Recovery of Proteases and Protease Inhibitors from Ganoderma spp. Cultivated in Amazonian Lignocellulose Wastes.

BACKGROUND: Ganoderma spp. are a great source of bioactive molecules. The production and recovery of bioactive molecules vary according to strain, growth substrate, and extraction solution. Variations in protease and their inhibitors in basidiomata from a commercial strain (G. lingzhi) and an Amazonian isolate (Ganoderma sp.) cultivated in Amazonian lignocellulosic wastes and extracted with different solutions are plausible and were investigated in our study. METHODS: Basidiomata from cultivation in substrates based on açaí seed, guaruba-cedro sawdust and three lots of marupá sawdust were submitted to extraction in water, Tris-HCl, and sodium phosphate. Protein content, proteases, and protease inhibitors were estimated through different assays. The samples were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Fourier transform infrared spectroscopy with attenuated total reflectance (FTIR-ATR). RESULTS: Tris-HCl provided higher protein extraction from Ganoderma sp. and higher caseinolytic, gelatinolytic, and fibrinolytic activity for G. lingzhi cultivated in açaí. Water extracts of Ganoderma sp., in general, exhibited higher trypsin and papain inhibitor activities compared to G. lingzhi. Extracts in Tris-HCl and sodium phosphate showed more intense protein bands in SDS-- PAGE, highlighting bands of molecular weights around 100, 50, and 30 kDa. FTIR spectra showed patterns for proteins in all extracts, with variation in transmittance according to substrate and extractor. CONCLUSION: Water extract from Amazonian Ganoderma sp. cultivated in marupá wastes are promising as a source of protease inhibitors, while the Tris-HCL extract of G. lingzhi from açaí cultivation stands out as a source of proteases with fibrinolytic, caseinolytic, and gelatinolytic activities.

Time-dependent change in Reishi (Ganoderma lingzhi) triterpenoids in culture with rumen fluid.

The purpose of this study was to investigate the time-dependent change in Reishi (Ganoderma lingzhi) triterpenoids in rumen fluid. G. lingzhi fruiting bodies were milled and incubated in a tube with rumen fluid for 0, 4, 8, 12, 24, and 48 h at 39°C. After incubation, all the tubes were freeze-dried and extracted by ethanol. The contents of 18 triterpenoids in the ethanol extract were quantitated by liquid chromatography-mass spectrometry (LC-MS/MS). Based on the results, triterpenoids were categorized into three groups: (1) rapid decrease, indicating reductions of more than 50% within 8 h; (2) mild decrease, with reductions of more than 50% within 48 h; and (3) minimal change, even after 48 h, there was not much change. Ganoderic acid C6, DM, H, K, and TR as well as Ganoderenic acid D were classified in (1); Ganoderic acid LM2 and T-Q as well as Ganoderiol F in (2); and Ganoderic acid A, B, C1, C2, I, and TN; Gnoderenic acid C; and Ganodermanontriol in (3). In addition, a relationship between chemical structure and metabolic speed was observed in some cases. The results of this study revealed that G. lingzhi triterpenoids are digested and metabolized at different speeds in ruminant fluid.

Ganonorsterone A, a norsteroid from the medicinal fungus Ganoderma lingzhi.

Phytochemical investigation on the fruiting bodies of the medicinal fungus Ganoderma lingzhi led to the isolation of a new norsteroid, namely ganonorsterone A (1), together with one known steroid, cyathisterol (2). The structure and absolute configuration of compound 1 were assigned by extensive analysis of MS, NMR data, and quantum-chemical calculations including electronic circular dichroism (ECD) and calculated 13C NMR-DP4+ analysis. Bioassay results showed that compound 1 displayed moderate inhibition on NO production in RAW 264.7 macrophages.

UPLC-ESI-MS/MS-based widely targeted metabolomics reveals differences in metabolite composition among four Ganoderma species.

