Next Article in Journal
Dual Anta-Inhibitors Targeting Protein Kinase CK1δ and A2A Adenosine Receptor Useful in Neurodegenerative Disorders
Previous Article in Journal
Background-Quenched Aggregation-Induced Emission through Electrostatic Interactions for the Detection of Poly(ADP-ribose) Polymerase-1 Activity
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Molecules
Volume 28
Issue 12
10.3390/molecules28124760
molecules-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Communication

DPPH Radical Scavenging Activity of New Phenolics from the Fermentation Broth of Mushroom Morehella importuna

by
Feifei Wang
1,2,
Jie Tan
1,2,
Ruixiang Jiang
1,2,
Feifei Li
1,2,
Renqing Zheng
1,2,
Linjun Yu
1,2,
Lianzhong Luo
3,* and
Yongbiao Zheng
1,2,*
1
Engineering Research Centre of Industrial Microbiology, Ministry of Education, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China
2
Provincial University Key Laboratory of Cellular Stress Response and Metabolic Regulation, College of Life Sciences, Fujian Normal University, Fuzhou 350117, China
3
Engineering Research Center of Marine Biopharmaceutical Resource, **amen Medical College, **amen 361023, China
*
Authors to whom correspondence should be addressed.
Molecules 2023, 28(12), 4760; https://doi.org/10.3390/molecules28124760
Submission received: 5 May 2023 / Revised: 11 June 2023 / Accepted: 11 June 2023 / Published: 14 June 2023
Browse Figures
Review Reports Versions Notes

Abstract

:
In recent years, wild morel mushroom species have begun to be widely cultivated in China due to their high edible and medicinal values. To parse the medicinal ingredients, we employed the technique of liquid-submerged fermentation to investigate the secondary metabolites of Morehella importuna. Two new natural isobenzofuranone derivatives (1–2) and one new orsellinaldehyde derivative (3), together with seven known compounds, including one o-orsellinaldehyde (4), phenylacetic acid (5), benzoic acid (6), 4-hydroxy-phenylacetic acid (7), 3,5-dihydroxybenzoic acid (8), N,N′-pentane-1,5-diyldiacetamide (9), and 1H-pyrrole-2-carboxylic acid (10), were obtained from the fermented broth of M. importuna. Their structures were determined according to the data of NMR, HR Q-TOF MS, IR, UV, optical activity, and single-crystal X-ray crystallography. TLC-bioautography displayed that these compounds possess significant antioxidant activity with the half DPPH free radical scavenging concentration of 1.79 (1), 4.10 (2), 4.28 (4), 2.45 (5), 4.40 (7), 1.73 (8), and 6.00 (10) mM. The experimental results would shed light on the medicinal value of M. importuna for its abundant antioxidants.

Graphical Abstract

1. Introduction

The genera Morchella (Morel), which includes these species M. importuna, M. esculenta, M. sextalata, and M. eximia, are edible medicinal fungi of the phylum Ascomycota with high gastronomic quality and potential therapeutic use [1]. The fruiting body of these species contained abundant bioactivities substances [2]. It was reported that the aqueous extract from M. importuna possessed antileishmanial activity, a novel peptide from M. importuna could induce apoptosis in HeLa cells, phenolic compounds from M. esculenta with high antioxidant activity [3], and sterols and trilinolein showed significant inhibition of NF-κB activation [4]. However, the wild morel is rare and could not be artificially cultivated until recent years. To utilize the natural resources of this genus mushroom, mycelia prepared by microbial fermentation technology were an alternative product for the investigation of its chemical constitution with pharmaceutical potential. Some reports provided the convincing truth that mycelia and other products prepared from submerged culture are a valid alternative to the fruiting body, such as M. esculenta [5]. Microbial fermentation was generally used to explore the metabolites of some wild and rare mushrooms in our lab. In this paper, we mainly focus on the isolation, chemical structures, and antioxidant properties of the low-molecule metabolites of M. importuna by microbial fermentation. As a result, two new isobenzofuranone derivatives and one new orsellinaldehyde derivative, together with five phenolics and two N-contained metabolites hexamethylene bisacetamide and 1H-pyrrole-2-carboxylic acid, were isolated from the fermentation broth. These natural metabolites show significant antioxidant activity.

