several pseudo-peptides with interesting biological activity. According to the molecular modeling, these compounds exhibit a neat parallel facing of the indolyl and naphthyl groups at about 3 Å distance. Ultraviolet absorption and steady state fluorescence measurements support this conclusion, showing a linear correlation between the spectral properties and the binding affinity of these analogues. Stacking of the indole ring with naphthalene gives rise to a complex characterized by a well-defined molar extinction coefficient. Consistently, steady state and lifetime fluorescence measurements suggest that the quenching process is ascribable to ground-state interactions between the chromophores. Implications of the π stacking propensity of aromatic groups in the biological activity of the compounds examined are briefly discussed. © 1995 John Wiley & Sons, Inc.     

Model for prediction of immobilizedAcetobacter cell number in calcium alginate gel droplets

The geometry of calcium alginate gel spheres is studied by fractal interpretation for prediction of number of cells to be immobilized. For alginate concentrations from 1 to 5% the simulated results are of 6.65×10⁸, 1.44×10⁸ and 5.27×10⁷ N cell mL⁻¹ for respectively 1.5, 2.5 and 3.5 mm gel sphere diameters. Simulated data are compared with those resulting from literature and particularly with experimental trials where from 10⁷ to 10⁸ N cell mL⁻¹, in 2% alginate beads with a diameter of 1.5 mm, is able to fulfil mechanical and swelling characteristics of alginate gel beads besides to supply good oxygenation.     

Simultaneous determination of NO and NO in fisheries wastewaters high in chloride concentration by capillary ion electrophoresis with direct UV detection

The simultaneous determination by capillary ion electrophoresis of nitrate and nitrite in wastewaters with chloride concentrations of 15 to 23g/l is described. Chloride concentrations over 200mg/l hampered the determination; thus, lead, mercuric and silver acetate were used to precipitate chloride. Silver acetate added in 1.5 times the stoichiometric amount for AgCl formation gave the best results in terms of nitrate and nitrite peak resolution.     

Mineralogical transformation and neoformation in granite caused by the lichens Tephromela atra and Ochrolechia parella

Tephromela atra and Ochrolechia parella are among the most abundant lichens colonizing granitic monuments in the region of Galicia (northwest Spain). In this work, their interaction with a two-mica granite used in the construction of the Toxosoutos Monastery (Noia, Galicia) was studied, using optical microscopy, scanning electron microscopy (with back-scattered-electron and energy-dispersive X-ray detection), X-ray diffractometry and atomic absorption spectroscopy to evaluate their physical, mineralogical and chemical effects. Both lichens contributed to physical weathering by penetrating intermineral voids and mineral cleavage planes, disaggregating the rock and entrapping the loosened mineral grains in their thalli. Significant chemical and mineralogical weathering also occurred, including depletion of potassium from biotite, transformation of this mica into hydroxyaluminium-vermiculite, and neoformation of whewellite and calcite in the lichen thalli. Neoformation of these calcium minerals on a calcium-poor rock such as granite is noteworthy, and this is the first time calcium carbonate has been detected within a lichen colonizing a granitic rock. Precipitation of the calcium carbonate was attributed to the local pH in the thalli having been raised due to release of sodium from nearby plagioclase during weathering.