Extracellular laccase activity among Ganoderma and Coriolopsis species grown on lignocellulosic wastes

Abstract

The extensive application of laccase in various fields depends on the supply of a large amount of laccase with high activity and low cost. Interest on the screening of fungal strains suitable for obtaining massive high-activity laccase in solid-state fermentation with suitable low-cost lignocellulosic wastes has increased. The present work determined the laccase secretion from different species belonging to genera Ganoderma and Coriolopsis fermented on different lignocellulosic wastes. Maximal laccase activity was obtained from Ganoderma lingzhi Han 1345 grown on rice straw for 9 days and indicated the capacity of secreting laccase of G. lingzhi Han 1345 was superior to that of Coriolopsis trogii Han 1211, C. strumosa Han 1356, and G. applanatum Han 1578. The presence of cottonseed hull was more favorable for Coriolopsis species secreting laccase, while Ganoderma species were more preferred to secrete laccase on rice straw. Further, the consistent substrate preference in laccase production of different species in the same genus was first exhibited. The results are useful to screen and obtain new species with superior capacity of secreting laccase and suitable low-cost lignocellulosic wastes as fermentation substrate for further industrial application of laccase.

Full Article

Extracellular Laccase Activity among Ganoderma and Coriolopsis Species Grown on Lignocellulosic Wastes

Mei-Ling Han,^a,# Jing Yang,^a,# Jian-Jun Ma,^a,b Chun-Rui Wang,^a Si-Yu Chen, ^a

Ming-Xin Xu,^a Qi-Yue Yang,^a Lu-Sen Bian,^c Xun-You Yan,^a,d and Qi An ^a,d,*

The extensive application of laccase in various fields depends on the supply of a large amount of laccase with high activity and low cost. Interest on the screening of fungal strains suitable for obtaining massive high-activity laccase in solid-state fermentation with suitable low-cost lignocellulosic wastes has increased. The present work determined the laccase secretion from different species belonging to genera Ganoderma and Coriolopsis fermented on different lignocellulosic wastes. Maximal laccase activity was obtained from Ganoderma lingzhi Han 1345 grown on rice straw for 9 days, and indicated the capacity of secreting laccase of G. lingzhi Han 1345 was superior to that of Coriolopsis trogii Han 1211, C. strumosa Han 1356, and G. applanatum Han 1578. The presence of cottonseed hull was more favorable for Coriolopsis species secreting laccase, while Ganoderma species were more preferred to secrete laccase on rice straw. Further, the consistent substrate preference in laccase production of different species in the same genus was first exhibited. The results are useful to screen and obtain new species with superior capacity of secreting laccase and suitable low-cost lignocellulosic wastes as fermentation substrate for further industrial application of laccase.

DOI: 10.15376/biores.17.3.5049-5064

Keywords: Extracellular laccase activity; Ganoderma; Coriolopsis; Lignocellulosic wastes

Contact information: a: College of Life Science, Langfang Normal University, Langfang 065000, Hebei, China; b: Forestry Engineering and Biodiversity Conservation Technology Research and Development Center in Langfang City, Langfang 065000, Hebei, China; c: Experimental Centre of Forestry in North China, Chinese Academy of Forestry, Beijing 102300, China; d: Technical Innovation Center for Utilization of Edible and Medicinal Fungi in Hebei Province, Langfang 065000, Hebei, China; # Mei-Ling Han and Jing Yang contributed equally to this article; * Corresponding author: fungiqian@163.com

