Abstract
Different Pleurotus ostreatus and Flammulina velutipes species were compared relative to their ability to produce laccase in submerged fermentation of various lignocellulosic wastes. Fungi cultivation in identical culture conditions revealed wide differences among both species and strains of the same species. The laccase secretion ability of P. ostreatus strains was superior to F. velutipes strains. Maximum laccase production on cottonseed hull, corncob, and poplar wood was secreted by P. ostreatus CY 568, P. ostreatus CCEF 89, and P. ostreatus CY 568, respectively. The nature of lignocellulosic materials played an important role in determining the expression of laccase potential of fungi. The presence of cottonseed hull improved laccase activity and accelerated the rate of enzyme production. Maximum laccase production on cottonseed hull was nearly 1.29-fold and 1.53-fold higher than that on corncob and poplar wood, respectively. Laccase activity was detected in almost all tested strains on cottonseed hull on the first day, while only a few strains on poplar wood and corncob were detected on the first day. These findings will be helpful for selecting the appropriate strain in industrial applications and for optimization of integrated industrial laccase production.
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Comparative Study on Laccase Activity of White Rot Fungi under Submerged Fermentation with Different Lignocellulosic Wastes
Qi An,a,b Jie Qiao,a Lu-Sen Bian,d Mei-Ling Han,a,c,* Xun-You Yan,a Zi-Zheng Liu,a and Chun-Yan Xie e
Different Pleurotus ostreatus and Flammulina velutipes species were compared relative to their ability to produce laccase in submerged fermentation of various lignocellulosic wastes. Fungi cultivation in identical culture conditions revealed wide differences among both species and strains of the same species. The laccase secretion ability of P. ostreatus strains was superior to F. velutipes strains. Maximum laccase production on cottonseed hull, corncob, and poplar wood was secreted by P. ostreatus CY 568, P. ostreatus CCEF 89, and P. ostreatus CY 568, respectively. The nature of lignocellulosic materials played an important role in determining the expression of laccase potential of fungi. The presence of cottonseed hull improved laccase activity and accelerated the rate of enzyme production. Maximum laccase production on cottonseed hull was nearly 1.29-fold and 1.53-fold higher than that on corncob and poplar wood, respectively. Laccase activity was detected in almost all tested strains on cottonseed hull on the first day, while only a few strains on poplar wood and corncob were detected on the first day. These findings will be helpful for selecting the appropriate strain in industrial applications and for optimization of integrated industrial laccase production.
Keywords: White rot fungi; Laccase production; Lignocellulosic wastes; Submerged fermentation
Contact information: a: College of Life Science, Langfang Normal University, Langfang 065000, Hebei, China; b: Technical Innovation Center for Utilization of Edible and Medicinal Fungi in Hebei Province, Langfang 065000, Hebei, China; c: Edible and Medicinal Fungi Research and Development Center of Universities/Colleges in Hebei Province, Langfang 065000, Hebei, China; d: Experimental Centre of Forestry in North China, Chinese Academy of Forestry, Beijing 102300, China; e: The Key Laboratory Study on Microbial Fermentation of Langfang city, Langfang 065000, Hebei, China;
* Corresponding author: meilinghan309@163.com
INTRODUCTION
Extensive research on fungi, especially white rot fungi (basidiomycetes), has been conducted to gain new productivity strains with great potential for lignocellulolytic enzymes secretion and better properties for application in industry (Elisashvili et al. 2008; An et al. 2018; Han et al. 2020). Among these lignocellulolytic enzymes, laccase (EC 1.10.3.2), a blue multi-copper oxidase, is one of the oldest known and important enzymes (Zerva et al. 2019). Laccase was first isolated from Rhus vernicifera (Yoshida 1883), and these enzymes are widely distributed in plants, some insects, bacteria, and fungi. Laccase has potential applications in numerous biotechnology, including delignification of pulp and paper, production of biofuels, drug analysis, removal of phenolic compounds in food and beverages, bioremediation, cosmetics, paints, furniture, agricultural and forestry waste disposal, nanobiotechnology, and biomedicine (Madhavi and Lele 2009; Kudanga and Le Roes-Hill 2014; Bertrand et al. 2017; Mate and Alcalde 2017; Singh and Arya 2019; Zerva et al. 2019; Han et al. 2020). Bioremediation mainly involves trichlorophenol, alkenes, industrial wastes, decolorisation of dyes, polluted soils, and herbicide degradation (Mayer and Staples 2002; Strong and Claus 2011; Ba and Kumar 2017). Furthermore, laccase plays an important role in lignin degradation, pigment biosynthesis, fruiting body formation, morphogenesis, and plant pathogenesis (Janusz et al. 2015; An et al. 2016a, 2018). In view of the rich effect in industry, agriculture, and environmental and biological processes, laccase has attracted much attention. However, all these effective applications are based on large-scale laccase production. Large-scale laccase production has become important from an economic standpoint despite the long standing technical difficulties in producing large amounts of laccase (Cardona et al. 2010; Agrawal et al. 2018; An et al. 2018). The capacity of producing laccase from different fungi is different, so it is necessary to compare the laccase production of different strains belonging to different species (Agrawal et al. 2018; An et al. 2018).
