Rare β-Resorcylic Acid Derivatives from a Halophyte-Associated Fungus Colletotrichum gloeosporioides JS0419 and Their Antifungal Activities

Six new β-resorcylic acid derivatives (1–5 and 7) were isolated from a halophyte-associated fungus, Colletotrichum gloeosporioides JS0419, together with four previously reported β-resorcylic acid lactones (RALs). The relative and absolute stereochemistry of 1 was completely established by a combination of spectroscopic data and chemical reactions. The structures of the isolated compounds were elucidated by analysis of HRMS and NMR data. Notably, compounds 1–3 had a β-resorcylic acid harboring a long unesterified aliphatic side chain, whereas the long aliphatic chains were esterified to form macrolactones in 4–9. Among the isolated compounds, monocillin I and radicicol showed potent antifungal activities against Cryptococcus neoformans, comparable to clinically available antifungal agents and radicicol showed weak antifungal activity against Candida albicans. These findings provide insight into the chemical diversity of fungal RAL-type compounds and their pharmacological potential.


Introduction
A variety of bioactive metabolites have been reported from plant-associated microorganisms [1]. Notably, the microorganisms have been reported to show mutualistic symbiosis with halophytes to help them survive in high saline conditions [2]. The discovery of bioactive secondary metabolites and the elucidation of relationship(s) between microbes and the host halophyte have become an attractive research aim, owing to their functional diversity [3].
As part of our efforts to discover new bioactive compounds from halophyte-associated fungi, we investigated the chemical profiles of fungal extracts using liquid chromatography coupled with a diode array detector and mass spectrometry (LC-DAD-MS). We selected the fungal strain C. gloeosporioides JS0419, isolated from the halophyte Suaeda japonica, for further analysis, because it produces a variety of secondary metabolites with unique UV patterns [21][22][23][24]. After fermentation and subsequent purification, we identified two distinct classes of new compounds in C. gloeosporioides JS0419. The first class comprised glycosylated cyclic lipodepsipeptides (colletotrichamides A-E) with moderate neuroprotective activities against glutamate-induced neurotoxicity in hippocampal HT22 cells, which we recently reported [25]. The other class comprised polyketides represented by β-RALs, which included non-lactonized and lactonized lactones. Chemical investigation of C. gloeosporioides JS0419 isolated from the leaves of S. japonica led to the isolation of nine polyketides (1-9). Among them, we identified the previously reported compounds, nordinonediol (6) [10], monocillin I (8) [26], and radicicol (9) [27] by comparing their spectroscopic data with those in previous reports. Here, we report the isolation, structural elucidation, and bioactivity of these polyketide compounds ( Figure 1).