INTRODUCTION

Lignocellulosic biomass, considered as the most abundant and renewable resource on earth, is mainly composed of cellulose (30 to 50% of dry matter weight), hemicellulose (20 to 40% of dry matter weight), and lignin (15 to 25% of dry matter weight), in addition to a small amount of structural proteins, lipids, and ash. Certainly, a part of it is usually considered as wastes, like corncob, leaves, sweet sorghum bagasse, coffee shell, corn stover, wheat straw, and sugar cane (Zhou et al. 2014; Sun et al. 2015; Moreira et al. 2016; Huang et al. 2019; Karagoz et al. 2019; Kumar et al. 2021). Generally, the main way of disposing of lignocellulosic wastes is to discard it to rot or burn it directly. Lignocellulose contains a variety of structural units, such as five-carbon sugars, six-carbon sugars, and aromatic compounds. The chemical diversity of these structural components allows production of different chemical products from lignocellulose. Another application, of course, is to grow fungi to produce various types of bioactive substances (Atilano-Camino et al. 2020; Gaikwad and Meshram 2020; Backes et al. 2022). Laccase is a biological active substance that has been widely studied by researchers because it is an ancient enzyme and has many applications (Unuofin et al. 2019; Zhou et al. 2021; Khatami et al. 2022).

Laccase (benzenediol: oxygen oxidoreductases, EC 1.10.3.2) belonging to a group of enzymes called blue multi-copper oxidases, is capable of oxidizing phenols and aromatic amines by reducing molecular oxygen to water (Hou et al. 2004). Laccase is widely distributed in nature, such as various plants, fungi, bacteria, and some insects (Unuofin et al. 2019; Zerva et al. 2019; Khatami et al. 2022). Laccase is one of the most important ligninolytic enzymes involved in lignin degradation (An et al. 2021a,b). Recent studies have indicated that laccases display broad substrate specificity and play an important role in the global carbon cycle (Wang et al. 2019; Teigiserova et al. 2021). Moreover, laccases are applied in many biotechnological processes and applications, including biodegradation, bioremediation, biosensors, biopharmaceuticals, decolorization and detoxification of textile effluents, biobleaching, and biopulping of paper industry, food chemistry and biofuels (Minussi et al. 2002; Sharma et al. 2019; Wang et al. 2019; Huang et al. 2020; Coria-Oriundo et al. 2021; Shokri et al. 2021; Sun et al. 2021a; Backes et al. 2022). Certainly, low-cost and high-activity are important characteristics for wide application of laccase in various aspects of industrial and environmental technology areas. Thus, obtaining low-cost laccase with high activity and high yield has attracted the attention of many researchers (Akpinar and Urek 2017; Chenthamarakshan et al. 2017).

White-rot fungi (Basidiomycetes) are of great interest because of their excellent laccase producing ability. Ganoderma and Coriolopsis spp. are white-rot fungi belonging to the family Polyporaceae. They typically grow on angiosperm wood and fallen angiosperm wood, respectively. Additionally, Ganoderma and Coriolopsis spp. are efficient lignin-degrading fungi that are able to colonize different lignocellulosic wastes as substrates. Submerged fermentation is suitable for fungi-producing laccase in an enzyme preparation factory due to the convenience of controlling fermentation conditions, while the main disadvantage of submerged fermentation is that the enzyme is diluted due to high water content. The method of solid-state fermentation (SSF) contains little or no free water, avoids the problem of enzyme dilution, and is more similar to the habitat of fungi living in the wild (Steudler and Bley 2015). The SSF is typically lignocellulosic wastes that contain rich nutrients for microorganism growth and enzyme production (Hatvani and Mécs 2001). Currently, numerous types of lignocellulosic wastes, e.g., sugarcane bagasse, wheat bran, oil palm trunk, rice husks, and straw, have been used for producing enzymes under solid-state fermentation (Singh et al. 2013; Karp et al. 2015; Thakur and Gupte 2015; Postemsky et al. 2017; Soccol et al. 2017; An et al. 2020; Han et al. 2021a, 2021c). Consequently, screening of the appropriate fungal species and lignocellulosic wastes for producing laccase is advisable. In the present study, four newly isolated strains of the wild white-rot fungi were obtained from their native habitats in northern and central China and examined for their ability to produce laccase. Furthermore, the capacity of secreting laccase of fungal strains collected from Central China and Northern China was also investigated for the first time. Four kinds of lignocellulosic wastes, Populus beijingensis, rice straw, cottonseed hull and corncob, were used as the only source of carbon, nitrogen, and mineral elements for fungi grown. The new isolated strains had the potential to serve as the alternative laccase-producing fungi, and lay a foundation for the optimization of the fermentation conditions for obtaining low cost laccase with high activity and the industrial application of laccase.

