The onset of starch degradation during banana ripening is concomitant to changes in the content of free and conjugated forms of indole-3-acetic acid

Banana (Musa acuminata AAA cv. Nanicão) slices were infiltrated with mannitol (control) and mannitol plus indole-3-acetic acid (IAA); then, some important ripening parameters like starch degradation, synthesis of ethylene and respiration were monitored. The contents of free-IAA and conjugated forms of IAA (ester and amide) were analyzed, by GC-MS-SIM, throughout the ripening in both banana slices and whole bananas. The starch degradation of IAA-treated slices was delayed for several days, but there was no difference between control and IAA-treated slices in the ethylene and respiration profiles. On day zero after infiltration, free-IAA levels were 500-fold higher in IAA-treated slices than in the control slices, but within 72 hours they declined to values 15-fold higher than those in the control group, with concomitant increase in IAA-ester. Similar to the banana slices, the onset of starch degradation occurred in whole bananas only when the free-IAA concentration was about 4 ng/g FW. The results herein suggest that IAA levels play a role during banana ripening in events like starch degradation with the consequence of banana sweetening.     

Susceptibility of the cattle tick Boophilus microplus (Acari: Ixodidae) to isolates of the fungus Beauveria bassiana

The susceptibility of the tick Boophilus microplus to Beauveria bassiana was evaluated by inoculating eggs, larvae and engorged females of the tick with five fungal isolates at concentrations of 10⁶, 10⁷ and 10⁸ conidia/ml. Tick eggs (0.25 g) were immersed in 1 ml of a suspension of the different conidial concentrations for 1 min. Similar exposure was performed by immersion of 2000 larvae and homogeneous groups of nine engorged females in 2 and 20 ml of conidial suspension, respectively. Treated eggs, larvae and adults were placed in an incubator at 27 ± 1 °C and relative humidity above 80% for evaluation of the fungal action. All fungal isolates applied at all conidial concentrations reduced the hatching rate of larvae from treated eggs by 1.36–65.58% and increased the mortality rate of inoculated larvae by 0.8–70.49%. In the bioassay with engorged females, oviposition period was reduced by 9.69–47.80%, egg mass weight by 4.71–53.87%, estimated reproduction by 8.3–60.62%, egg production index by 5.03–54.20%, percent larval hatching by 0.27–13.96%, and the mortality rate of treated females was increased by 96.60–100%. The reduction of the estimated reproduction obtained for the treated groups ranged from 8.37 to 64.52%. The sporulation of the pathogen on dead females ranged from 3.70 to 88.88% depending on the isolate and concentration used. Isolates AM 09, CB 7 and JAB 07 were the most effective and effectiveness increased with increasing concentrations of conidia in the suspensions.     

The mechanism of reaction of nitrosyl with met- and oxymyoglobin: an ESR study

In this ESR work we have studied the pentacoordinate symmetry in horse, whale and sperm-whale myoglobin (Mb) in different physical states such as solution and powder. Experiments were performed in which the following parameters were varied: the sample temperature, pH, reaction time with NO, and NO concentration. The results enabled us to explain the NO reaction mechanism in the oxy and met forms of myoglobin. The study of powder samples at different degrees of hydration allowed us to identify the diamagnetic intermediate species existent in the reaction of NO with met-Mb proposed in the literature. The results presented explain adequately the pH effect and temperature dependence observed in the ESR spectra obtained using the met-Mb sample solutions from Sigma Chemical Co., which consist of a mixture (13%) of Mb-O2.     

Toxoplasma gondii Chitinase Induces Macrophage Activation

Toxoplasma gondii is an obligate intracellular protozoan parasite found worldwide that is able to chronically infect almost all vertebrate species, especially birds and mammalians. Chitinases are essential to various biological processes, and some pathogens rely on chitinases for successful parasitization. Here, we purified and characterized a chitinase from T. gondii. The enzyme, provisionally named Tg_chitinase, has a molecular mass of 13.7 kDa and exhibits a Km of 0.34 mM and a Vmax of 2.64. The optimal environmental conditions for enzymatic function were at pH 4.0 and 50°C. Tg_chitinase was immunolocalized in the cytoplasm of highly virulent T. gondii RH strain tachyzoites, mainly at the apical extremity. Tg_chitinase induced macrophage activation as manifested by the production of high levels of pro-inflammatory cytokines, a pathogenic hallmark of T. gondii infection. In conclusion, to our knowledge, we describe for the first time a chitinase of T. gondii tachyzoites and provide evidence that this enzyme might influence the pathogenesis of T. gondii infection.     

Effect of hydration in metHb: Reversible changes monitored by ESR of iron

The dehydration of human and bovine methemoglobins was monitored using ESR spectroscopy of the iron signal. The interconversion of the Fe(III) signal between the high spin form (at g 6) in solution and low spin form (at g 2) was quantitatively studied as a function of hydration. The dehydration process leads also to a loss of paramagnetism resulting in the appearance of about 40% Fe(II) below 0.40 grH₂O/grHb. The remaining 60% of Fe(III) ESR signal is distributed as the residual high spin form (at g 6, 5%) and low spin form (hemichromes H and P, 55%). The formation of hemichrome P was explained as resulting from the coordination of the cysteine residue at β⁹³ with the iron atom which follows the rupture of the proximal histidine bond. Experiments with hemoglobins where the sulphur atom of cysteine β was blocked (N-ethylmaleimide) did not showed the hemichrome P, confirming the involvement of the sulphur atom. This implies that the dehydration process induces displacements and torsion of the F helix, drastically changing the iron coordination at proximal site. In agreement with this proposition the Fe(II) symmetry is pentacoordinated with the disrupted bond to the proximal histidine at fifth coordination. This is also supported by ESR experiments with nitrosyl complex at low hydrations. All conformational changes were reversibly modulated by hydration degree and partially by lyophilization rate. A one-cycle dehydration of bovine hemoglobin followed by solubilization shows 100% reversibility of hemichrome P. Increasing the number of cycles of dehydration-hydration reduces the reversibility degree. With three cycles a reversibility of 70%–75% is observed. The level of 0.40 grH₂O/grHb was the critical hydration for the molecules to return to aquo met form and correspond also to a minimal water content necessary to cover all protein surface as obtained from other techniques.     

UV-Light Effects on Cytochrome C Modulated by the Aggregation State of Phenothiazines

The present study shows the factors that modulate the photodamage promoted by phenothiazines. Cytochrome c was irradiated with UV light for 120 min, over a pH range from 4.0 to 8.0, in the absence and in the presence of different concentrations of thioridazine (TR) and fluphenazine (FP). In the absence of phenothiazines, the maximal rate of a Soret band blue shift (nm/min) from 409 to 406 nm was obtained at pH 4.0 (0.028 nm/min). The presence of phenothiazines at the concentration range 10-25 µmol/L amplified and accelerated a cytochrome c blue shift (409 to 405 nm, at a rate = 0.041 nm/min). Above 25 µmol/L, crescent concentrations of phenothiazines contributed to cytochrome c protection with (maximal at 2500 µmol/L). Scanning electronic microscopy revealed the formation of nanostructures. The pH also influenced the effect of low phenothiazine concentrations on cytochrome c. Thus, the predominance of phenothiazine-promoted cytochrome c damage or protection depends on a balance of the following factors: the yield of photo-generated drug cation radicals, which is favored by acidic pH; the stability of the cation radicals, which is favored by the drug aggregation; and the cytochrome c structure, modulated by the pH.