The Chinese name "Lingzhi" refers to Ganoderma genus, which are increasingly used in the food and medical industries. Ganoderma species are often used interchangeably since the differences in their composition are not known. To find compositional metabolite differences among Ganoderma species, we conducted a widely targeted metabolomics analysis of four commonly used edible and medicinal Ganoderma species based on ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry. Through pairwise comparisons, we identified 575-764 significant differential metabolites among the species, most of which exhibited large fold differences. We screened and analyzed the composition and functionality of the advantageous metabolites in each species. Ganoderma lingzhi advantageous metabolites were mostly related to amino acids and derivatives, as well as terpenes, G. sinense to terpenes, and G. leucocontextum and G. tsugae to nucleotides and derivatives, alkaloids, and lipids. Network pharmacological analysis showed that SRC, GAPDH, TNF, and AKT1 were the key targets of high-degree advantage metabolites among the four Ganoderma species. Analysis of Gene Ontology and Kyoto Encyclopedia of Genes and Genomes demonstrated that the advantage metabolites in the four Ganoderma species may regulate and participate in signaling pathways associated with diverse cancers, Alzheimer's disease, and diabetes. Our findings contribute to more targeted development of Ganoderma products in the food and medical industries.

Determination of selenium-containing species, including nanoparticles, in selenium-enriched Lingzhi mushrooms.

Mushrooms are considered a valuable food source due to their high protein and fibre and low fat content, among the other health benefits of their consumption. Selenium is an essential nutrient and is renowned for its chemo-preventative properties. In this study, batches of selenium-enriched Lingzhi mushrooms were prepared by growing mycelium and fruit in substrates containing various concentrations of sodium selenite. The mushroom fruit accumulated low levels of selenium with selenomethionine being the most abundant form in all enriched samples. Conversely, the mycelium showed significant selenium accumulation but relatively low proportions of selenomethionine. The red colour of the selenium-enriched mycelia indicated the probable presence of selenium nanoparticles, which was confirmed by single-particle inductively coupled plasma-mass spectrometry. Mean particle diameters of 90-120 nm were observed, with size distributions of 60-250 nm. Additional analysis with transmission electron microscopy confirmed this size distribution and showed that the biogenic selenium nanoparticles were roughly spherical in shape and contained elemental selenium.

Application of Miscanthus substrates in the cultivation of Ganoderma lingzhi.

Ganoderma lingzhi is a traditional Chinese medicine that has been used to improve health and longevity for thousands of years. It is usually cultivated on hardwood log- or sawdust-based formulations. Conversely, in this study, we used Miscanthus sacchariflorus (MSF), M. floridulus, and M. sinensis (MSS), fast-growing perennial grasses widely distributed in China, for G. lingzhi cultivation. Mycelial growth rate, activities of lignin-degrading enzymes on colonized mushroom substrates, and expression levels of CAZymes and laccase genes based on different substrates were analyzed. Total triterpenoids, sterols, and polysaccharides content of fruiting bodies obtained from different substrates were investigated. The activities of laccase and manganese peroxidase in mycelia increased in the MSF- and MSS-based formulations compared with that in the sawdust-based formulation. The results of mycelial growth- and cultivation-related experiments showed that the Miscanthus substrates could be used as the substrates for cultivating G. lingzhi. The content of active ingredients, namely triterpenoids, sterols, and polysaccharides, in fruiting bodies cultivated on the Miscanthus substrates did not decrease compared with those in substrate obtained from the sawdust-based formulation. Therefore, the present study provides alternative substrates for the cultivation of G. lingzhi, and a reference for better utilization of inexpensive substrate in future.

Pharmacological Activities and Safety of Ganoderma lucidum Spores: A Systematic Review.