EXPERIMENTAL

Materials

Microorganisms

Two Ganoderma sp. strains Han 1345 and Han 1578, and two Coriolopsis sp. strains Han 1211 and Han 1356 were used in the present study. Among them, Han 1345 and Han 1578 were collected in Ma’an Mountain Forest Park of Hubei Province of central China and Wulingshan National Nature Reserve of Hebei Province of northern China, respectively. Another two strains, Han 1211 and Han 1356 were collected from Langfang Normal University of Hebei Province of northern China and the Wuhan Botanical Garden of the Chinese Academy of Sciences, Hubei Province, China, respectively. All strains were isolated and purified on complete yeast medium (CYM), which was composed of 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 15 g/L. All strains were maintained in 2% (w/v) malt extract agar (MEA) plates at the Experimental Centre of Forestry in North China, Chinese Academy of Forestry.

Lignocellulosic wastes

Lignocellulosic wastes used in the present study were Populus beijingensis, rice straw, cottonseed hull, and corncob. Among them, Populus beijingensis was obtained from Langfang City, while rice straw, cottonseed hull, and corncob were kindly provided by farmers from Chengde City. To obtain lignocellulosic particles with particle size of 20- to 60-mesh, all lignocellulosic wastes were air-dried, then ground.

Methods

Culture conditions of fungi and inoculum preparation

A liquid medium contained per liter: 20 g glucose, 2 g peptone, 2 g yeast extract, 0.5 g MgSO₄·7H₂O, 1 g K₂HPO₄·3H₂O, and 0.46 g KH₂PO₄, was used for growing mycelium pellets of fungi. All cultures were inoculated with five mycelial plugs (5 mm diameter) taken from the periphery of a colony grown on CYM at 26 °C for 7 days. The cultures were incubated at 26 °C for 7 days on a rotary shaker at 150 rpm. Then, mycelial suspensions were prepared with modular homogenizer S10 (Ningbo Xinzhi Biotechnology Co., Ltd., Ningbo, China) to homogenize under 8,000 rpm for 3 min. These homogenized mycelial suspensions were used as the inoculum.

Solid-state fermentation for laccase production

For the production of laccase through SSF, two Ganoderma sp. strains Han 1345 and Han 1578, and two Coriolopsis sp. strains Han 1211 and Han 1356 were inoculated separately on the substrate containing single lignocellulosic waste. A total of 3 g of lignocellulosic waste was taken into a 250-mL flasks and 9 mL of deionized water was added to moisten the lignocellulosic waste and then autoclaved (121 °C, 30 min). Then, 3 mL of homogenized inoculum was added into each Erlenmeyer flask and kept for 1 to 10 days at 26 °C under constant agitation at 150 rpm.

Sampling, extraction and assay of laccase

To obtain the crude enzyme, 100 mL of acetate-sodium acetate buffer (50 mM, pH 5.5) was added to solid lignocellulosic wastes. Then, all flasks were transferred and agitated on a rotary shaker at 120 rpm for 4 h at 10 °C (An et al. 2021; Han et al. 2021a,c). Cell free supernatants, considered as a crude laccase, were obtained by filtering the liquid containing lignocellulosic waste through a filter paper and centrifuging the culture liquid at 4 °C, 12,000 rpm for 20 min. Supernatant obtained after centrifugation was determined for the laccase.

Quantitative estimation of laccase activity was determined by observing the change of substrate in absorbance at 415 nm (Ɛ₄₁₅ = 3.16 × 10⁴ M^-1 cm^-1). The substrate used for laccase was 2,2-azino-bis-[3-ethylbenzthiazoline-6-sulfonate] (ABTS). Laccase activity was assessed by the reaction mixture containing 1 mM ABTS and 50 mM acetate-sodium acetate buffer (pH 4.2) (Han et al. 2021a, 2021c) and detected by an iMark^TM microplate absorbance reader (Bio-Rad, Hercules, CA, USA). One unit of laccase activity was defined as the amount of crude enzyme required to transform 1 µmol of ABTS per minute into product.