2. Results

2.1. Structure Identification

Ten compounds were purified from the organic extract prepared from the fermentation broth of M. importuna as reported in detail in the experimental part.
Inspection of the spectral data of compounds 1–2 (Table 1) indicated that they are derivatives of isobenzofuranone, which possess one phenyl group substituted by two meta phenolic hydroxyls according to two meta-coupled aromatic protons (Table 1). Compound 1 was isolated as a white amorphous substance with [ a ] D 20 − 23.0 (c 0.36, MeOH) and UV (methanol) λmax, nm: 209. The molecular formula was determined to be C9H8O5 by High-resolution Quadrupole Time-of-Flight Mass Spectrometry (HR Q-TOF MS) at 197.0441 for [M + H]+ (calculated for C9H9O5, 197.0450). The IR spectra showed the absorptions for hydroxyl (3147 cm−1), ester carbonyl, and aromatic unsaturated double bond (1623 cm−1). The 13C NMR data revealed six carbon signals (δ 103.1, 109.6, 123.4, 131.3, 155.5, and 162.5) for one phenyl group, which was substituted by two meta phenolic hydroxyls according to the chemical shift and coupling constant of two protons [δ 6.74 (d, J = 2.0 Hz) and 6.68 (d, J = 2.0 Hz)]. The key heteronuclear multiple-bond correlation (HMBC) spectrum of 1 exhibited correlations from H-7 to C-1, C-5, C-6, and C-3a, H-5 to C-4, C-6, C-7, and C-3a, and H-3 to C-1 and C-8 and allowed the establishment of isobenzofuranone substituted with two hydroxyls and one methoxyl groups in 1. The planar structure of 1 was characterized as one new derivative of isobenzofuranone, namely 4,6-dihydroxy-3-methoxyisobenzofuran-1(3H)-one. The absolute configuration of C-3 was assigned by the optical rotation test. Rubralide A [6] isolated from Penicillium rubrum showed the dextral optical rotation with the value of [ a ] D 20 + 4.0 (c 0.5, EtOH), whose only one chiral carbon has an (R)-configuration at C-3. The sinistral rotation value of [ a ] D 20 − 23.0 (c 0.36, MeOH) of 1, indicated the (S)-configuration at C-3. Therefore, compound 1 was concluded as (S)-4,6-dihydroxy-3-methoxyisobenzofuran-1(3H)-one. Compound 2 was isolated as white amorphous substance with the UV (methanol) λmax, nm: 224. The molecular formula of 2 was determined to be C9H8O5 by HR Q-TOF MS at 197.0443 for [M + H]+ (calculated for C9H9O5, 197.0450). The IR spectra showed the absorptions for hydroxyl (3312 cm−1), ester carbonyl (1684 cm−1), and aromatic unsaturated double bond (1637 cm−1). The NMR spectrum of 2 was similar to those of 1, except for the presence of the proton at δH 10.33 (H-3) and carbon signal at δC 196.6 (C-3) (Table 1). The downfield chemical shift of H-3 and C-3 indicated the presence of a formaldehyde group in compound 2, compared with the hemiacetal group at same position in compound 1. HMBC correlations from H-7 to C-5, C-6, and C-3a, H-5 to C-4, C-6, and C-3a, H-3 to C-1 and C-5, and H-8 to C-1 were observed in HMBC spectrum of compound 2, indicating the opening of the furanone ring of an isobenzofuranone skeleton. Compound 2 was deduced as methyl 2-formyl-3,5-dihydroxybenzoate according to the IUPAC system, a hydrolysis product of 1 (Figure 1). Compound 2 was often used as the building block for the synthetic medicinal chemicals [7] and reported as a natural product for the first time in this paper. Isobenzofuranone is also known as phthalide and is widely distributed in microbes and plants [8]. In recent years, many natural molecules with this skeleton were discovered and shown diverse biological activities, such as antibacterial [9], antifungal [10], insecticidal [11], cytotoxic [12], anticyclooxygenase-2 [13], anti-acetylcholinesterase [14], α-glucosidase inhibitory effect [15], anticoagulation [16], hepatoprotective [17], and neurodegenerative prevention [18], etc. So, phthalide analogs exhibit a potential pharmacological value. Except for Hericium erinaceus [19,20] and Pleurotus djamor [21], few reports refer to phthalide-related metabolites isolated from mushrooms. New isobenzofuranone derivative 1 from mushroom M. importuna merits further investigation for its bioactivities.
Compound 4 was isolated as a white powder. The 13C NMR data (recorded in (CD3)2OD, 500 M) revealed eight carbon signals for one methyl (δC 18.2), six sp2 carbons (δC 113.2, 167.1, 101.4, 166.5, 111.6, 146.0), and one carbonyl carbon (δC 194.2) in compound 4. Six protons’ peaks for δH 10.09 (s, 1H), δH 6.30 (dd, J = 2.3, 0.9 Hz, 1H), 6.17 (d, J = 2.3 Hz, 1H) and δH 2.53 (s, 3H) were observed in 1H NMR spectra of 4. Based on the above 1H and 13C NMR data, 4 was identified as 2,4-dihydroxy-6-methylbenzaldehyde, namely o-orsellinaldehyde, which was confirmed by X-ray diffraction (Figure 1). Crystallographic data (CCDC 2214836) for 4: C8H8O3, monoclinic, space group P21/c, a = 7.4221(4) Å, b = 13.1943(8) Å, c = 7.1941(4) Å, α = 90°, β = 90.425 (5)°, γ = 90°, V = 704.50 (7) Å3, Z = 4, Dc = 1.434 g·cm−3, F(000) = 320, 16,396 reflections measured, 1218 unique (Rint = 0.0862) which were used in all calculations. The final ωR2 was 0.1361 (all data) and R1 was 0.0495 (I ≥ 2σ (I)).
Compound 3 was isolated as a white powder with UV (methanol) λmax, nm: 202. The molecular formula of compound 3 was determined to be C9H10O4 based on the HR Q-TOF MS peak at m/z: 261.1823 (calculated for C9H10O4Na, 261.1830) and NMR data. The IR spectra showed the absorptions for hydroxyl (3433 cm−1) and an aromatic unsaturated double bond (1630 cm−1). Additionally, 1H NMR [(CD3)2OD, 500 M)]: 6.25 (s, 1H, H-5), 10.09 (s, 1H, H-7), 2.51 (s, 3H, H-8), 3.70 (s, 3H, H-9). Moreover, 13C NMR [(CD3)2OD, 125 M)]:113.1 (C-1), 135.9 (C-2), 140.5 (C-3), 160.1 (C-4), 100.9 (C-5), 162.3 (C-6), 194.6 (C-7), 10.5 (C-8), 61.1 (C-9). HMBC correlations from H3-8 to C-1/2/3, from H-7 to C-6, from H-5 to C-1/3/4/6, and from H3-9 to C-3 could be observed in HMBC spectra. Compound 3 was a new derivative of 4 and was identified as 4,6-dihydroxy-3-methoxy-2-methylbenzaldehyde (Figure 1). o-Orsellinaldehyde was reported as a bioactive metabolite produced by the mushroom Grifola frondosa with a selective cytotoxic effect and anti-inflammatory and pro-apoptotic properties [22,23], and Phlebiopsis gigantea with antifungal activity [24].
Analysis of 1H and 13C NMR spectra data (Figures S9–S16), compounds 5–8 were identified as four aromatic carboxylic acids containing the carboxylic group and the benzene ring, named as 2-phenylacetic acid, benzoic acid, 2-(4-hydroxyphenyl) acetic acid, 3,5-dihydroxybenzoic acid (Figure 1). The experimental results indicated M. importuna could yield a wide variety of phenolics. Phenolics are the vital chemical ingredient of some culinary and medicinal mushroom, such as Phellinus pini [25], Flammulina velutipes [26], Inonotus obliquus [27], Ganoderma lucidum [28], Pleurotus citrinopileatus [29], Hypsizygus marmoreus [30], Porodaedalea chrysoloma [31], Antrodia cinnamomea [32], and Tuber indicum [33], etc. Phenolics exhibit antioxidant activity and provide health benefits.
Compound 9 was isolated as a white powder with ESI-MS m/z: 187 [M + H]+. Additionally, 1H NMR (600 MHz, CDCl3) δ: 5.92 (s, 2H, 2 × NH), 3.23 (m, 4H, H-1, 5), 1.52 (m, 4H, H-2, 4), 1.36 (m, 2H, H-3), 1.98 (s, 6H, 2 × CH3). Furthermore, 13C NMR (151 MHz, CDCl3) δ: 170.6 (2 × CO), 39.3 (C-1, 5), 29.0 (C-2, 4), 23.8 (C-3), 23.4 (2 × CH3). Compared with the literature NMR data, 9 was identified as N,N’-pentane-1, 5-diyldiacetamide which was isolated as a natural product from Blaps japonensis [34]. The hybrid polar-planar compound 9 is a hexamethylene bisacetamide (HMBA) analog. It was reported HMBA was a potent inducer of erythroleukemic differentiation [35] and an inhibitor of vascular smooth muscle cell proliferation [36]. HMBA and its analogs induce hexamethylene bis-acetamide inducible protein I (HEXIM1) expression in cancer cells and achieve its biological activity. Although the molecule failed at Phase II clinical trial because of the dose-dependent toxicity, HMBA analogs showed the pharmaceutical potential.
Compound 10 was isolated as a white powder. Additionally, 1H NMR (600 MHz, CDCl3) δ: 7.34 (d, J = 3.5 Hz, 1H, H-3), 6.57 (dd, J = 3.5, 1.7 Hz, 1H, H-4), 7.65 (d, J = 1.7 Hz, 1H, H-5). Moreover, 13C NMR (151 MHz, CDCl3) δ: 143.9 (C-2), 120.3 (C-3), 112.4 (C-4), 147.6 (C-5), 163.3 (COOH). Compound 10 was identified as 1H-pyrrole-2-carboxylic acid (PCA) and further confirmed by X-ray diffraction (Figure 1). Crystallographic data (CCDC 2214809) for 10: C5H5O2, brown crystal, monoclinic, space group C2/c, a = 13.2599(14) 10−10 m, b = 5.0253(5) 10−10 m, c = 14.8921(16) 10−10 m, α = 90°, β = 99.199(11)°, γ = 90°, V = 979.57(18) 10−30 m3, Z = 4, Dc = 1.507 g∙cm−3, μ (Mo-Kα) = 0.119 mm−1, F(000) = 464.3, 2112 reflections measured, 1025 unique (Rint = 0.0128) which were used in all calculations. The final ωR2 was 0.1426 (all data) and R1 was 0.0366 (I ≥ 2σ (I)). PCA was previously reported as an antimicrobial natural product obtained from bacteria, the sponge Agelas nakamurai, and the Chinese herb Pseudostellaria heterophylla [37,38]. It was discovered for the first time as a mushroom metabolite in this paper.

