Effect of microplastics on the activity of carboxylesterase and phosphatase enzymes in Scinax squalirostris tadpoles.

Microplastics (MPs) are critical emerging pollutants around the world. There is a growing interest in the effects of MP ingestion, non-digestion, and toxicity on aquatic organisms. Amphibian tadpoles are the vertebrate group that has received the least attention regarding this issue. The aim of the present study was to determine the ingestion of polyethylene MPs by Scinax squalirostris tadpoles by atomic force microscopy (AFM) and to evaluate the activities of carboxylesterase (CbE, using 4-naphthyl butyrate-NB-, and 1-naphthyl acetate -NA- as substrates) and alkaline phosphatase (ALP) under MP exposure. Enzyme activities were analyzed spectrophotometrically at 2 and 10 days of exposure. Tadpoles were exposed to two different treatments during 10 days: a negative control (CO, dechlorinated water) and MP (60 mg L-1). AFM images of the digestive contents of tadpoles revealed the presence of MPs. After 10 days of MP exposure, CbE (NB) activity was significantly higher and CbE (NA) activity was significantly lower in MP treatments than in controls. ALP activity decreased in MP treatments after 2 and 10 days of exposure. The detection of MP particles in the intestinal contents and the effects on metabolic enzymes in a common frog species evidenced the potential health risk of MP to aquatic vertebrates. Thus, the differential response in enzymes and substrates demonstrate the need for considering the complex effects of contaminants and nutrients on ecosystems for ecotoxicological risk characterization.

Biochemical characterization and immunolocalization studies of a Capsicum chinense Jacq. protein fraction containing DING proteins and anti-microbial activity.

The DING protein family consists of proteins of great biological importance due to their ability to inhibit carcinogenic cell growth. A DING peptide with Mr ∼7.57 kDa and pI ∼5.06 was detected in G10P1.7.57, a protein fraction from Capsicum chinense Jacq. seeds. Amino acid sequencing of the peptide produced three smaller peptides showing identity to the DING protein family. G10P1.7.57 displayed a phosphatase activity capable of dephosphorylating different phosphorylated substrates and inhibited the growth of Saccharomyces cerevisiae cells. Western immunoblotting with a custom-made polyclonal antibody raised against a sequence (ITYMSPDYAAPTLAGLDDATK), derived from the ∼7.57 kDa polypeptide, immunodetected an âˆ¼ 39 kDa polypeptide in G10P1.7.57. Purification by electroelution followed by amino acid sequencing of the ∼39 kDa polypeptide yielded seven new peptide sequences and an additional one identical to that of the initially identified peptide. Western immunoblotting of soluble proteins from C. chinense seeds and leaves revealed the presence of the ∼39 kDa polypeptide at all developmental stages, with increased accumulation when the organs reached maturity. Immunolocalization using Dabsyl chloride- or Alexa fluor 488-conjugated antibodies revealed a specific fluorescent signal in the cell cytoplasm at all developmental stages, giving support to the idea that the ∼39 kDa polypeptide is a soluble DING protein. Thus, we have identified and characterized a protein fraction with a DING protein from C. chinense.