The presence of laccase, alone or together with lignin peroxidase (Lip) and manganese peroxidase (Mnp), has been demonstrated in a wide variety of white rot fungi (Mayer and Staples 2002), such as the genus Trametes, Coriolus, Pleurotus, Lentinus, Flammulina and Ganoderma (Galhaup et al. 2002; Jang et al. 2006; Elissetche et al. 2007; Arockiasamy et al. 2008; An et al. 2016a, b; Guo et al. 2017; Gupta and Jana 2018; Sadeghian-Abadi et al. 2019; Han et al. 2020). Some Trametes and Pleurotus species are efficient producers of laccase (Lorenzo et al. 2002; Castanera et al. 2015), such as T. versicolor, T. pubescens, and P. ostreatus (Galhaup et al. 2002; Dominguez et al. 2007; Park et al. 2015). Laccase activity depends on the type of fungal species or strains (Elisashvili et al. 2008; Janusz et al. 2015; An et al. 2016a, 2018; Han et al. 2017, 2018, 2020). The secretion of laccase from fungi is affected by many factors, including the concentration, ratio and complexity of carbon and nitrogen sources, fermentation method, the type and concentration of metal ions, pH, temperature, co-cultivation of different fungi, and aromatic compounds (Stajić et al. 2006; Elisashvili and Kachlishvili 2009; Janusz et al. 2015; Metreveli et al. 2017; Filipe et al. 2019; Rajavat et al. 2020).
Research on the complexity of carbon and nitrogen sources has mainly focused on lignocellulosic biomass. Lignocellulosic biomass, the most abundant and renewable eco-friendly resources on earth, is attractive because it can be used to make valuable industrial products (Pinar et al. 2017; Han et al. 2020). The area of agricultural and forestry crops is vast in China, and the types and quantities of agricultural and forestry wastes are huge. However, the majority of biomass resources are wasted. White rot fungi can utilize a variety of lignocellulosic residues by producing several extracellular secreted enzymes to grow fruiting bodies for human consumption or play an important role in the process of conversion of lignocellulosic biomass to bioethanol. Furthermore, using lignocellulosic residues to produce enzymes is an environmentally friendly and economical method (Lamia et al. 2017). The common fermentation methods are solid-state fermentation (SSF) and submerged fermentation (SmF) (An et al. 2016b). The advantage of solid-state fermentation is that it avoids dilution of enzyme products due to the high content of water in submerged fermentation (Oostra et al. 2000). However, it is easier to regulate the fermentation conditions of submerged fermentation compared with solid-state fermentation. Submerged fermentation is the main method used in industry.