Statistical analysis

The presented data was exhibited as the mean ± standard error (SE) of three replicates (n = 3). Significant differences in laccase activity between the lignocellulosic wastes and fungi were assessed using two-way analysis of variance (ANOVA) with Tukey’s honest significance difference (HSD) post hoc test by SPSS 22.0 (IBM Corp., version 22.0, Armonk, NY, USA).

Identification of the Ganoderma sp. strains and Coriolopsis sp. strains

Ganoderma sp. strains and Coriolopsis sp. strains were identified by amplification of the ITS rRNA gene. The mycelium was scraped with a surgical blade from the surface of the Petri dishes cultivated Ganoderma sp. strains and Coriolopsis sp. strains for 7 days. Then, the total genomic DNA was extracted using the cetyltrimethylammonium bromide (CTAB) rapid plant genome extraction kit-DN14 (Aidlab Biotechnologies Co., Ltd., Beijing, China) according to the instructions (Han et al. 2016, 2021b). ITS5 and ITS4 primers were used for polymerase chain reaction (PCR) amplification in a thermocycler. Then, the PCR products were sent to the Beijing Genomics Institute (Beijing, China) for sequencing. The obtained sequence was queried against ITS sequences available in GenBank, using BLAST. Finally, all new ITS sequences of Han 1211, Han 1345, Han 1356, and Han 1578 were submitted to GenBank and corresponding returned GenBank numbers were ON262337, ON262338, ON262339, and ON262340, respectively.

RESULTS AND DISCUSSION

Identification of the Selected Fungal Strain

The taxonomic status of strains Han 1211, Han 1345, Han 1356, and Han 1578 identified by ITS sequence was Coriolopsis trogii, Ganoderma lingzhi, Coriolopsis strumosa, and Ganoderma applanatum, respectively.

Statistical Analysis

As shown in Table 1, it was not difficult to find that species, lignocellulosic wastes, and the interaction of species, and lignocellulosic wastes were significantly affecting the laccase activity (P < 0.001) during the whole fermentation process (Table 1).

Table 1. Two-way ANOVA to Analyze Effects of Different Species, Lignocellulosic Wastes, and the Interactions of Different Species and Lignocellulosic Wastes on Laccase Activity

Effect of Different Lignocellulosic Wastes on Laccase Activity

Certain lignocellulosic wastes contain natural inducers that have the potential ability to enhance laccase productivity, diminish the cost of production, and reduce pollution in solid-state fermentation (Wang et al. 2019). To select lignocellulosic wastes for producing laccase by SSF, lignocellulosic wastes should be a suitable support for fungi growth. There have been few studies on laccase production of the genus Coriolopsis grown on lignocellulosic wastes that indicate newly isolated Coriolopsis sanguinaria belonging to genus of Coriolopsis was an excellent laccase-producing strain (An et al. 2021b). Hence, assessment the effect of different lignocellulosic wastes to different newly isolated fungi strains secreting laccase was a key and sustainable task.