Ganoderma lucidum is traditionally used to prevent and treat some diseases such as liver disorders, hypertension, insomnia, diabetes, and cancer. G. lucidum spore extracts are also reported to share similar bioactivities as extracts from its other parts. However, there is no systematic review that elucidates its pharmacological effect. Our aim is to comprehensively summarise current evidence of G. lucidum spore extracts to clarify its benefits to be applied in further studies. We searched five primary databases: PubMed, Virtual Health Library (VHL), Global Health Library (GHL), System for Information on Grey Literature in Europe (SIGLE), and Google Scholar on September 13, 2021. Articles were selected according to inclusion and exclusion criteria. A manual search was applied to find more relevant articles. Ninety studies that reported the pharmacological effects and/or safety of G. lucidum spores were included in this review. The review found that G. lucidum spore extracts showed quite similar effects as other parts of this medicinal plant including anti-tumor, anti-inflammatory, antioxidant effects, and immunomodulation. G. lucidum sporoderm-broken extract demonstrated higher efficiency than unbroken spore extract. G. lucidum extracts also showed their effects on some genes responsible for the body's metabolism, which implied the benefits in metabolic diseases. The safety of G. lucidum should be investigated in depth as high doses of the extract could increase levels of cancer antigen (CA)72-4, despite no harmful effect shown on body organs. Generally, there is a lot of potential in the studies of compounds with pharmacological effects and new treatments. Sporoderm breaking technique could contribute to the production of extracts with more effective prevention and treatment of diseases. High doses of G. lucidum spore extract should be used with caution as there was a concern about the increase in CA.

Lanostane triterpenoids with anti-proliferative and anti-inflammatory activities from medicinal mushroom Ganoderma lingzhi.

Eight previously undescribed lanostane triterpenoids and nine known ones were identified from the fruiting bodies of Ganoderma lingzhi S.H. Wu, Y. Cao & Y.C. Dai. Their structures were determined based on spectroscopic data and quantum chemical calculations. Structurally, ganoderane GL-1, featuring a hydrogenated tetramethyls-phenanthraquinone, represents the first example in lanostane nor-triterpenoid group. Biologically, ganoderanes GL-2 and GL-3, distinguished by the presence of a rare "1,11-epoxy" moiety, exhibited significant inhibition against nitric oxide production induced by lipopolysaccharide in RAW264.7 macrophage cells, while ganoderanes GL-4 and GL-8 exhibited bifunctional activities of anti-proliferation and anti-inflammation.

Ganoderma lingzhi culture enhance growth performance via improvement of antioxidant activity and gut probiotic proliferation in Sanhuang broilers.

Introduction: The experiment was conducted to evaluate the effects of Ganoderma lingzhi culture (GLC) as a fermented feed on growth performance, serum biochemical profile, meat quality, and intestinal morphology and microbiota in Sanhuang broilers. In addition, the association between gut bacteria and metabolites was investigated via untargeted metabolomic analysis. Methods: A total of 192 Sanhuang broilers (112 days old) with an initial body weight of 1.62 ± 0.19 kg were randomly allocated to four treatments, six replicate pens per treatment with 8 broilers per pen. The four treatments contain a control diet (corn-soybean meal basal diet, CON), a positive control diet (basal diet + 75 mg/kg chlortetracycline, PCON), and the experimental diets supplemented with 1.5 and 3% of GLC, respectively. The trial includes phase 1 (day 1-28) and phase 2 (day 29-56). Results: The results showed that broilers in PCON and GLC-added treatments showed a lower FCR (P < 0.05) in phase 2 and overall period and a higher ADG (P < 0.05) in phase 2. On day 56, the concentrations of serum SOD (P < 0.05), and HDL (P < 0.05) and cecal SCFA contents (P < 0.05) were increased in broilers fed GLC diets. Broilers fed GLC also showed a higher microbiota diversity and an elevated abundance of SCFA-related bacteria in the caecum. The association between intestinal bacteria and metabolites was investigated via correlation analysis. The differential metabolites in the caecum, such as L-beta-aspartyl-L-aspartic acid and nicotinamide riboside, were identified. Conclusion: In summary, dietary GCL supplementation could increase growth performance to some extent. Moreover, GLC might benefit broilers' health by improving serum HDL content, antioxidant status, SCFAs contents, bacterial diversity, and probiotic proliferation in the caecum.