Previous studies investigated the laccase production secreted by strains belonging to the genus Pleurotus with wood, tree leaves, corncob, and wheat straw (Elisashvili et al. 2008; Han et al. 2017). Laccase production of strains belonging to the genus Flammulina have been discussed in the context of the effect of metal ions and aromatic compounds (Janusz et al. 2015; An et al. 2016a). However, studies on laccase secretion of the genus Pleurotus and Flammulina strains cultured by lignocellulosic waste with different lignin content have not been reported. The present work compared the laccase activity of different P. ostreatus and F. velutipes strains by lignocellulosic wastes with different lignin content in submerged fermentation. The strains and lignocellulosic wastes that contribute to the high yield of laccase were screened to provide a basis for expanding the number of strains used in industrial application and for obtaining low-cost laccase.
EXPERIMENTAL
Materials
Microorganisms
Four white-rot basidiomycete Pleurotus ostreatus strains CCEF 89, CY 568, CCMSSC 00322, and CCMSSC 00406, and four white-rot basidiomycete Flammulina velutipes CCMSSC 00114, CCMSSC 00118, CCMSSC 05317, and CCMSSC 05331 were obtained from Institute of Microbiology, Beijing Forestry University (Beijing, China). All organisms were maintained on Complete Yeast Medium (CYM) agar medium (glucose 20 g/L, peptone 2 g/L, yeast extract 2 g/L, MgSO4·7H2O 0.5 g/L, K2HPO4·3H2O 1 g/L, KH2PO4 0.46 g/L, and agar 20 g/L) at 4 °C in the College of Life Science, Langfang Normal University.
Lignocellulosic materials
Poplar wood was obtained from Langfang Normal University (Hebei, China). Corncob and cottonseed hull were kindly provided by farmers in Chengde city, Hebei province, China. All lignocellulosic wastes were chopped into small pieces, air-dried, and ground. The particles size of lignocellulosic wastes was between 20- and 60-mesh.
Methods
Organism and inoculum preparation
The microorganism was transferred to new CYM agar medium plates and incubated at 26 °C for 7 days. Inoculants with a diameter of 5 mm were made on the new flat plate covered with mycelium by a hole punch. Under aseptic conditions, 5 inoculants were placed in 250 mL flasks containing 100 mL of CYM medium (glucose 20 g/L, peptone 2 g/L, yeast extract 2 g/L, MgSO4·7H2O 0.5 g/L, K2HPO4·3H2O 1 g/L, and KH2PO4 0.46 g/L). The medium was cultured on a rotary shaker at 26 °C and 150 rpm. After 7 days, mycelial pellets were harvested and homogenized with a laboratory blender for 2 min at 5000 rpm. The resulting suspension was used as an inoculum.
Culture conditions
The submerged fermentation (SmF) of the poplar sawdust, corncob, and cottonseed hull was performed individually at 26 °C in 250 mL flasks containing 3 g of the substrate with 100 mL of deionized water. All Erlenmeyer flasks were autoclaved at 121 °C for 30 min; after cooling, 3 mL of homogenized mycelium were used to inoculate each flask. All flasks were incubated at 26 °C.
To determine the laccase dynamics under different lignocellulosic wastes and strains conditions in SmF, laccase activities were tested from fungi growth on the 1st day to the 10th day.
Enzyme activity was measured in the culture broth from each inoculated flask, at different growth times after removing the mycelia and lignocellulosic substrate. Fermentation liquor was filtered by Whatman No. 1 filter paper and then centrifuged at 4 °C and 12000 rpm for 20 min. The supernatant was the crude enzyme liquid, which was used for measurement of enzyme activity.
Enzyme activities assays
All measurements were derived from three independent experiments, and the mean values were taken. The standard deviations for the experiments were less than ± 10%.
Laccase activity was assayed by examining the oxidation of 1 mM 2,2’-azinobis-[3-ethyltiazoline-6-sulfonate] (ABTS) to its cation radical ABTS+ at 420 nm (Ɛ420 = 3.6 × 104 M-1 cm-1). The reaction mixture contained 1 mM ABTS in 50 mM sodium acetate buffer, pH 4.2, and 50 μL of adequately diluted culture liquid (Woolfenden and Wilson 1982; An et al. 2018). One unit of enzyme activity was defined as the amount of enzyme forming 1 µmol of ABTS+ per minute.