2.2. DPPH Free Radical Scavenging Activity

In the TLC-bioautography experiments, the various degrees of white spot observed in the different lanes indicated that these compounds 1, 2, 4, 5, 7, 8, and 10 possessed significant antioxidant activity (Figure 2). Their antioxidant activities were further assessed by the Brand-Williams’ method. DPPH radical scavenging rate of these compounds with the different doses were shown in Figure 3. The half DPPH scavenging concentration (SC50) of these compounds were 1.79 (1), 4.10 (2), 4.28 (4), 2.45 (5), 4.40 (7), 1.73 (8), and 6.00 (10) mM, compared with the value of 0.216 mM of vitamin C, which were estimated by using Probit analysis in SPSS 18.0 with p value less than 0.01 except for 0.064 for 2. Most of these compounds displayed significant anti-DPPH radical potency. These data suggest that M. importuna could be used as a source of abundant natural antioxidants.
The DPPH radical (DPPH•) bearing a stable unpaired electron is regarded as one convenient method for determining the antioxidant activity of a wide variety of organic molecules [39]. The intracellular free radicals were believed to be involved in a diverse range of diseases and accelerating the aging process [40]. Antioxidants with the ability to scavenge free radicals and reduce oxidative stress are reported to be consecrated with all types of pharmacological applications [41]. In recent years, the antioxidant substance obtained from mushrooms were discovered to confer a beneficial effect on human health, such as antidiabetic and antihyperglycemic [42], neuroprotective [43], hepatoprotective [44], anti-inflammatory [45], immunomodulatory [46], anti-aging [47], anticancer [48], antimicrobial [49], anti-melanogenic [50] and hypopigmentation [51], anticoagulant [52], nephroprotective [53], and hyperuricemia treatment [54], etc. The abundant natural antioxidants of M. importuna will lay the foundation for its medicinal benefits.

3. Materials and Methods

3.1. General Experimental Procedures

NMR spectra were acquired on a Bruker AVANCE Ⅲ 500 spectrometer operating at 500/125 MHz and a Bruker AVANCE NEO 600 spectrometer operating at 600/150 MHz. UV spectra were recorded on a Shimadzu UV-2401PC spectrophotometer in nm (λmax). IR spectra were measured with a Bruker Vertex 70 FT-IR spectrophotometer with KBr cells in cm–1. Optical rotations were measured on a Rudolph Research Analytical polarimeter (Autopol VI). HR QTOF MS spectra were recorded on an Agillent 6520 mass spectrometer in the positive mode (4.0 KV) over the mass range m/z 200 to 800. X-ray single diffraction was performed on an Oxford Gemini S Ultra diffractor. SpectraMax® i3x multi-mode microplate reader (Molecular Devices) was used to measure the absorbance. Column chromatography was accomplished over silica gel (Qingdao Marine Chemical Company, Qingdao, China), reverse phase octadecyl-silica RP-C18 (Merck, Darmstadt, Germany), and Sephadex LH-20 (Amersham Biosciences, Piscataway, NJ, USA). Medium-pressure liquid chromatography (MPLC) was performed using a Quiksep-50IID (H&E, Bei**g, China). Thin layer chromatography (TLC) was performed on the precoated silica gel GF254 plates (Qingdao Marine Chemical Company, Qingdao, China). Organic solvents used were from Sino-pharm Chemical Reagent Co., Ltd. (Shanghai, China). Additionally, 2,2-diphenyl-1-picrylhydrazyl (DPPH) was the product of Aladdin Industrial Corporation (D1227007).

3.2. Fungus Material

The strain M. importuna was collected by **e Bao-gui (Fungal Research Centre, Fujian Agriculture and Forestry University, Fuzhou, China) and was further identified by the DNA sequence of internal transcribed spacers (ITS) region (Figure S21). The strain has been deposited in College of Life Sciences, Fujian Normal University and has been deposited in the China Centre for Type Culture Collection (CCTCC M 2014324).