Comparative proteomic analysis of two divergent strains provides insights into thermotolerance mechanisms of Ganoderma lingzhi.

Heat stress (HS) is a major abiotic factor influencing fungal growth and metabolism. However, the genetic basis of thermotolerance in Ganoderma lingzhi (G. lingzhi) remains largely unknown. In this study, we investigated the thermotolerance capacities of 21 G. lingzhi strains and screened the thermo-tolerant (S566) and heat-sensitive (Z381) strains. The mycelia of S566 and Z381 were collected and subjected to a tandem mass tag (TMT)-based proteome assay. We identified 1493 differentially expressed proteins (DEPs), with 376 and 395 DEPs specific to the heat-tolerant and heat-susceptible genotypes, respectively. In the heat-tolerant genotype, upregulated proteins were linked to stimulus regulation and response. Proteins related to oxidative phosphorylation, glycosylphosphatidylinositol-anchor biosynthesis, and cell wall macromolecule metabolism were downregulated in susceptible genotypes. After HS, the mycelial growth of the heat-sensitive Z381 strain was inhibited, and mitochondrial cristae and cell wall integrity of this strain were severely impaired, suggesting that HS may inhibit mycelial growth of Z381 by damaging the cell wall and mitochondrial structure. Furthermore, thermotolerance-related regulatory pathways were explored by analyzing the protein-protein interaction network of DEPs considered to participate in the controlling the thermotolerance capacity. This study provides insights into G. lingzhi thermotolerance mechanisms and a basis for breeding a thermotolerant germplasm bank for G. lingzhi and other fungi.

Optimization of mycelial growth and cultivation of wild Ganoderma sinense.

Ganoderma sinense, a well-known medicinal macrofungus of Basidiomycetes, is widely used in traditional medicine for promoting health and longevity in East Asia. The fruiting bodies of G. sinense contain polysaccharides, ergosterol, and coumarin, which have antitumor, antioxidant, and anticytopenia activities. Mushroom cultivation requires suitable conditions for the formation of fruiting bodies and yield. However, little is known about the optimal culture conditions for mycelial growth and cultivation of G. sinense. In this study, the successful cultivation of a G. sinense strain collected from the wild was reported. The optimal culture conditions were identified by examining one factor at a time. The results of this study revealed that the nutritional requirements for the optimal mycelial growth of G. sinense were fructose (15 g/l) as the carbon source and yeast extract (1 g/l) as the nitrogen source. The optimal pH and temperature for G. sinense were 7 and 25-30°C, respectively. The mycelia grew fastest in treatment II (69% rice grains + 30% sawdust + 1% calcium carbonate). G. sinense produced fruiting bodies under all tested conditions and showed the highest biological efficiency (2.95%) in treatment B (96% sawdust, 1% wheat bran, 1% lime). In summary, under optimal culture conditions, G. sinense strain GA21 showed satisfactory yield and a high potential for commercial cultivation.

Transcriptome analysis of Ganoderma lingzhi (Agaricomycetes) response to Trichoderma hengshanicum infection.

Green mold caused by Trichoderma spp. has become one of the most serious diseases which threatening the production of Ganoderma lingzhi. To understand the possible resistance mechanism of the G. lingzhi response to T. hengshanicum infection, we examined the G. lingzhi transcript accumulation at 0, 12, and 24 h after T. hengshanicum inoculation. The gene expression analysis was conducted on the interaction between G. lingzhi and T. hengshanicum using RNA-seq and digital gene expression (DGE) profiling methods. Transcriptome sequencing indicated that there were 162 differentially expressed genes (DEGs) at three infection time points, containing 15 up-regulated DEGs and 147 down-regulated DEGs. Resistance-related genes thaumatin-like proteins (TLPs) (PR-5s), phenylalanine ammonia-lyase, and Beta-1,3-glucan binding protein were significantly up-regulated. At the three time points of infection, the heat shock proteins (HSPs) genes of G. lingzhi were down-regulated. The down-regulation of HSPs genes led to the inhibition of HSP function, which may compromise the HSP-mediated defense signaling transduction pathway, leading to G. lingzhi susceptibility. Pathway enrichment analyses showed that the main enriched pathways by G. lingzhi after infection were sphingolipid metabolism, ether lipid metabolism, and valine, leucine and isoleucine degradation pathway. Overall, the results described here improve fundamental knowledge of molecular responses to G. lingzhi defense and contribute to the design of strategies against Trichoderma spp.