Statistical analysis
To examine the effects of lignocellulosic materials and strains on laccase activities, two-way analysis of variance followed by the Tukey post hoc test was applied to these variables, with lignocellulosic materials and strains as fixed factors (PROC GLM, IBM SPSS software version 22.0; Armonk, NY, USA). All statistical figures were generated using the program Origin 2016 (OriginLab Corporation, Northampton, MA, USA).
RESULTS AND DISCUSSION
Effects of Lignocellulosic Materials on Production of Laccase Enzymes
Many studies have indicated the effect of nutrient medium on laccase activity (Isikhuemhen and Mikiashvilli 2009), including the kinds, concentration, and complexity of carbon/nitrogen sources. The complex carbon and nitrogen sources used are mainly lignocellulose materials. The selection of lignocellulosic materials is one of the important factors affecting the secretion of laccase by fungi. Fungi growing on different lignocellulosic materials show different laccase activities (Han et al. 2017, 2018, 2020; Leite et al. 2019). Previous studies have used a single lignocellulose material, such as wood chips, tree leaves or coffee shells, as the substrate for fungal growth. The comparative studies of different lignocellulosic materials on laccase secretion by white-rot fungi are relatively few (Elisashvili et al. 2008). Thus, this paper presents a comparative study of laccase production by P. ostreatus and F. velutipes strains in different lignocellulose materials. Poplar wood, corncob, and cottonseed hull are commonly used as lignocellulosic materials in the cultivation of P. ostreatus and F. velutipes (Han et al. 2017). Their lignin contents differ greatly. The lignin content of corncobs, hardwood stems, and cottonseed hull is 15%, 18 to 25%, and 31.7%, respectively (Sánchez 2009; Tian et al. 2013). While monitoring the extracellular laccase activities of P. ostreatus and F. velutipes strains during the submerged fermentation of cottonseed hull, most strains showed laccase activity after 1 day of incubation, except for one, the P. ostreatus CCMSSC 00406. This suggested that the presence of cottonseed hull improved laccase activity and accelerated the rate of enzyme production (Fig. 1). As shown in Table 1, the effect of the lignocellulosic material on laccase activities of tested P. ostreatus and F. velutipes strains was significant (P < 0.001) throughout the fermentation stage.
Laccase activity was detected in almost all tested P. ostreatus and F. velutipes strains on the substrate of cottonseed hull on the 1st day, except P. ostreatus CCMSSC 00406 (Fig. 1). Half of the tested strains, as P. ostreatus CCEF 89, CY568 and F. velutipes CCMSSC 00114, CCMSSC 00118, exhibited laccase activity on 1st day on the substrate of corncob (Fig. 2). Only two strains, P. ostreatus CCEF 89 and CY568, demonstrated laccase activity on 1st day on poplar wood substrate (Fig. 3). Many strains showed detectable laccase activity after being cultivated for 3 days on poplar wood substrate, for example, F. velutipes CCMSSC 05317 and CCMSSC 00118. None of these strains showed this phenomenon on corncob or cottonseed hull substrate.
Table 1. Effects of Strains, Lignocellulosic Materials, and Strains ×Lignocellulosic Materials Interactions on Laccase Activities of tested P. ostreatus and F. velutipes strains (Two-Way ANOVA)
In terms of laccase production and maximum laccase activity, the tested strains showed variations on cottonseed hull, corncob, and poplar wood. Laccase production of P. ostreatus CCEF 89 in cottonseed hull, corncob, and poplar wood ranged from 61.38 ± 4.09 U/L to 748.24 ± 9.53 U/L, 26.12 ± 2.28 U/L to 699.12 ± 44.91 U/L, and 3.32 ± 0.30 U/L to 509.75 ± 15.43 U/L, respectively (Figs. 1, 2, and 3). Maximum laccase activity for strain CCEF 89 obtained from cottonseed hull was higher than that obtained from corncob and poplar wood, by 1.07-fold and 1.47-fold, respectively (Table 2). The time of maximum laccase activity for strain CCEF 89 from cottonseed hull, corncob, and poplar wood was the 6th day, 8th day, and 7th day, respectively (Table 2). Han et al. (2017) reported that laccase activity in synthetic medium containing cottonseed hull was higher than that in synthetic medium containing sawdust or corncob, indicating that the cottonseed hull could be considered as a better inducer for enhancing the laccase activities of P. ostreatus CCEF 89 and CCEF 99. There was a similar result in the present study. The maximum laccase activity for P. ostreatus strain CY 568 obtained from cottonseed hull was 902.92 ± 25.42 U/L on 7th day, which was higher than that obtained from corncob (611.71 ± 24.21 U/L, 7th day) and poplar wood (590.72 ± 14.98 U/L, 7th day), by 1.48-fold and 1.53-fold, respectively (Table 2).