The laccase activity of Coriolopsis trogii Han 1211 stimulated by cottonseed hull, rice straw, corncob, and Populus beijingensis at the 1^st day was 28.03 ± 2.09 U/L, 8.34 ± 0.92 U/L, 1.61 ± 0.17 U/L, and 7.84 ± 0.52 U/L, respectively (Fig. 1). Clearly, the existence of cottonseed hull was conducive to accelerate the production of laccase by C. trogii Han 1211. Similarly, An et al. (2020) compared the laccase activity of different Pleurotus ostreatus and Flammulina velutipes strains on poplar wood, corncob, and cottonseed hull, and indicated that the presence of cottonseed hull was beneficial to secrete laccase via testing P. ostreatus and F. velutipes strains in submerged fermentation. Maximum laccase activity of C. trogii Han 1211 grown on cottonseed hull, rice straw, corncob, and Populus beijingensis was 223.03 ± 11.51 U/L at 8^th day, 118.24 ± 3.48 U/L at 3^rd day, 151.50 ± 3.32 U/L at 3^rd day, and 75.85 ± 3.62 U/L at 8^th day (Table 2). Thus, the presence of cottonseed hull was helpful for improving laccase activity of C. trogii Han 1211 and the existence of Populus beijingensis was disadvantaged for C. trogii Han 1211 secreting laccase. The trend of laccase activity of C. trogii Han 1211 on cottonseed hull was difference from that on rice straw, corncob, and Populus beijingensis due to the peak number of laccase activity (Fig. 1). In contrast, rice straw and corncob contributed to produce laccase in a short time, although the highest laccase activities on rice straw and corncob were not the highest of the four tested lignocellulosic wastes. Furthermore, the most cost-effective method of producing laccase using C. Trogii Han 1211 on the four lignocellulosic wastes was to perform the fermentation process on corn cob for 3 days.

Fig. 1. Time course of laccase activity of Coriolopsis trogii Han 1211 grown on cottonseed hull, rice straw, corncob, and Populus beijingensis by solid-state fermentation

Table 2. Maximum Laccase Activity and Corresponding Occurrence Time of Coriolopsis sp. and Ganoderma sp. Strains on Different Lignocellulosic Wastes

At the 1^st day, laccase activity of Ganoderma lingzhi Han 1345 on cottonseed hull, rice straw, corncob, and Populus beijingensis was 7.03 ± 0.63 U/L, 0 ± 0 U/L, 0.30 ± 0 U/L, and 13.66 ± 0.46 U/L, respectively (Fig. 2). Therefore, Populus beijingensis was helpful for improving laccase activity at the beginning of fermentation. Maximum level of laccase activity of G. lingzhi Han 1345 on rice straw was 745.83 ± 8.99 U/L at the 9^th day, nearly 8.87-fold, 2.23-fold, and 6.26-fold higher than that on cottonseed hull (84.09 ± 7.69 U/L, at 5^th day), corncob (333.74 ± 11.41 U/L, at 8^th day), and Populus beijingensis (119.05 ± 4.98 U/L, at 4^th day) (Table 2). Clearly, the presence of rice straw was conducive to improving laccase activity of G. lingzhi Han 1345, but it was not helpful for obtaining high laccase activity in a shorten time. At the end of solid-state fermentation, the laccase activity of G. lingzhi Han 1345 on rice straw and corncob was 573.34 ± 17.33 U/L and 212.48 ± 13.39 U/L, while the laccase activity detected on cottonseed hull and Populus beijingensis was 50.53 ± 2.18 U/L and 0 ± 0 U/L. This showed G. lingzhi Han 1345 could secret stable and high-level activity of laccase when it was stimulated by rice straw and corncob (Fig. 2). An et al. (2021a) evaluated the laccase activity of G. lingzhi Han 500 on cottonseed hull, Toona sinensis, Sophora japonica, Salix babylonica, and straw of Oryza sativa, and found that the maximum laccase activity was 788.73 ± 6.12 U/L measured on cottonseed hull at 8^th day. However, the maximum laccase activity of G. lingzhi Han 1345 on cottonseed hull was merely 84.09 ± 7.69 U/L at 5^th day in the present work. Actually, G. lingzhi Han 1345 was more preferred to produce laccase on rice straw. This also reflects the specificity of enzyme production and substrate preference of different strains belonging to one species. Of course, G. lingzhi Han 500 in the previous study was collected from northern China and G. lingzhi Han 1345 in the present study was collected from central China. The difference of habitat environment between central China and northern China may also be one of the reasons for the difference in enzyme production and substrate preference.