Re-Examination of the Holotype of Ganoderma sichuanense (Ganodermataceae, Polyporales) and a Clarification of the Identity of Chinese Cultivated Lingzhi.

The widely cultivated Chinese Lingzhi is a famous fungus with significant medicinal and economic value, which has commonly been misidentified as Ganoderma lucidum for a long period of time. The scientific binomial of the fungus is always a hotly debated question that revolves around G. lingzhi and G. sichuanense. To interpret the species concept of the taxon, six specific primers for G. sichuanense and one universal primer were designed. Through directed and nested PCRs, we obtained nine ITS sequences from the holotype (HMAS 42798) of G. sichuanense. By genome sequencing, the ITS sequence of the first cultivated Lingzhi (HMAS 25103) was assembled. Based on a phylogenetic study of the genus Ganoderma, the correct name for widely cultivated Ganoderma species in China was confirmed as G. sichuanense, and G. lingzhi should be a later synonym.

Characteristics of the Genome, Transcriptome and Ganoderic Acid of the Medicinal Fungus Ganoderma lingzhi.

Ganoderma (Ganodermaceae) is a genus of edible and medicinal mushrooms that create a diverse set of bioactive compounds. Ganoderma lingzhi has been famous in China for more than 2000 years for its medicinal properties. However, the genome information of G. lingzhi has not been characterized. Here, we characterized its 49.15-Mb genome, encoding 13,125 predicted genes which were sequenced by the Illumina and PacBio platform. A wide spectrum of carbohydrate-active enzymes, with a total number of 519 CAZymes were identified in G. lingzhi. Then, the genes involved in sexual recognition and ganoderic acid (GA, key bioactive metabolite) biosynthesis were characterized. In addition, we identified and deduced the possible structures of 20 main GA constituents by UPLC-ESI-MS/MS, including a new special ganochlearic acid A. Furthermore, 3996 novel transcripts were discovered, and 9276 genes were predicted to have the possibility of alternative splicing from RNA-Seq data. The alternative splicing genes were enriched for functional categories involved in protein processing, endocytosis, and metabolic activities by KEGG. These genomic, transcriptomic, and GA constituents' resources would enrich the toolbox for biological, genetic, and secondary metabolic pathways studies in G. lingzhi.

Species diversity of Ganoderma (Ganodermataceae, Polyporales) with three new species and a key to Ganoderma in Yunnan Province, China.

Ganoderma is a globally distributed genus that encompasses species with forestry ecological, medicinal, economic, and cultural importance. Despite the importance of this fungus, the studies on the species diversity of Ganoderma in Yunnan Province, China (YPC) have poorly been carried out. During this study, opportunistic sampling was used to collect 21 specimens of Ganoderma from YPC. Morphology and multigene phylogeny of the internal transcribed spacer (ITS) regions, the large subunit of nuclear ribosomal RNA gene (nrLSU), the translation elongation factor 1-α gene (TEF1-α), and the second largest subunit of RNA polymerase II (RPB2) were used to identify them. Morphological and molecular characterization of the 21 specimens showed that they belong to 18 species of Ganoderma, of which three are novel viz. G. artocarpicola, G. obscuratum and G. yunnanense. Ganoderma artocarpicola is characterized by the sessile and concrescent basidiomata, reddish brown to yellowish brown pileus surface, heterogeneous context, wavy margin, and ovoid basidiospores. Ganoderma obscuratum is distinguished by small pores (6-9 per mm), dorsolaterally sub-stipitate basidiomata which become greyish-brown when dry, and narrow ellipsoid basidiospores. Ganoderma yunnanense is characterized by cream color pore surface and context, centrally to laterally stipitate basidiomata with reddish-brown to violet-brown strongly laccate pileus surface, and broadly ellipsoid basidiospores. With the help of an extensive literature survey and the results of this study, a checklist of 32 Ganoderma species from YPC was established, which accounts for 71.11% of the known species in China. In addition, a key to the Ganoderma in YPC is also provided.