3.3. Fermentation and Preparation of Extracts

M. importuna was cultured by submerged liquid fermentation [55], which was carried out in Erlenmeyer flasks (250 mL) containing 100 mL of potato dextrose medium with a total volume of 40 L. These flasks were incubated at 28 °C with a shaking speed of 180 rpm. The culture broth was centrifuged at 5000 rpm for 30 min to remove the mycelia and then extracted in batches with equal volumes of ethyl acetate. The organic phase was concentrated under reduced pressure by a rotary evaporator to afford the crude extract E1 (3.4 g). Following the same step, another potato dextrose medium which added 1 g bran and 20 g bean sprout juice per liter was used to culture M. importuna with a total volume of 36 L and obtained the crude extract E2 (8.8 g).

3.4. Isolation and Purification of Compounds 1–10 [55]

E1 (3.4 g) was firstly separated into three subfractions E1.1 (287.1 mg), E1.2 (107.4 mg), and E1.3 (15.8 mg) by MPLC over RP-C18 silica gel (170 g) using a stepwise gradient CH3OH in H2O (v/v 0:100, 5:100, 30:100, 50:50, 70:100, 100:0) based on TLC analyses. Fraction E1.1 was further chromatographed over Sephadex LH-20 (120 g) eluting with methanol and afforded the subfraction E1.11 (20 mg). E1.11 was subjected to silica gel (1.0 g) chromatography using a CHCl3-CH3OH solvent gradient to yield compound 7 (5 mg). E1.2 was separated by column chromatography over Sephadex LH-20 (120 g) with methanol eluant and afforded the subfractions E1.21 (22 mg). E1.21 was next separated by column chromatography over Sephadex LH-20 (120 g) with acetone eluant and afforded E1.211 (11.4 mg) and E1.212 (7.9 mg). Compounds 5 (5.5 mg) and 6 (5.4 mg) were, respectively, purified from E1.211 and E1.212 by column chromatography over silica gel (1 g) with the eluant of CHCl3-CH3OH solvent gradient. E1.3 was separated by column chromatography over Sephadex LH-20 (120 g) with methanol eluant and afforded E1.31 (5.4 mg) and E1.32 (7.9 mg). Then, E1.31 and E1.32 were, respectively, subjected to column chromatography over Sephadex LH-20 (120 g) with acetone eluant and yielded pure compounds 4 (2 mg) and 3 (0.7 mg).
Three fractions E2.1 (3.65 g), E2.2 (313.4 mg), and E2.3 (1.6 g) were obtained from the crude extract E2 by MPLC over RP-C18 silica gel (170 g) using 5% and 30% CH3OH in H2O as the eluant. E2.1 was then subjected to column chromatography over Sephadex LH-20 (120 g) with methanol eluant and afforded subfractions E2.11 (175.4 mg). Compound 10 (4.1 mg) was yielded from E2.11 by column chromatography over silica gel (1 g) eluting with petroleum ether-acetone solvent. E2.2 was then subjected to column chromatography over Sephadex LH-20 (120 g) with methanol eluant and afforded two subfractions E2.21 (236.6 mg) and E2.22 (68.3 mg). Compound 1 (4.2 mg) was yielded from E2.21 by column chromatography over silica gel (1 g) eluting with petroleum ether-acetone solvent. Compound 8 (6.4 mg) was acquired from E2.22 by column chromatography over silica gel (1 g) with the eluant of CHCl3-CH3OH solvent gradient. Two subfractions E2.31 (37.9 mg) and E2.32 (1.2 g) were afforded from E2.3 by column chromatography over Sephadex LH-20 (120 g) eluting with methanol. Compound 9 (7.4 mg) was purified from E2.31 (37.9 mg) by successive column chromatography over Sephadex LH-20 (120 g) using acetone eluant, and silica gel (1 g) eluting with the chloroform and methanol solution (80:1). The subfraction E2.321 (36.8 mg) was yielded from E2.32 by MPLC over RP-C18 silica gel using 20% CH3OH in H2O as the eluant. Compound 2 (2.4 mg) was obtained from E2.321 by successive column chromatography over Sephadex LH-20 (120 g) using methanol eluant and silica gel (1 g) eluting with petroleum ether-acetone solvent.

3.5. DPPH Free Radical Scavenging Activity

TLC-bioautography [56] was used to determine the free radical scavenging activity of compounds 1–10. These compounds were dispensed to the concentration of 2 mg/mL. Then, 2 μL solution of each compound was spotted onto a precoated silica gel GF254 plate. The TLC plate was left to let the solvent evaporate completely. Then, the plate was sprayed with 0.04% DPPH in ethanol and incubated at 40 °C for 30 min. Any antioxidant compounds could be seen as white spots against the blue background. Ascorbic acid was set as the positive control sample.
The DPPH scavenging rate of these compounds was further assayed by the method reported by Blois [57] and modified by Brand-Williams [58]. Each compound was dissolved in methanol and diluted to 6 different concentrations with the same volume of 94 μL in a 96-well microtiter plate. Then, 20 μL of DPPH dissolved in methanol with a concentration of 0.714 μg/mL was added to these wells. The final concentration of these compounds in each well was 9.8, 19.6, 39.3, 78.6, 157.2, and 314.4 μg/mL, respectively. Ascorbic acid was set as the positive control group with the final concentration of 0.5, 1.0, 2.0, 4.0, 8.0, and 16.0 μg/mL. The reaction mixtures were incubated for 30 min in a dark room at room temperature. The absorbance was read by a microplate reader at 519 nm. The percentage of the DPPH scavenging effect was calculated with the following equation:
DPPH scavenging effect (%) = [(A0 − A1)/A0 × 100%]
where A1 is the absorbance of the samples and the standards, and A0 is the absorbance of the vehicle.

3.6. X-ray Single Crystal Diffraction for Compound 4 and 10

The single crystal of 4 and 10 was obtained from aqueous acetone. A suitable crystal was selected and subjected to an Oxford Gemini S Ultra diffractometer using Cu-Kα (λ = 1.54184 Å) radiation at 99 K for 4 and using Mo-Kα (λ = 0.7073 Å) radiation at 273 K for 10. Their structures were determined using the direct method and refined with full-matrix least-squares calculations on F2 using olex2-2.1 [59]. Crystallographic data have been deposited with the Cambridge Crystallographic Data Centre (CCDC).

