Actividad antifúngica del quitosano y aceites esenciales sobre Rhizopus stolonifer -Ehrenb: Fr Vuill, agente causal de la pudrición blanda del tomate / Antifungal activity of chitosan and essential oils on Rhizopus stolonifer Ehrenb: Fr Vuill causal agent of soft rot of tomato

Rhizopus stolonifer es el agente causal de la pudrición blanda, enfermedad poscosecha que ocasiona pérdidas económicas importantes. Se han empleado fungicidas sintéticos como el dicloran para controlar a este microorganismo, sin embargo, se ha demostrado que los fungicidas representan un riesgo para el ambiente y la salud humana. Actualmente se buscan alternativas naturales para el control de las pudriciones poscosecha. Se evaluó in vitro e in situ el efecto antifúngico del quitosano y de los aceites esenciales de canela (Cinnamomum zeylanicum), clavo (Syzygium aromaticum) y tomillo (Thymus vulgaris) y dicloran sobre Rhizopus stolonifer. Los tratamientos más efectivos para inhibir in vitro a Rhizopus stolonifer fueron obtenidos con quitosano a 10 mg mL-1, con los tres aceites esenciales probados a la concentración de 0,3 mg mL-1, las mezclas de quitosano a 10 mg mL-1 con los aceites a 0,3 mg mL-1 y el dicloran a 1 mg mL-1. Los experimentos in situ mostraron que el tratamiento individual con quitosano fue el mejor para reducir el porcentaje de infección de los frutos de tomate (Lycopersicon esculentum) inoculados con Rhizopus stolonifer y que la mezcla de quitosano con aceites esenciales no mejora la actividad antifúngica. El quitosano y el dicloran fueron los mejores tratamientos para reducir la pérdida de peso de los frutos. Los tratamientos individuales con quitosano representan una alternativa natural para controlar la pudrición blanda en frutos de tomate.

Rhizopus stolonifer is the causal agent of soft rot, postharvest disease that causes important economic losses. Synthetic fungicides such as dichloran have been used to control this microorganism; however, it has been shown that fungicides represent a risk for the environment and human health. Actually, natural alternatives are looked for the control of postharvest rotting. In vitro and in situ experiments the antifungal effect of chitosan, essential oils of cinnamon (Cinnamomum zeylanicum), clove (Syzygium aromaticum), and thyme (Thymus vulgaris) and dichloran on Rhizopus stolonifer were evaluated. The most effective treatments for in vitro inhibition of Rhizopus stolonifer were obtained by quitosano to 10 mg mL-1, with the three essential oils proved to the concentration of 0.3 mg mL-1, the mixtures chitosan to 10 mg mL-1 with the oils at 0.3 mg mL-1 and dichloran at 1 mg mL-1. In situ experiments showed that the individual treatment with chitosan was the best to reduce the infection percentage of the tomato (Lycopersicon esculentum) fruits inoculated with Rhizopus stolonifer and chitosan mixture with essential oils did not improve its antifungal activity. Chitosan and dichloran were the best treatments to reduce the weight loss of the fruits. Individual treatments with chitosan represent a natural alternative for the control of soft rot on tomato fruits.

Ferric reductase, superoxide dismutase and alkaline phosphatase activities in siderophore producing fungi.

Enzymes associated with release of iron from internalized ferrated siderophore (ferrisiderophore reductase), with damage to the cell at high iron concentration (superoxide dismutase) and siderophore synthesis (alkaline phosphatase), were examined in 3 test fungi viz., Aspergillus sp. ABp4, Aureobasidium pullulans and Rhizopus sp. Extracellular ferrisiderophore reductase activity was present in all the three fungi, but Aureobasidium pullulans, that showed the highest activity (84.3 microM min(-1)), was the only one to produce intra-cellular ferric reductase (147.9 microM min(-1)). Superoxide dismutase was produced by Aureobasidium pullulans and Rhizopus sp., but not by Aspergillus sp. ABp4, that showed intra-cellular enzyme activity in case of ferric reductase and alkaline phosphatase. Maximum SOD activity was seen in Aureobasidium pullulans both extra-cellularly (93.83 ng ml(-1)) and intra-cellularly (57.14 ng ml(-1)). All the test fungi examined, produced intra-cellular alkaline phosphatase. There was no extracellular alkaline phosphatase. Among the three fungi, Aureobasidium pullulans showed highest alkaline phosphatase activity (129.9 microM min(-1)) and Aspergillus sp. ABp4 the least (76.4 microM min(-1)).