Investigating the cellular antioxidant and anti-inflammatory effects of the novel peptides in lingzhi mushrooms.

The lingzhi mushroom (Ganoderma lucidum) is well known for its medicinal properties and has long played a role in traditional oriental medicine due to its health-giving benefits and potential to extend life expectancy. The mushroom contains a number of highly bioactive compounds and can also act as an excellent source of protein. This research investigated the peptides obtained from the protein hydrolysates of lingzhi mushrooms to assess their free radical scavenging abilities. These peptides were acquired via different proteases (Alcalase, Neutrase, papain, and pepsin-pancreatin) and were tested at a range of different concentrations (1.0%, 2.5%, and 5.0% w/v). The highest levels of 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and nitric oxide (NO) radical scavenging activities were presented by lingzhi mushroom hydrolysate using 2.5% (w/v) pepsin-pancreatin after 6 h of digestion. The hydrolysate was then fractionated using 10, 5, 3, and 0.65 kDa molecular weight cut-off membranes. The results showed that the MW 0.65 kDa fraction had the highest level of free radical scavenging activity. Further analysis of this MW 0.65 kDa fraction began with another RP-HPLC fractionation technique to obtain three further sub-fractions. De novo peptide sequencing using electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-Q-TOF-MS/MS) was chosen as the optimum method for studying the F3 sub-fraction. DRVSIYGWG and ALLSISSF were discovered as new peptides with different antioxidant properties. Adenocarcinoma colon (Caco-2) cells showed the antioxidant action of these synthesized peptides. This activity was linked to peptide concentration. The peptides and their pure synthetic counterparts were found to reduce NO generation by RAW 264.7 macrophages without causing cytotoxicity. The results of gene expression reveal that the DRVSIYGWG and ALLSISSF peptides were able to cut the expression of the proinflammatory cytokine genes iNOS, IL-6, TNF-α, and COX-2 in the context of RAW 264.7 macrophages.

Intraspecific comparison of mitochondrial genomes reveals the evolution in medicinal fungus Ganoderma lingzhi.

Several mitogenomes of the genus Ganoderma have been assembled, but intraspecific comparisons of mitogenomes in Ganoderma lingzhi have not been reported. In this study, 19 G. lingzhi mitogenomes were assembled and analyzed combined with three mitogenomes of G. lingzhi from GenBank in term of the characteristics, evolution, and phylogeny. The results showed that the mitogenomes of the G. lingzhi strains are closed circular ranging from 49.23 kb to 68.37 kb. The genetic distance, selective pressure, and base variation indicate that the 14 common protein coding genes were highly conserved. The differences in introns, open reading frames, and repetitive sequences in the mitogenome were the main factors leaded to the variations in mitogenome. The introns were horizontally transferred in mitogenomes, and the differences between introns in the same insertion, which were primarily caused by the repetitive sequence, showed that the introns may be under degeneration. Besides, the frequent insertion and deletion of introns showed an evolutionary rate faster than protein coding genes. Phylogenetic analysis showed that the G. lingzhi strains gathered with high support, and those with the same intron distribution law had closer clustering relationships.

GlPP2C1 Silencing Increases the Content of Ganodermalingzhi Polysaccharide (GL-PS) and Enhances Slt2 Phosphorylation.