4. Conclusions

In summary, M. importuna can produce abundant phenolics, including two new isobenzofuranone derivatives and one new orsellinaldehyde derivative. These natural metabolites show the significant antioxidant activity of TLC-bioautography and DPPH free radical scavenging assay. The experimental results provided a scientific foundation for the development of the medicinal value of M. importuna.

Supplementary Materials

The following supporting information can be downloaded at: https://mdpi.longhoe.net/article/10.3390/molecules28124760/s1, Figures S1–S20: 1H and 13C NMR spectra for all compounds; Figure S21: ITS DNA sequence of the strain Morchella importuna.

Author Contributions

Funding acquisition, Y.Z.; Investigation, Y.Z., F.W., J.T., R.J., F.L., R.Z. and L.Y.; Methodology, Y.Z., L.Y. and L.L.; Project administration, Y.Z., and L.L.; Writing—original draft, Y.Z.; Writing—review and editing, Y.Z. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by Natural Science Foundation of Fujian Province (2020J01172), Engineering Research Center of Marine Biopharmaceutical Resource, Fujian Province University, **amen Medical College (MBS-201902 and MNPR-2022002), and the scientific research innovation program “**yuanjiang River Scholarship” of College of Life Sciences, Fujian Normal University (XRS-202217).

Data Availability Statement

The data has supplied in Supporting Informations.

Acknowledgments

The authors thank the Testing Center of Fuzhou University for the NMR and mass data and the State Key Laboratory of Physical Chemistry of Solid Surface at **amen University for the single crystal X-ray diffraction data.

Conflicts of Interest

The authors declare no conflict of interest.

Sample Availability

Samples of compounds 1–10 are available from the authors.