Fig. 2. Time course of laccase activity of Ganoderma lingzhi Han 1345 grown on cottonseed hull, rice straw, corncob, and Populus beijingensis by solid-state fermentation

No laccase activity of Coriolopsis strumosa Han 1356 was detected in all four tested lignocellulosic wastes on the first day (Fig. 3). Laccase activity on cottonseed hull, rice straw, corncob, and Populus beijingensis was first detected on day 2 (9.24 ± 0.76 U/L), day 3 (40.18 ± 0.76 U/L), day 3 (16.68 ± 1.77 U/L), and day 2 (42.70 ± 3.60 U/L). Based on this, it seems likely that Populus beijingensis was suitable for C. strumosa Han 1356 secreting laccase. However, maximum laccase activity detected on cottonseed hull was 203.64 ± 5.89 U/L for 5 days, nearly 1.66-fold, 1.74-fold, and 3.39-fold higher than that on rice straw (122.51 ± 8.21 U/L, 7 days), corncob (116.94 ± 2.09 U/L, 6 days), and Populus beijingensis (60.08 ± 2.60 U/L, 3 days), respectively (Table 2). Clearly, the presence of cottonseed hull was conducive to improve laccase activity of C. strumosa Han 1356 in a shorten time.

Fig. 3. Time course of laccase activity of Coriolopsis strumosa Han 1356 grown on cottonseed hull, rice straw, corncob, and Populus beijingensis by solid-state fermentation

At the 1^st day, laccase activity of Ganoderma applanatum Han 1578 on cottonseed hull was 19.09 ± 0.76 U/L, and was undetected on rice straw, corncob, and Populus beijingensis (Fig. 4). Maximum laccase activity from cottonseed hull was 41.09 ± 3.60 U/L at the 2^nd day, nearly 7.30-fold, 9.30-fold, and 9.30-fold on rice straw (5.63 ± 0.35 U/L), corncob (4.42 ± 0.46 U/L), and Populus beijingensis (4.42 ± 0.17 U/L), respectively. To some extent, cottonseed hull was truly effective in promoting rapid production of laccase from G. applanatum Han 1578. Maximum level of laccase activity on rice straw, corncob, and Populus beijingensis and was 53.04 ± 1.31 U/L at 8^th day, 35.16 ± 2.93 U/L at 6^th day, and 14.67 ± 0.63 U/L at 7th day (Table 2). Furthermore, the laccase activity detected on cottonseed hull, rice straw, corncob, and Populus beijingensis was 1.61 ± 0.17 U/L, 33.96 ± 1.49 U/L, 14.77 ± 0.60 U/L, and 4.52 ± 0.30 U/L. Thus, rice straw was conducive to improve laccase activity of G. applanatum Han 1578, but it was not helpful for obtaining high laccase activity in a short time.

Fig. 4. Time course of laccase activity of Ganoderma applanatum Han 1578 grown on cottonseed hull, rice straw, corncob, and Populus beijingensis by solid-state fermentation

Effect of Fungi in Same or Different Genus on Laccase Activity

Previous study has shown that the laccase activity of fungi can vary greatly under identical culture conditions among both species and strains of the same species (Elisashvili et al. 2008). Commonly, species is a classification noun, but strains can be used to refer to fungi of the same or different species. White-rot fungi (species or strains of Basidiomycota) have the ability to degrade lignocellulosic wastes with high efficiency because they could secret various extracellular enzymes (Kuhar et al. 2015; Rodrigues et al. 2019). Recently, there has been increased research attention on screening fungal or bacterial strains suitable for obtaining massive high-activity laccase in solid-state fermentation (Al-balawi et al. 2017; Sun et al. 2017; Boruah et al. 2019; Wang et al. 2019; Yasar et al. 2019; Dullah et al. 2021; Sun et al. 2021b). In recent years, novel fungi with the ability to produce ligninolytic enzymes have been isolated from different natural habitats. Meanwhile, the capacity of secreting laccase from different species belonging to the same genus was compared. Huang et al. (2019) compared the differences in degrading crop straw from different edible fungi due to their differences in the ability of secreting lignocellulolytic enzymes. An et al. (2021a) analyzed the laccase activity of Pleurotus ostreatus CY 568 and Ganoderma lingzhi Han 500, and found that the laccase activity of newly isolated G. lingzhi Han 500 was higher than that of other Ganoderma strains in previous studies. Research on laccase of genus Coriolopsis has rarely been reported, and the only relevant studies focused on recombinant expression and heterologous expression of laccase (Songulashvili et al. 2016; Avelar et al. 2017; Glazunova et al. 2018). Based on this, screening new strains with the ability of producing laccase enzyme is of great value for obtaining high activity and high yield of laccase. Furthermore, the laccase production of fungal strains collected from central China and northern China was also investigated for the first time.