Polysaccharides have attracted much attention in the food industry due to their diverse biological activities. To date, research on the mechanism of polysaccharide synthesis has mainly focused on the role of crucial enzymes in the polysaccharide synthesis pathway, but other genes that regulate polysaccharide synthesis have not been well explored. In this study, the GlPP2C1 gene, encoding a phosphoprotein type 2C phosphatase, was cloned, and PP2C-silenced strains (PP2C1i-1 and PP2C1i-3) were screened. Measurements of the polysaccharide content and cell wall tolerance revealed that GlPP2C1 silencing increased the polysaccharide content and enhanced cell wall resistance in Ganoderma lingzhi. The contents of intracellular polysaccharides (IPS), extracellular polysaccharides (EPS) and ß-1,3-D-glucan in PP2C-silenced strains were increased by 25%, 33% and 36%, respectively, compared with those in the WT strains and strains transformed with an empty vector. Further mechanistic studies showed that GlPP2C1 silencing increased the content of Ganoderma lingzhi polysaccharides (GL-PS) through Slt2. In summary, this study revealed the mechanism through which protein phosphatase regulates GL-PS biosynthesis for the first time.

Anticancer and Antioxidant Activities in Ganoderma lucidum Wild Mushrooms in Poland, as Well as Their Phenolic and Triterpenoid Compounds.

The goal of this study was to the assess anti-cancer and antioxidant properties of the Ganoderma lucidum fruiting body, and to identify bioactive compounds found in their extracts. Significant antiproliferative activity was observed against MCF-7, MCF-7/DX, LOVO, LOVO/DX, MDA-MB 231, SW 620, and NHDF cell lines. With IC50 values of 25.38 µg/mL and 47.90 µg/mL, respectively, the extract was most effective against MDA-MB 231 and SW 620 cell lines. The bioactive compounds were identified using an ACQUITY UPLC-PDA-MS system. The extracts contained 13 triterpenoids and 28 polyphenols from the flavonols, phenolic acids, flavones, flavan-3-ols, and stilbenes families. Ganoderic acid derivative was found to be the most abundant triterpenoid (162.4 mg/g DW), followed by ganoderic acid B (145.6 mg/g DW). Resveratrol was the most abundant phenolic in the extract (5155.7 mg/100 g DM). The findings could explain why G. lucidum extracts are used in folk medicine.

Discovery of a Multifunctional Octapeptide from Lingzhi with Antioxidant and Tyrosinase Inhibitory Activity.

Ganoderma lucidum or Lingzhi is a fungus species widely known as a traditional medicine. Exploring the beneficial peptides by hydrolysis using pepsin and trypsin has been extensively performed to identify new bioactive natural products. A multifunctional peptide that expresses potential scavenging activity and tyrosinase inhibition is valuable in therapeutic and cosmetic applications. This study aimed to identify and investigate the effects of a novel multifunctional peptide from Lingzhi on the melanogenic enzymes in melanoma cells by a targeted-proteomics approach. The multifunctional peptide was de novo sequenced by LC-MS/MS to be NH2-PVRSSNCA-CO2H (octapeptide). This sequence was chemically synthesized by solid-phase peptide synthesis (SPPS). The antioxidant ability of the synthesized octapeptide was measured by the DPPH, ABTS, and FRAP assays. The results showed that the peptide exhibited an antioxidant activity equal to 0.121 ± 0.01 mg equivalent to ascorbic acid, 0.173 ± 0.03 mg equivalent to gallic acid, and 2.21 ± 0.23 mM equivalent to FeSO4, respectively, which is comparable to these well-known antioxidants. The proteomics approach identified a total of 5804 proteins and several pathways involved in the effects of the octapeptide in melanoma cells. Targeted proteomics revealed three specific proteins associated with pigmentation including Rab29, Dct, and Tyrp1. The Rab29 and Dct were upregulated whereas Tyrp1 was downregulated in the octapeptide treatment group. These findings could be used in the understanding of the molecular functions of the multifunctional octapeptide on melanogenic enzymes, supporting its potential as a therapeutic and cosmetic ingredient.