References

  1. Sunil, C.; Xu, B. Mycochemical profile and health-promoting effects of morel mushroom Morchella esculenta (L.) A review. Food Res. Int. 2022, 159, 111571. [Google Scholar] [CrossRef] [PubMed]
  2. Sambyal, K.; Singh, R.V. A comprehensive review on Morchella importuna: Cultivation aspects, phytochemistry, and other significant applications. Folia Microbiol. 2021, 66, 147–157. [Google Scholar] [CrossRef] [PubMed]
  3. Yildiz, O.; Can, Z.; Laghari, A.Q.; Sahin, H.; Malkoc, M. Wild Edible Mushrooms as a Natural Source of Phenolics and Antioxidants. J. Food Biochem. 2015, 39, 148–154. [Google Scholar] [CrossRef]
  4. Kim, J.A.; Lau, E.; Tay, D.; De Blanco, E.J.C. Antioxidant and NF-κB inhibitory constituents isolated from Morchella esculenta. Nat. Prod. Res. 2011, 25, 1412–1417. [Google Scholar] [CrossRef] [PubMed]
  5. Ma, T.W.; **ao, B.Y.; Yang, F.C. Establishment of cultivating strategy for highly aggregated mycelia of Morchella esculenta in a stirred-tank bioreactor. Bioprocess Biosyst. Eng. 2012, 35, 1627–1635. [Google Scholar] [CrossRef]
  6. Kimura, Y.; Yoshinari, T.; Koshino, H.; Fujioka, S.; Okda, K.; Shimada, A. Rubralactone, Rubralides A, B and C, and Rubramin Produced by Penicillium rubrum. Biosci. Biotechnol. Biochem. 2007, 71, 1896–1901. [Google Scholar] [CrossRef]
  7. Katoh, T.; Ohmori, O.; Iwasaki, K.; Inoue, M. Synthetic studies on Sch 202596, an antagonist of the galanin receptor subtype GalR1: An efficient synthesis of (±)-geodin, the spirocoumaranone part of Sch 202596. Tetrahedron 2002, 58, 1289–1299. [Google Scholar] [CrossRef]
  8. León, A.; Del-Ángel, M.; Ávila, J.L.; Delgado, G. Phthalides: Distribution in Nature, Chemical Reactivity, Synthesis, and Biological Activity. Prog. Chem. Org. Nat. Prod. 2017, 104, 127–246. [Google Scholar]
  9. Chen, Q.; Yu, J.J.; He, J.; Feng, T.; Liu, J.K. Isobenzofuranones and isocoumarins from kiwi endophytic fungus Paraphaeosphaeria sporulosa and their antibacterial activity against Pseudomonas syringae pv. actinidiae. Phytochemistry 2022, 195, 113050. [Google Scholar] [CrossRef]
  10. Sánchez-Fernández, R.E.; Sánchez-Fuentes, R.; Rangel-Sánchez, H.; Hernández-Ortega, S.; López-Cortés, J.G.; Macías-Rubalcava, M.L. Antifungal and antioomycete activities and modes of action of isobenzofuranones isolated from the endophytic fungus Hypoxylon anthochroum strain Gseg1. Pestic. Biochem. Physiol. 2020, 169, 104670. [Google Scholar] [CrossRef]
  11. Farias, E.S.; Silva, E.M.P.; Teixeira, M.G.; Ferreira, J.S.; Alvarenga, E.S.; Picanço, M.C. Phthalides as promising insecticides against Tuta absoluta (Lepidoptera: Gelechiidae). J. Environ. Sci. Health Part B 2018, 53, 49–56. [Google Scholar] [CrossRef]
  12. Zhang, Y.; Yang, Y.; Ruan, J.; Chen, Q.; Li, J.; Guo, Y.; Han, L.; Wang, T. Isobenzofuranones from the aerial parts of Leontopodium leontopodioides (Wild.) Beauv. Fitoterapia 2018, 124, 66–72. [Google Scholar] [CrossRef]
  13. Lv, J.L.; Zhang, L.B.; Guo, L.M. Phthalide dimers from Angelica sinensis and their COX-2 inhibition activity. Fitoterapia 2018, 129, 102–107. [Google Scholar] [CrossRef]
  14. Dai, Y.; Li, K.; She, J.; Zeng, Y.; Wang, H.; Liao, S.; Lin, X.; Yang, B.; Wang, J.; Tao, H.; et al. Lipopeptide Epimers and a Phthalide Glycerol Ether with AChE Inhibitory Activities from the Marine-Derived Fungus Cochliobolus Lunatus SCSIO41401. Mar. Drugs 2020, 18, 110547. [Google Scholar] [CrossRef]
  15. Yan, Z.; Huang, C.; Guo, H.; Zheng, S.; He, J.; Lin, J.; Long, Y. Isobenzofuranone monomer and dimer derivatives from the mangrove endophytic fungus Epicoccum nigrum SCNU-F0002 possess α-glucosidase inhibitory and antioxidant activity. Bioorg. Chem. 2020, 94, 103407. [Google Scholar] [CrossRef]
  16. Zhang, L.B.; Lv, J.L.; Liu, J.W. Phthalide Derivatives with Anticoagulation Activities from Angelica sinensis. J. Nat. Prod. 2016, 79, 1857–1861. [Google Scholar] [CrossRef]
  17. Zhang, X.; Yan, H.W.; Feng, Z.M.; Yang, Y.N.; Jiang, J.S.; Zhang, P.C. Neophathalides A and B, two pairs of unusual phthalide analog enantiomers from Ligusticum chuanxiong. Org. Biomol. Chem. 2020, 18, 5453–5457. [Google Scholar] [CrossRef]
  18. Dos Reis Teixeira, A.; Teixeira, R.R.; Ribeiro, I.M.L.; Pereira, W.L.; Manhabosco, T.M.; de Brito, A.C.F.; Oliveira, L.A.M.; Coelho Nogueira, K.O.P. Association of electroanalytical and spectrophotometric methods to evaluate the antioxidant activity of isobenzofuranone in primary cultures of hippocampal neurons. Toxicol. In Vitro 2020, 68, 104970. [Google Scholar] [CrossRef]
  19. Li, J.; Wang, X.L.; Li, G.; Xu, P.S.; Xu, K.P.; Tan, G.S. Two new isobenzofuranone derivatives from the fruiting bodies of Hericium erinaceus. J. Asian Nat. Prod. Res. 2017, 19, 1108–1113. [Google Scholar] [CrossRef]
  20. Wang, X.L.; Gao, J.; Li, J.; Long, H.P.; Xu, P.S.; Xu, K.P.; Tan, G.S. Three new isobenzofuranone derivatives from the fruiting bodies of Hericium erinaceus. J. Asian Nat. Prod. Res. 2017, 19, 134–139. [Google Scholar] [CrossRef]
  21. Colmenares-Cruz, S.; González-Cortazar, M.; Castañeda-Ramírez, G.S.; Andrade-Gallegos, R.H.; Sánchez, J.E.; Aguilar-Marcelino, L. Nematocidal activity of hydroalcoholic extracts of spent substrate of Pleurotus djamor on L(3) larvae of Haemonchus contortus. Vet. Parasitol. 2021, 300, 109608. [Google Scholar] [CrossRef] [PubMed]
  22. Tomas-Hernandez, S.; Garcia-Vallvé, S.; Pujadas, G.; Valls, C.; Ojeda-Montes, M.J.; Gimeno, A.; Cereto-Massagué, A.; Roca-Martinez, J.; Suárez, M.; Arola, L.; et al. Anti-inflammatory and Proapoptotic Properties of the Natural Compound o-Orsellinaldehyde. J. Agric. Food Chem. 2018, 66, 10952–10963. [Google Scholar] [CrossRef] [PubMed]
  23. Lin, J.T.; Liu, W.H. ο-Orsellinaldehyde from the Submerged Culture of the Edible Mushroom Grifola frondosa Exhibits Selective Cytotoxic Effect Against Hep 3B Cells Through Apoptosis. J. Agric. Food Chem. 2006, 54, 7564–7569. [Google Scholar] [CrossRef] [PubMed]
