65ª Reunião Anual da SBPC

C. Ciências Biológicas - 3. Bioquímica - 3. Enzimologia

CORN STEEP LIQUOR: A AGROINDUSTRIAL WASTE USED AS A SOURCE OF INDUCTION OF PRODUCTION ENZYMES BY MICROALGAE Chlorella vulgaris

Páblo Eugênio da Costa e Silva - Laboratory of Immunopathology Keizo Asami – LIKA Raquel Pedrosa Bezerra - Department of Animal Physiology and Morphology – UFRPE Daniela de Araújo Viana Marques - Department of Animal Physiology and Morphology – UFRPE João Carlos Monteiro de Carvalho - Department of Biochemical and Pharmaceutical Technology – USP Ana Lúcia Figueiredo Porto - PhD Advisor – Department of Animal Physiology and Morphology – UFRPE

INTRODUÇÃO:

Enzymes are biomolecule which play a very important role in various industries such as food, textile and pharmaceutical industries. Several industrial wastes can be used as nutrient source to enzyme production by microorganisms. The corn steep liquor is a residue obtained by maceration of the corn grain and contains very high protein content. It can be used as nitrogen source for protease production and cell growth of various micro-organisms, including microalgae. The advantages to use agroindustrial wastes is their low cost, prevent its disposal in environmental, besides of add values to these wastes due to its applicability in the biotechnological production of enzymes, such as in this work.

OBJETIVO DO TRABALHO:

The objective of this study was to evaluate the proteolytic enzymes production from the microalgae Chlorella vulgaris using corn steep liquor as nitrogen source.

MÉTODOS:

Microalgae Chlorella vulgaris (UTEX 1803) was grown in Bold's Basal medium (BBM) (medium 1) and in BBM containing 0.5% corn steep liquor (medium 2). At the stationary phase of growth, both cultures were harvested, centrifuged and lyophilized. For enzymatic extraction, 50 mg biomass was homogenized in phosphate buffer for 30 minutes, centrifuged at 5000 rpm for 10 minutes and the supernatant was used for protease activity. The protease activity was performed using azocasein as substrate. Was added to microcentrifuge tubes, 50 microliters of azocasein 1% (w/v) prepared in 0.2 M Tris-HCl, pH 7.2. The solution was incubated with 30 µL of enzymatic extract for 60 minutes at 25 ºC. Forward, 240 µL of trichloroacetic acid (TCA) 10% (w/v) was added. After 15 minutes, the tubes were centrifuged for 5 minutes at 10,000 rpm. 350 microliters of supernatant was added to 650 µL of NaOH (1 M), and the absorbance of the mixture was measured in a spectrophotometer at 450 nm against a blank. One unit (U) of enzyme activity was defined as the amount of enzyme able to hydrolyze azocasein giving an increase of 0.001 units of absorbance per minute. The cultivation was done in duplicated for 15 days.

RESULTADOS E DISCUSSÃO:

C. Vulgaris biomass cultivated in the medium 1 and 2 presented extract cell with proteolytic activity of 2.9 ± 0.8 U/mL and 21.3 U/mL ± 0.8 U/mL, respectively. The results show that the presence of 0.5 % corn steep liquor in the medium culture induced the of microalgae Chlorella vulgaris to enzymes production, significatly increasing the enzymatic activity, suggesting that the corn steep liquor can be used as a substrate for production of biotechnologically proteases.

CONCLUSÕES:

It is concluded that the substrate used for enzyme production from the microalgae Chlorella vulgaris is quite feasible, besides contributing to the use of a agroindustrial waste minimizing serious problems with their disposal at the environmental. The microalgae proved suitable for cultivation with corn steep liquor, besides being a source producing proteolytic enzymes able to hydrolyze this substrate. Comparing the values obtained from the proteolytic activity, it is found that the cultivation of the microalga with 0.5% resulted in a very significant increase in protease production, suggesting be an excellent substrate for industrial application. In later stages the present study will evaluate the activity of these proteases against pathogenic micro-organisms from human to confirm its applicability in health biotechnology.

Palavras-chave: Protease, Biomolecule, Biotechnology.