Laccase activity of Coriolopsis trogii Han 1211, Ganoderma lingzhi Han 1345, Coriolopsis strumosa Han 1356, and Ganoderma applanatum Han 1578 detected on cottonseed hull was 28.03 ± 2.09 U/L, 7.03 ± 2.09 U/L, 0 ± 0 U/L, and 19.09 ± 0.76 U/L at the first day (Figs. 1 through 4). Similarly, the value of laccase activity from C. trogii Han 1211, G. lingzhi Han 1345, C. strumosa Han 1356, and G. applanatum Han 1578 on rice straw was 8.34 ± 0.92 U/L, 0 ± 0 U/L, 0 ± 0 U/L, and 0 ± 0 U/L for the first day. Laccase activity of C. trogii Han 1211, G. lingzhi Han 1345, C. strumosa Han 1356, and G. applanatum Han 1578 on corncob was 1.61 ± 0.17 U/L, 0.30 ± 0 U/L, 0 ± 0 U/L, and 0 ± 0 U/L at the 1^st day. Based on the above description, the capacity of secreting laccase of C. trogii Han 1211 was superior to other three strains in the early stages of fermentation. However, laccase activity of C. trogii Han 1211, G. lingzhi Han 1345, C. strumosa Han 1356, and G. applanatum Han 1578 on Populus beijingensis was 7.84 ± 0.52 U/L, 13.66 ± 0.46 U/L, 0 ± 0 U/L, and 0 ± 0 U/L. Thus, G. lingzhi Han 1345 was more in favor of secreting laccase than other tested strains when Populus beijingensis as a substrate at the beginning of fermentation. In terms of laccase activity on cottonseed hull, the maximum value from C. trogii Han 1211 was 223.03 ± 11.51 U/L for the 8^th day, nearly 2.65-fold, 1.10-fold, and 5.43-fold higher than that from G. lingzhi Han 1345 (84.09 ± 7.69 U/L, for 5^th day), C. strumosa Han 1356 (203.64 ± 5.89 U/L, for 5^th day), and G. applanatum Han 1578 (41.09 ± 3.60 U/L, for 2^th day), respectively. Furthermore, C. trogii Han 1211 and C. strumosa Han 1356 belonged to the genus of Coriolopsis. Hence, the presence of cottonseed hull was conducive to secret laccase of the genus of Coriolopsis (Figs. 1 through 4). Maximum laccase activity of G. lingzhi Han 1345 on rice straw was 745.83 ± 9.00 U/L at the 9^th day, nearly 6.31-fold, 6.09-fold, and 14.06-fold higher than that from C. trogii Han 1211 (118.24 ± 3.48 U/L, 3^rd day), C. strumosa Han 1356 (122.51 ± 8.21 U/L, 7^th day), and G. applanatum Han 1578 (53.04 ± 1.31 U/L, 8^th day). Clearly, rice straw was helpful for G. lingzhi Han 1345 secreting laccase. The value of laccase activity obtained from C. trogii Han 1211 and C. strumosa Han 1356 was similar, but C. trogii Han 1211 reached the maximum laccase activity faster. In terms of laccase activity on corncob, maximum laccase activity of C. trogii Han 1211, G. lingzhi Han 1345, C. strumosa Han 1356, and G. applanatum Han 1578 was 151.50 ± 3.32 U/L for 3^rd day, 333.74 ± 11.41 U/L for 8^th day, 116.94 ± 2.09 U/L for 6^th day, and 35.16 ± 2.93 U/L for 6^th day. The phenomenon of laccase activity was similar to that on rice straw. In other words, corncob was helpful for G. lingzhi Han 1345 secreting laccase, and the maximum value of laccase activity obtained from C. trogii Han 1211 was close to that from C. strumosa Han 1356. Maximum laccase activity from C. trogii Han 1211, G. lingzhi Han 1345, C. strumosa Han 1356, and G. applanatum Han 1578 on Populus beijingensis was 75.85 ± 3.62 U/L (8^th day), 119.05 ± 4.98 U/L (4^th day), 60.08 ± 2.60 U/L (3^rd day), and 14.67 ± 0.63 U/L (7^th day). Thus, the ability of secreting laccase of G. lingzhi Han 1345 was superior to other strains during fermentation on Populus beijingensis.