  24. Kälvö, D.; Menkis, A.; Broberg, A. Secondary Metabolites from the Root Rot Biocontrol Fungus Phlebiopsis gigantea. Molecules 2018, 23, 1417. [Google Scholar] [CrossRef] [Green Version]
  25. Ryang, J.; Liu, F.; Ng, T.B. Purified antioxidant from the medicinal mushroom Phellinus pini protects rat H9c2 cell against H(2) O(2) -induced oxidative stress. J. Food Biochem. 2021, 45, e13818. [Google Scholar] [CrossRef]
  26. Ma, S.; Zhang, H.; Xu, J. Characterization, Antioxidant and Anti-Inflammation Capacities of Fermented Flammulina velutipes Polyphenols. Molecules 2021, 26, 6205. [Google Scholar] [CrossRef]
  27. Zhao, W.; Huang, P.; Zhu, Z.; Chen, C.; Xu, X. Production of phenolic compounds and antioxidant activity via bioconversion of wheat straw by Inonotus obliquus under submerged fermentation with the aid of a surfactant. J. Sci. Food Agric. 2021, 101, 1021–1029. [Google Scholar] [CrossRef]
  28. Kebaili, F.F.; Tahar, N.; Esseddik, T.M.; Redouane, R.; Chawki, B.; Pablo, A.; Massimiliano, P. Antioxidant Activity and Phenolic Content of Extracts of Wild Algerian Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum (Agaricomycetes). Int. J. Med. Mushrooms 2021, 23, 79–88. [Google Scholar] [CrossRef]
  29. Yin, C.; Fan, X.; Liu, C.; Fan, Z.; Shi, D.; Yao, F.; Cheng, W.; Gao, H. The Antioxidant Properties, Tyrosinase and α-Glucosidase Inhibitory Activities of Phenolic Compounds in Different Extracts from the Golden Oyster Mushroom, Pleurotus citrinopileatus (Agaricomycetes). Int. J. Med. Mushrooms 2019, 21, 865–874. [Google Scholar] [CrossRef]
  30. Xu, Q.; Wang, H.; Li, T.; Chen, L.; Zheng, B.; Liu, R.H. Comparison of phenolics, antioxidant, and antiproliferative activities of two Hypsizygus marmoreus varieties. J. Food Sci. 2020, 85, 2227–2235. [Google Scholar] [CrossRef]
  31. Sárközy, A.; Kúsz, N.; Zomborszki, Z.P.; Csorba, A.; Papp, V.; Hohmann, J.; Vanyolos, A. Isolation and Characterization of Chemical Constituents from the Poroid Medicinal Mushroom Porodaedalea chrysoloma (Agaricomycetes) and Their Antioxidant Activity. Int. J. Med. Mushrooms 2020, 22, 125–131. [Google Scholar] [CrossRef] [PubMed]
  32. Chu, J.; Ming, Y.; Cui, Q.; Zheng, N.; Yang, S.; Li, W.; Gao, H.; Zhang, R.; Cheng, X. Efficient extraction and antioxidant activity of polyphenols from Antrodia cinnamomea. BMC Biotechnol. 2022, 22, 9. [Google Scholar] [CrossRef]
  33. Li, Y.; Liang, H.; Zhou, D.; **ng, Y.; Chen, J. Phenolics, Flavonoids Content and Antioxidant Activities of Tuber indicum at Different Maturity Stages. Chem. Biodivers. 2022, 19, e202100830. [Google Scholar] [CrossRef] [PubMed]
  34. Yan, Y.M.; Dong, X.P.; Li, Y.; Cheng, Y.X. Chemical constituents in Yi Medicine Blaps japonensis and their cytotoxic activities. Zhongcaoyao 2013, 44, 269–271. [Google Scholar]
  35. Reuben, R.C.; Khanna, P.L.; Gazitt, Y.; Breslow, R.; Rifkind, R.A.; Marks, P.A. Inducers of erythroleukemic differentiation. Relationship of structure to activity among planar-polar compounds. J. Biol. Chem. 1978, 253, 4214–4218. [Google Scholar] [CrossRef]
  36. Ouchida, R.; Kusuhara, M.; Shimizu, N.; Hisada, T.; Makino, Y.; Morimoto, C.; Handa, H.; Ohsuzu, F.; Tanaka, H. Suppression of NF-κB-dependent gene expression by a hexamethylene bisacetamide-inducible protein HEXIM1 in human vascular smooth muscle cells. Genes Cells 2003, 8, 95–107. [Google Scholar] [CrossRef]
  37. Yue, Y.; Chen, C.; Zhong, K.; Wu, Y.; Gao, H. Purification, Fermentation Optimization, and Antibacterial Activity of Pyrrole-2-carboxylic Acid Produced by an Endophytic Bacterium, Bacillus cereus ZBE, Isolated from Zanthoxylum bungeanum. Ind. Eng. Chem. Res 2022, 61, 1267–1276. [Google Scholar] [CrossRef]
  38. Chen, H.; Yang, C.; Ke, T.; Zhou, M.; Li, Z.; Zhang, M.; Gong, G.; Hou, T. Antimicrobial activity of secondary metabolites from Streptomyces sp. K15, an endophyte in Houttuynia cordata Thunb. Nat. Prod. Res. 2015, 29, 2223–2225. [Google Scholar] [CrossRef]
  39. Munteanu, I.G.; Apetrei, C. Analytical Methods Used in Determining Antioxidant Activity: A Review. Int. J. Mol. Sci. 2021, 22, 3380. [Google Scholar] [CrossRef]
  40. Di Meo, S.; Venditti, P. Evolution of the Knowledge of Free Radicals and Other Oxidants. Oxid. Med. Cell. Longev. 2020, 2020, 9829176. [Google Scholar] [CrossRef] [Green Version]
  41. Zarkovic, N. Antioxidants and Second Messengers of Free Radicals. Antioxidants 2018, 7, 158. [Google Scholar] [CrossRef] [Green Version]
  42. Xu, Z.; Fu, L.; Feng, S.; Yuan, M.; Huang, Y.; Liao, J.; Zhou, L.; Yang, H.; Ding, C. Chemical Composition, Antioxidant and Antihyperglycemic Activities of the Wild Lactarius deliciosus from China. Molecules 2019, 24, 1357. [Google Scholar] [CrossRef] [Green Version]
  43. El Sayed, N.S.; Ghoneum, M.H. Antia, a Natural Antioxidant Product, Attenuates Cognitive Dysfunction in Streptozotocin-Induced Mouse Model of Sporadic Alzheimer’s Disease by Targeting the Amyloidogenic, Inflammatory, Autophagy, and Oxidative Stress Pathways. Oxid. Med. Cell. Longev. 2020, 2020, 4386562. [Google Scholar] [CrossRef]
  44. Song, X.; Shen, Q.; Liu, M.; Zhang, C.; Zhang, L.; Ren, Z.; Wang, W.; Dong, Y.; Wang, X.; Zhang, J.; et al. Antioxidant and hepatoprotective effects of intracellular mycelium polysaccharides from Pleurotus geesteranus against alcoholic liver diseases. Int. J. Biol. Macromol. 2018, 114, 979–988. [Google Scholar] [CrossRef]
  45. Stastny, J.; Marsik, P.; Tauchen, J.; Bozik, M.; Mascellani, A.; Havlik, J.; Landa, P.; Jablonsky, I.; Treml, J.; Herczogova, P.; et al. Antioxidant and Anti-Inflammatory Activity of Five Medicinal Mushrooms of the Genus Pleurotus. Antioxidants 2022, 11, 1569. [Google Scholar] [CrossRef]
  46. Liu, Y.; Zhou, Y.; Liu, M.; Wang, Q.; Li, Y. Extraction optimization, characterization, antioxidant and immunomodulatory activities of a novel polysaccharide from the wild mushroom Paxillus involutus. Int. J. Biol. Macromol. 2018, 112, 326–332. [Google Scholar] [CrossRef]