Fig. 1. Effects of cottonseed hull on laccase production by tested P. ostreatus and F. velutipes strains. Average values were calculated from individual measurements for each of three parallel cultures of eight strains.
Fig. 2. Effects of corncob on laccase production by tested P. ostreatus and F. velutipes strains. Average values were calculated from individual measurements for each of three parallel cultures of eight strains.
The maximum laccase activity for P. ostreatus strain CCMSSC 00322 from cottonseed hull (612.82 ± 13.16 U/L, 6th day) was higher than that from corncob (370.20 ± 13.51 U/L, 8th day) and poplar wood (371.81 ± 7.01 U/L, 8th day), by 1.66-fold and 1.65-fold, respectively (Table 2). Similarly, maximum laccase activity for P. ostreatus strain CCMSSC 00406 on cottonseed hull was 490.66 ± 11.87 U/L on 7th day, which was higher than that from corncob (156.62 ± 3.17 U/L, 7th day) and poplar wood (270.44 ± 5.98 U/L, 8th day), by 1.48-fold and 1.53-fold, respectively (Table 2). It was easy to see that laccase activity of four P. ostreatus strains on cottonseed hull was higher than that of the substrate of poplar wood and corncob. Meanwhile, the laccase activity of P. ostreatus CCEF 89 and CY 568 obtained from corncob was higher than that from poplar wood. The laccase activity of P. ostreatus CCMSSC 00322 and CCMSSC 00406 from corncob was lower than that from poplar wood.
A previous study showed that the inducement ability of wood chips and corncob to laccase production was different in different strains of P. ostreatus (Han et al. 2017). The maximum laccase activity for F. velutipes CCMSSC 00114 from cottonseed hull, corncob, and poplar wood was 79.67 ± 1.22 U/L on 6th day, 60.98 ± 1.49 U/L on 7th day and 45.61 ± 0.97 U/L on 7th day. The maximum laccase activities were up to 71.83 ± 0.35 U/L, 58.27 ± 1.14 U/L, and 42.50 ± 0.80 U/L for F. velutipes CCMSSC 00118 from substrates of cottonseed hull, corncob, and poplar wood, respectively (Table 2), the maximum values occurring after 6 d, 7 d, and 7 d, respectively. A previous study found that the laccase activity of F. velutipes CCMSSC 00118 was 10.1 U/L in the glucose-free CYM medium with 5% cottonseed hull (An et al. 2015). The laccase activity was lower than in this study. The maximum laccase activity for F. velutipes CCMSSC 05317 obtained from cottonseed hull was 70.42 ± 0.35 U/L, which was higher than that obtained from corncob (55.96 ± 1.22 U/L) and poplar wood (40.18 ± 0.46 U/L), by 1.26-fold and 1.75-fold, respectively (Table 2).
The time of maximum laccase activity for strain CCMSSC 05317 from cottonseed hull, corncob, and poplar wood was 6th day, 7th day and 7th day (Table 2). Maximum laccase activity for F. velutipes CCMSSC 05331 from cottonseed hull, corncob, and poplar wood was 68.11 ± 1.09 U/L, 50.13 ± 3.68 U/L, and 39.98 ± 0.17 U/L, respectively (Table 2). The time of maximum laccase activity was 6th day, 7th day and 7th day, respectively (Table 2). Furthermore, the appearance times for maximum laccase activity of tested strains on cottonseed hull was earlier than that on corncob or poplar wood, except P. ostreatus CY 568 and CCMSSC 00406.