Production and some properties of proteinase by fungi utilizing whey

Ten fungal strains were screened on different kinds of whey and permeate for their ability to produce proteinase enzymes. Rhizopus sp. NRC 5 was selected as a new isolate from acid whey characterized by its high proteolytic productivity when grown in submerged culture of acid whey for 5 days at 30C +/- 2. Addition of CaC12 to the fermentation medium [acid whey] stimulated the enzymes formation, while MgC12 inhibited the enzymes biosynthesis. The produced enzymes gave high proteolytic activity at acidic and alkaline pH [5.5 and 8]. Optimum reaction temperature and time for acidic and alkaline proteinase enzymes were 40C for 15 and 25 min, respectively. Maximum acidic and alkaline proteolytic enzymes activity was obtained at enzyme concentrations of 0.54 mg/ml besides substrate concentration of 2.0 g%

Extracellular production of alpha-rhamnosidase by Rhizopus nigricans.

Extracellular production of alpha-rhamnosidase [EC 3.2.1.40] from an indigeneous fungal strain of R. nigricans has been demonstrated. The enzyme has been shown to follow Michaelis-Menten kinetics using p-nitrophenyl-alpha-L-rhamnopyranosid as a substrate. The pH and the temperature optima of the enzyme have been found to be around 6.5 and 60 degrees-80 degrees C respectively.

Production of extracellular acidic lipase by Rhizopus arrhizus as a function of culture conditions.

Thermophilic strain of Rhizopus arrhizus accumulates an acidic lipase in culture fluid when grown in a medium containing ground nut oil, milk powder and inorganic salts. Addition of 2.0% ground nut oil yielded the highest productivity of enzyme. Soyabean meal and arabinose were found to be the best nitrogen and carbon sources for enzyme production respectively. Addition of metal ions such as MnCl2, SnCl2 and CaCl2 increased the enzyme productivity by 4 fold. The enzyme productivity in the fermenter was much higher (310 U/ml) than in shake-flask (180 U/ml). Crude lipase preparation showed pH and temperature activity optima at 3.5 and 45 degrees C respectively. The enzyme is thermostable and highly active in hydrolysing triglycerides and failed to hydrolyse-methyl esters of caprylate and palmitate.

Caracterizaçäo da lipase secretada por Rhizopus oligosporus

A lipase secretada no meio de cultura por Rhizopus oligosporus apresenta maior velocidade de hidrólise nos tempos iniciais da reaçäo enzimática ou em concentraçöes menores de enzima. O efeito do pH da reaçäo enzimática sobre a atividade lipolítica propicia uma curva com maior velocidade de hidrólise em valor de pH 8,5 e nitidamente descendente do lado alcalino do pH da reaçäo. Sais minerais, como o cloreto de sódio e o cloreto de cálcio, adicionados à mistura de reaçäo, aumentam a atividade lipolítica em cerca de 45% em média. Por sua vez, os sais biliares (taurocolato e desoxicolato de sódio) ativam a enzima em 22-25% quando em concentraçöes baixas e inibem acentuadamente quando em concentraçöes altas. A enzima secretada no meio de cultura suplementado com extrato de farelo de soja, óleo de soja ou Tween 80 apresenta uma pequena atividade de esterase em níveis correspondentes à atividade de lipase