  47. Yuan, F.; Gao, Z.; Liu, W.; Li, H.; Zhang, Y.; Feng, Y.; Song, X.; Wang, W.; Zhang, J.; Huang, C.; et al. Characterization, Antioxidant, Anti-Aging and Organ Protective Effects of Sulfated Polysaccharides from Flammulina velutipes. Molecules 2019, 24, 3517. [Google Scholar] [CrossRef] [Green Version]
  48. Altannavch, N.; Zhou, X.; Khan, M.A.; Ahmed, A.; Naranmandakh, S.; Fu, J.J.; Chen, H.C. Anti-oxidant and Anticancerous Effect of Fomitopsis officinalis (Vill. ex Fr. Bond. et Sing) Mushroom on Hepatocellular Carcinoma Cells In Vitro through NF-kB Pathway. Anticancer Agents Med. Chem. 2022, 22, 1561–1570. [Google Scholar]
  49. Garcia, J.; Rodrigues, F.; Castro, F.; Aires, A.; Marques, G.; Saavedra, M.J. Antimicrobial, Antibiofilm, and Antioxidant Properties of Boletus edulis and Neoboletus luridiformis Against Multidrug-Resistant ESKAPE Pathogens. Front. Nutr. 2021, 8, 773346. [Google Scholar] [CrossRef]
  50. Smeriglio, A.; D’Angelo, V.; Denaro, M.; Trombetta, D.; Raimondo, F.M.; Germanò, M.P. Polyphenol Characterization, Antioxidant and Skin Whitening Properties of Alnus cordata Stem Bark. Chem. Biodivers. 2019, 16, e1900314. [Google Scholar] [CrossRef]
  51. Kong, S.; Choi, H.R.; Kim, Y.J.; Lee, Y.S.; Park, K.C.; Kwak, S.Y. Milk Protein-Derived Antioxidant Tetrapeptides as Potential Hypopigmenting Agents. Antioxidants 2020, 9, 1106. [Google Scholar] [CrossRef] [PubMed]
  52. Nataraj, A.; Govindan, S.; Ramani, P.; Subbaiah, K.A.; Sathianarayanan, S.; Venkidasamy, B.; Thiruvengadam, M.; Rebezov, M.; Shariati, M.A.; Lorenzo, J.M.; et al. Antioxidant, Anti-Tumour, and Anticoagulant Activities of Polysaccharide from Calocybe indica (APK2). Antioxidants 2022, 11, 091694. [Google Scholar] [CrossRef]
  53. Okolo, K.O.; Orisakwe, O.E.; Siminialayi, I.M. Nephroprotective and Antioxidant Effects of King Tuber Oyster Medicinal Mushroom, Pleurotus tuber-regium (Agaricomycetes), on Carbon Tetrachloride-Induced Nephrotoxicity in Male Sprague Dawley Rats. Int. J. Med. Mushrooms 2018, 20, 419–429. [Google Scholar] [CrossRef] [PubMed]
  54. Quy, T.N.; Xuan, T.D. Xanthine Oxidase Inhibitory Potential, Antioxidant and Antibacterial Activities of Cordyceps militaris (L.) Link Fruiting Body. Medicines 2019, 6, 20. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  55. Li, W.; Liu, Q.; Li, S.; Zheng, Y. New Sesquiterpenoids from the Fermented Broth of Termitomyces albuminosus and their Anti-Acetylcholinesterase Activity. Molecules 2019, 24, 2980. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  56. Kassim, N.K.; Lim, P.C.; Ismail, A.; Awang, K. Isolation of antioxidative compounds from Micromelum minutum guided by preparative thin layer chromatography-2,2-diphenyl-1-picrylhydrazyl (PTLC-DPPH) bioautography method. Food Chem. 2019, 272, 185–191. [Google Scholar] [CrossRef]
  57. Blois, M.S. Antioxidant Determinations by the Use of a Stable Free Radical. Nature 1958, 181, 1199–1200. [Google Scholar] [CrossRef]
  58. Brand-Williams, W.; Cuvelier, M.E.; Berset, C. Use of a free radical method to evaluate antioxidant activity. LWT-Food Sci. Technol. 1995, 28, 25–30. [Google Scholar] [CrossRef]
  59. Dolomanov, O.V.; Bourhis, L.J.; Gildea, R.J.; Howard, J.A.K.; Puschmann, H. OLEX2: A complete structure solution, refinement and analysis program. J. Appl. Crystallogr. 2009, 42, 339–341. [Google Scholar] [CrossRef]
Molecules 28 04760 g001 550
Figure 1. The chemistry structures of compounds 1–10 and the crystal form of compound 4 and 10.
Figure 1. The chemistry structures of compounds 1–10 and the crystal form of compound 4 and 10.
Molecules 28 04760 g001
Molecules 28 04760 g002 550
Figure 2. TLC-bioautography test for DPPH radical scavenging activity of compounds 1–10.
Figure 2. TLC-bioautography test for DPPH radical scavenging activity of compounds 1–10.
Molecules 28 04760 g002
Molecules 28 04760 g003 550
Figure 3. DPPH radical scavenging activity of compounds. (The associated p value of SC50 of these compounds estimated by using Probit analysis in SPSS 18.0 were less than 0.01, except for 2 with p value of 0.064).
Figure 3. DPPH radical scavenging activity of compounds. (The associated p value of SC50 of these compounds estimated by using Probit analysis in SPSS 18.0 were less than 0.01, except for 2 with p value of 0.064).
Molecules 28 04760 g003
Table
Table 1. The 1H and 13C NMR Spectral Data of 1–2.
Table 1. The 1H and 13C NMR Spectral Data of 1–2.
Position12
δH (Mult, J in Hz) *δC **δH (Mult, J in Hz) *δC **
1 169.2 167.2
36.36 (s, 1H)103.210.33 (s, 1H)196.6
3a 123.4 113.3
4 155.5 165.4
56.68 (d, J = 2.0 Hz, 1H)109.66.96 (d, J = 2.4 Hz, 1H)112.1
6 162.5 167.0
76.74 (d, J = 2.0 Hz, 1H)103.16.52 (d, J = 2.4 Hz, 1H)106.7
7a 131.3 137.8
83.49 (s, 3H)56.23.93 (s, 3H)53.4
* Recorded at 600 MHz in (CD3)2OD, λ in ppm; ** Recorded at 150 MHz in (CD3)2OD; λ in ppm.
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Wang, F.; Tan, J.; Jiang, R.; Li, F.; Zheng, R.; Yu, L.; Luo, L.; Zheng, Y. DPPH Radical Scavenging Activity of New Phenolics from the Fermentation Broth of Mushroom Morehella importuna. Molecules 2023, 28, 4760. https://doi.org/10.3390/molecules28124760

AMA Style

Wang F, Tan J, Jiang R, Li F, Zheng R, Yu L, Luo L, Zheng Y. DPPH Radical Scavenging Activity of New Phenolics from the Fermentation Broth of Mushroom Morehella importuna. Molecules. 2023; 28(12):4760. https://doi.org/10.3390/molecules28124760

Chicago/Turabian Style

Wang, Feifei, Jie Tan, Ruixiang Jiang, Feifei Li, Renqing Zheng, Linjun Yu, Lianzhong Luo, and Yongbiao Zheng. 2023. "DPPH Radical Scavenging Activity of New Phenolics from the Fermentation Broth of Mushroom Morehella importuna" Molecules 28, no. 12: 4760. https://doi.org/10.3390/molecules28124760

Article Metrics

Back to TopTop