A High Redox Potential Laccase from Pycnoporus sanguineus RP15: Potential Application for Dye Decolorization

Abstract

Laccase production by Pycnoporus sanguineus RP15 grown in wheat bran and corncob under solid-state fermentation was optimized by response surface methodology using a Central Composite Rotational Design. A laccase (Lacps1) was purified and characterized and the potential of the pure Lacps1 and the crude culture extract for synthetic dye decolorization was evaluated. At optimal conditions (eight days, 26 °C, 18% (w/w) milled corncob, 0.8% (w/w) NH4Cl and 50 mmol·L−1 CuSO4, initial moisture 4.1 mL·g−1), the laccase activity reached 138.6 ± 13.2 U·g−1. Lacps1 was a monomeric glycoprotein (67 kDa, 24% carbohydrate). Optimum pH and temperature for the oxidation of 2,2’-azino-bis(3-ethylbenzthiazoline-6-sulfonate) (ABTS) were 4.4 and 74.4 °C, respectively. Lacps1 was stable at pH 3.0–8.0, and after two hours at 55–60 °C, presenting high redox potential (0.747 V vs. NHE). ABTS was oxidized with an apparent affinity constant of 147.0 ± 6.4 μmol·L−1, maximum velocity of 413.4 ± 21.2 U·mg−1 and catalytic efficiency of 3140.1 ± 149.6 L·mmol−1·s−1. The maximum decolorization percentages of bromophenol blue (BPB), remazol brilliant blue R and reactive blue 4 (RB4), at 25 or 40 °C without redox mediators, reached 90%, 80% and 60%, respectively, using either pure Lacps1 or the crude extract. This is the first study of the decolorization of BPB and RB4 by a P. sanguineus laccase. The data suggested good potential for treatment of industrial dye-containing effluents.

Use of sugarcane molasses by Pycnoporus sanguineus for the production of laccase for dye decolorization

“…However, in our study the enzymatic extract produced 80% decolorization of this dye (Figure 4), which was the highest level (24 h; pH 5). Similar results were reported by Iracheta-Cárdenas et al (2016) when the enzymatic extract of P. sanguineus caused RBBR decolorization of 90% (4 h; pH 5), and by Zimbardi et al (2016) who used a crude enzymatic extract of P. sanguineus RP15 to decolorize RBBR by 70 or 80% (2 h; 25 or 40°C, respectively).…”

“…sanguineus has been successfully cultivated for the production of laccases in a variety of lignocellulosic waste materials or by-products from the agro-industry (Zimbardi et al, 2016). The production of laccase at peak activity and after culture condition optimization using sugarcane molasses was 8.5-fold higher than that reported by Zimbardi et al (2016) for the RP15 strain of P. sanguineus cultivated in wheat bran (5900 U/L) under solid state fermentation. The production of laccase by P. sanguineus U13-4 was 5-fold higher than laccase production by Agaricus blazei (9700 U/L), also cultivated with sugarcane molasses (Valle et al, 2014a).…”

Fungal Laccase Production from Lignocellulosic Agricultural Wastes by Solid-State Fermentation: A Review

“…They were first described in Toxicodendron vernicifluum (Japanese lacquer tree; formerly called Rhus vernicifera) and subsequently in a wide variety of organisms, including bacteria, insects, and fungi (notably, white rot fungi) [3]. Laccases display broad substrate specificity and are applied in many industrial and environmental technology areas, including in textile effluents (decolorization, detoxification), paper production (biobleaching, biopulping), and biopharmaceuticals (transformation of antibiotics, steroids) [4][5][6]. Their ability to remove xenobiotic substances and generate polymeric products makes them useful in bioremediation processes [7,8].…”

PRODUÇÃO DE LACASE DE Pycnoporus sanguineus EM MEIO DE CULTIVO À BASE DE MELAÇO SOJA

“…Entretanto, a produção dessa enzima em meios contendo fontes alternativas e mais baratas de nutrientes ainda é pouco conhecida e explorada. Poucos são os estudos que avaliaram o cultivo de P. sanguineus em diferentes subprodutos agroindustriais para produção de lacase (MARIM et al, 2016;ZIMBARDI et al, 2016). A produção com melaço de soja após a otimização do cultivo foi 1,5 menor do que a produção relatada por Marim et al (2016) com essa mesma linhagem, mas cultivada com melaço de cana--de-açúcar 3,4 g/L de nitrogênio (ureia) e sem cobre como indutor.…”

“…A produção com melaço de soja após a otimização do cultivo foi 1,5 menor do que a produção relatada por Marim et al (2016) com essa mesma linhagem, mas cultivada com melaço de cana--de-açúcar 3,4 g/L de nitrogênio (ureia) e sem cobre como indutor. Entretanto, a produção nesse estudo foi seis vezes maior que a obtida por Zimbardi et al (2016) para a linhagem de P. sanguineus RP15 em cultivo no estado sólido com farelo de trigo (5900 U/L). O melaço de soja possui entre 50 e 60% carboidratos com uma mistura de diferentes sacaríde-os como sacarose (17-28%), estaquiose (17-19%), rafinose (9-10%), frutose (0,5-1,5%) e glicose (0,2-0,5%), além de quantidade significativa de proteína (9,5%) e lipídeos (20%) (SIQUEIRA et al, 2008).…”