Qiu et al. (2014) investigated the effect of Kudzu vine root for Funalia trogii secreting laccase and found the laccase activity was 42.5 IU/g via solid-state fermentation with Kudzu vine root. The maximum value of laccase activity of Coriolopsis rigida was 3 × 10⁵ nkat/L via fermentation with barley bran and proved that C. rigida was an excellent producer for secreting laccase (Gomez et al. 2005; Alcantara et al. 2007). Maximum laccase activity of C. trogii Han 751 on stalk of Sorghum bicolor was 799.03 ± 40.89 U/L (An et al. 2021b). It was noteworthy that two strains of genus Coriolopsis, C. trogii Han 1211 and C. strumosa Han 1356, had a preference on cottonseed hull for secreting laccase. Additionally, two strains of genus Ganoderma, G. lingzhi Han 1345 and G. applanatum Han 1578, showed the consistency of secreting laccase on rice straw. An et al. (2016) analyzed the laccase activity from the Genus Flammulina (Agaricomycetes) and indicated different fungi preferred different substrate for secreting laccase. However, strains belonging to Ganoderma or Coriolopsis used in the present study showed a consistent preference for laccase secretion on lignocellulosic wastes. Maximum laccase production of G. lingzhi Han 500 on cottonseed hull was higher than that on Salix babylonica, corncob, Populus beijingensis, Toona sinensis, and straw of Oryza sativa (An et al. 2021a). While the maximum laccase of G. lingzhi Han 1345 and G. applanatum Han 1578 on rice straw was higher than that on cottonseed hull, it showed the consistency of secreting laccase on cottonseed hull, corncob, and Populus beijingensis. Overall, the capacity of secreting laccase of G. lingzhi Han 1345 was superior to that of C. trogii Han 1211, C. strumosa Han 1356, and G. applanatum Han 1578.

CONCLUSIONS

1. Different from previous studies, the present study found that two strains belonging to Ganoderma showed a consistent preference for laccase secretion in the presence of lignocellulosic wastes, and two strains belonging to Coriolopsis showed a consistent preference for laccase secretion with lignocellulosic wastes.
2. The existence of cottonseed hull was conducive to secret laccase of Coriolopsis trogii Han 1211 and C. strumosa Han 1356. Additionally, cottonseed hull was beneficial to improve laccase activity of C. strumosa Han 1356 in a short time.
3. Ganoderma lingzhi Han 1345 and G. applanatum Han 1578 were more preferred to secret laccase on rice straw.
4. The capacity of secreting laccase of G. lingzhi Han 1345 was superior to that of C. trogii Han 1211, C. strumosa Han 1356, and G. applanatum Han 1578.

ACKNOWLEDGMENTS

The authors gratefully acknowledge the financial support of the National Natural Science Foundation of China (31900009), Special Project of Cultivating Scientific and Technological Innovation Ability of College and Middle School Students (2021H100404), Special Project of Cultivating Scientific and Technological Innovation Ability of College and Middle School Students (S202110100006), the Fundamental Research Funds for the Universities in Hebei Province (JYQ202201 & JYQ201901), and the Top-notch Youth Project of Langfang City, China.

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Article submitted: April 19, 2022; Peer review completed: July 11, 2022; Revised version received and accepted: July 12, 2022; Published: July 13, 2022.

DOI: 10.15376/biores.17.3.5049-5064