Elisashvili et al. (2008) reported that the laccase activity in tree leaves solid-state fermentation or in wheat straw solid-state fermentation by P. ostreatus 2175, P. ostreatus IBB8, P. ostreatus IBB108, and P. ostreatus 2191 has almost no difference. The maximum laccase activity with a powdered walnut shell as the inducer by F. trogii or Trametes versicolor is higher than with powdered wheat straw as the inducer in submerged fermentation (Birhanli and Yeşilada 2013). The laccase activity of P. ostreatus CCEF 99 and CY 568 on sawdust substrate was sometimes higher than that on corncob substrate, but sometimes lower than that on corncob substrate on solid-state fermentation (Han et al. 2020). All the above studies indicate that the lignin content is not directly proportional to the laccase production secreted of fungi induced by these lignocellulosic materials. A roughly similar phenomenon was found in this study.
Fig. 3. Effects of poplar wood on laccase production by tested P. ostreatus and F. velutipes strains. Average values were calculated from individual measurements for each of three parallel cultures of eight strains.
Table 2. Maximum Laccase Activities, Lignocellulosic Material, Strains, and Time of tested P. ostreatus and F. velutipes Strains
Effects of Strains on Production of Laccase Enzymes
Laccase is necessary for lignin degradation by white-rot fungi such as Ganoderma lucidum, Flammulina velutipes, Fomes fomentarius, and Pleurotus ostreatus (Badalyan and Sakeyan 2004; An et al. 2016a; Han et al. 2020). Laccase production is significantly affected by different species or different strains of the same species (Janusz et al. 2015; An et al. 2016a, 2018; Huang et al. 2019; Han et al. 2020). Analysis the capacity of laccase production from different species or strains is helpful to obtain strains with high yield of laccase and provide more valuable strains for industrial production. In this study, the effect of the strains on laccase activities of tested P. ostreatus and F. velutipes strains was significant (P < 0.001) throughout the fermentation stage (Table 1). Eight strains belonging to P. ostreatus and F. velutipes showed their unique ability of secreting laccase (Figs. 1, 2 and 3). On the whole, the laccase secretion ability of P. ostreatus strains was stronger than that of F. velutipes strains in this study.
Laccase activity was detected in almost all tested P. ostreatus and F. velutipes strains on cottonseed hull on the 1st day, except P. ostreatus CCMSSC 00406 (Fig. 1). Laccase secreted by P. ostreatus strains exceeded 35 U/L on the first day, except for strain CCMSSC 00406 (Fig. 1), while production of laccase secreted by F. velutipes strains were not exceeding 7 U/L on the first day. The laccase activity of four F. velutipes strains was not high throughout the fermentation stage, and the trend of laccase expression was similar (Fig. 1). The maximum laccase activity for P. ostreatus CY 568 on cottonseed hull was nearly 1.21-fold, 1.47-fold, 1.84-fold, 11.33-fold, 12.57-fold, 12.82-fold, and 13.26-fold than that for P. ostreatus CCEF 89, P. ostreatus CCMSSC 00322, P. ostreatus CCMSSC 00406, F. velutipes CCMSSC 00114, F. velutipes CCMSSC 00118, F. velutipes CCMSSC 05317 and F. velutipes CCMSSC 05331, respectively. Different from the substrate of cottonseed hull, only four strains on corncob demonstrated laccase activity on the first day, and the activity was lower than that under the condition of cottonseed hull (Fig. 2). The maximum laccase activity for P. ostreatus CCEF 89 on corncob was nearly 1.14-fold, 1.89-fold, 4.46-fold, 11.46-fold, 12.00-fold, 12.49-fold, and 13.95-fold than that for P. ostreatus CY 568, P. ostreatus CCMSSC 00322, P. ostreatus CCMSSC 00406, F. velutipes CCMSSC 00114, F. velutipes CCMSSC 00118, F. velutipes CCMSSC 05317 and F. velutipes CCMSSC 05331, respectively. The trend of laccase expression for four F. velutipes strains was similar (Fig. 2). Different from the substrate of cottonseed hull and corncob, only two strains showed laccase activity on poplar wood on the first day, and the activity was lower than that under the condition of cottonseed hull and corncob. In particular, some strains, such as F. velutipes CCMSSC 05331 and 05317, did not exhibit laccase activity until day 4 (Fig. 3). The maximum laccase activity for P. ostreatus CY 568 on poplar wood was nearly 1.16-fold, 1.59-fold, 2.18-fold, 12.95-fold, 13.90-fold, 14.70-fold, and 14.78-fold that for P. ostreatus CCEF 89, P. ostreatus CCMSSC 00322, P. ostreatus CCMSSC 00406, F. velutipes CCMSSC 00114, F. velutipes CCMSSC 00118, F. velutipes CCMSSC 05317, and F. velutipes CCMSSC 05331, respectively. The trend of laccase expression for four F. velutipes strains was similar (Fig. 3). Previous studies about F. velutipes were mainly focused on the effects of metal ions and aromatic compounds on laccase (Janusz et al. 2015; An et al. 2016a). Previous studies reported the laccase activity of F. velutipes under lignocellulosic materials with simple carbon and nitrogen sources (An et al. 2015). The lignocellulosic material used for studying laccase production by F. velutipes was ramie stalk (Xie et al. 2017). This study was the first to compare the laccase activity of different strains of F. velutipes in submerged fermentation of lignocellulosic wastes of different composition. A previous study indicated that continuous and stable laccase production was an extremely important advantage of solid-state fermentation with poplar sawdust (Han et al. 2020). Unfortunately, no stable and continuous laccase activity was found on substrate of poplar sawdust under submerged fermentation. A previous study showed that cottonseed hull combined with simple carbon and nitrogen sources were more effective than corncobs and wood chips combined with simple carbon and nitrogen sources (Han et al. 2017). The result of this study also confirmed that laccase production of fungi cultured by cottonseed hull with deionized water was better than that by poplar wood or corncob with deionized water.
CONCLUSIONS
- This study indicated that the biosynthetic potential of basidiomycetes was highly dependent on the type of fungi and the lignocellulosic substrate. Different species of fungi or different strains of the same fungus had a preference of lignocellulosic materials to some extent.
- On the whole, the laccase secretion ability of P. ostreatus strains was superior to F. velutipes strains. The maximum laccase production of four P. ostreatus strains on cottonseed hull, corncob, and poplar wood was much higher than that of four F. velutipes strains on the same lignocellulosic waste.
- The presence of cottonseed hull was conducive to secreting laccase by P. ostreatus and F. velutipes strains in submerged fermentation. Maximum laccase production of tested P. ostreatus and F. velutipes strains on cottonseed hull was nearly 1.29-fold and 1.53-fold higher than that on corncob and poplar wood, respectively.
- The presence of cottonseed hull was helpful for accelerating the rate of enzyme production. Laccase activity was detected in almost all tested P. ostreatus and F. velutipes strains on cottonseed hull on the first day, while only a few strains on poplar wood and corncob could be detected on the first day.
ACKNOWLEDGMENTS
This research was supported by the National Natural Science Foundation of China (31700024), the Fundamental Research Funds for the Universities in Hebei Province (JYQ201901), the Fundamental Research Funds for the Universities in Hebei Province (JQ201905), Science Technology Research and Guidance Project of Colleges and Universities in Hebei Province (Z2019001), and Funds for Undergraduate Innovation and Entrepreneurship Training Project in Langfang Normal University (S202010100020).
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Article submitted: July 22, 2020; Peer review completed: August 29, 2020; Revised version received: September 1, 2020; Accepted: September 2, 2020; Published: October 16, 2020.
DOI: 10.15376/biores.15.4.9166-9179