Structure Determination of the Compounds
Compound 1 (colletogloeic acid A) was isolated as a yellowish amorphous solid and analyzed for its molecular formula (C 18 H 26 O 8 ) based on (+)HRESIMS (observed [M + Na − H 2 O] + at m/z 375.1420, calcd for m/z 375.1414), which was supported by 1 H-NMR, 13 C-NMR, and heteronuclear single quantum coherence (HSQC) data (Table 1). The 1 H NMR spectrum of 1 displayed one olefinic proton (δ H 6.31), two meta-coupled aromatic protons (δ H 6.29 (d, J = 2.0 Hz) and 6.28 (d, J = 2.0 Hz)), three carbinol protons (δ H 4.00, 3.66, and 3.42), ten aliphatic protons (δ H 2.52-1.42), and one methyl proton (δ H 1.20). The 13 C NMR spectrum together with the HSQC data of 1 revealed eighteen carbons, one carboxylic acid carbon (δ C 168.1); eight sp 2 carbons (δ C 167.7~99.6), three of which were protonated; three oxygenated sp 3 methine carbons (δ C 76.1, 72.9, and 65.6), five aliphatic sp 3 carbons (δ C 42. 4, 34.1, 33.4, 28.1, and 26.5), and one methyl carbon (δ C 24.6). The presence of β-resorcylic acid (2,4-dihydroxybenzoic acid) was determined based on the meta-coupled proton signals (J = 2.0 Hz) together with two oxygenated sp 2 carbons and a carboxylic carbon signal. Interpretation of 1 H-1 H COSY allowed the assignment of a long aliphatic Mar. Drugs 2022, 20, 195 3 of 15 3 ] corresponding to C-10 to C-18, and we confirmed its connection with the β-resorcylic acid moiety according to the HMBC spectrum ( Figure 2). Although there are many overlapping signals for CH 2 in the 1 H NMR spectra, all the methylene protons and carbons could be assigned by interpreting HMBCs from isolated methylene protons (H 2 -10) and oxymethines (H-14, H-15, and H-17), as shown in Figure 2. HMBCs of an olefinic proton H-8 (δ H 6.31) of the aliphatic chain with C-2 (δ C 99.6) and C-6 (δ C 103.9) of the β-resorcylic acid moiety indicated that the long-chain moiety was attached to C-7. Additionally, we considered the possibility that the long aliphatic chain of 1 was macrolactonylated through an ester bond between C-1 and C-17, as many RAL-type compounds have previously been reported in fungi. However, it was more probable to have a free hydroxyl group at C-17 rather than to be esterified to form a macrolactone ring due to the chemical resonance of H-17 (δ H 4.0) in the long chain. Evidently, no HMBC correlation from H-17 to the carbonyl carbon C-1 supported the presence of free hydroxyl groups. Although a number of RAL-type compounds have been reported from fungi, to our knowledge, this is a rare case to isolate β-resorcylic acid with an unesterified long aliphatic side chain. We then confirmed the presence of β-resorcylic acid with a non-esterified aliphatic chain by a simple chemical reaction. Compound 1 was acetylated to confirm the number of exchangeables in the side chain. Acetylation of 1 caused downfield shifts of three oxymethine protons (H-14, H-15, and H-17) in the aliphatic side chain by~1 ppm (compared with those in compound 1), thus completing the planar structure of compound 1 ( Figure S7). Compound 1 was enolated at C-8 and C-9, whereas most of the previously reported RAL-type compounds have a keto group at C-9. To the best of our knowledge, the only precedent example of RAL with an enol group at C-8 and C-9 is pochonin P [26]. Therefore, the present study represents the second report on the natural occurrence of β-resorcylic acid derivatives with an enol chain.  (14, 9.8, 2.8) 42.0, CH 2 1.69, ddd (14.5, 5.5, 3. shifts of three oxymethine protons (H-14, H-15, and H-17) in the aliphatic side chain by ~1 ppm (compared with those in compound 1), thus completing the planar structure of compound 1 ( Figure S7). Compound 1 was enolated at C-8 and C-9, whereas most of the previously reported RAL-type compounds have a keto group at C-9. To the best of our knowledge, the only precedent example of RAL with an enol group at C-8 and C-9 is pochonin P [26]. Therefore, the present study represents the second report on the natural occurrence of β-resorcylic acid derivatives with an enol chain.   Compound 2 (colletogloeic acid B) was a yellowish amorphous solid and determined to be chlorinated according to the characteristic isotopic pattern in its MS spectrum. Its molecular formula was found to be C 18 H 25 ClO 9 based on (+)HRESIMS (observed [M + Na − H 2 O] + at m/z 425.1010) and spectroscopic data. This chlorinated compound had one aromatic proton, one olefinic proton, and four oxymethine proton signals. The C-6 of the β-resorcylic acid moiety was confirmed to be chlorinated based on HMBCs of the aromatic proton H-4 (δ H 6.42) and the olefinic proton H-8 (δ H 6.72) with C-6 (δ C 108.3). Detailed analysis of 1 H-1 H COSY revealed the presence of a 1,3,4,6-tetraol system in the aliphatic chain instead of the 1,2,4-triol system shown in 1. The position of the additional carbinol group (δ H 3.91; δ C 70.4) was confirmed to be C-12 by HMBCs of H 2 -10 (δ H 2.75 and 2.67) with C-8 (δ C 101.9) and C-12 (δ C 70.4), suggesting its planar structure ( Figure 2 Although the NMR spectral data of 3 were similar to those of 1, an additional double bond in trans at δ H 6.54 (1H, dt, J = 15.5, 2.5 Hz) and 6.16 (1H,d,J = 15.5 Hz) and the lack of the oxygenated methine group revealed a distinct structural difference between 1 and 3. The additional double bond was positioned at C-10 and C-11 based on the HMBC correlations of H-10 (δ H 6.16) with C-8 (δ C 106.0) and C-12 (δ C 33.8), as well as H-11 (δ H 6.54) with C-9 (δ C 153.5) and C-13 (δ C 26.1). Additionally, a spin system CH(O)-CH 2 -CH(O)-CH 3 corresponding to C15(O)-C16-C17(O)-C18 obtained by 1 H-1 H COSY indicated hydroxylation at C-15 and C-17, which was also supported by HMBCs.
Compound 4 (Colletoresorcylic lactone) was a yellowish amorphous solid and its molecular formula was found to be C 18 H 24 O 7 based on the HSQC NMR and (+)HRESIMS (observed [M + Na] + at m/z 375.1419). Compound 4 showed different UV profiles from compounds 1-3. While 1-3 had a strong UV absorption at 244 nm and a medium absorption at 320 nm, 4 showed three UV absorptions at 211, 262, and 298 nm, suggestive of a lactonized resorcylic acid. The 1 H, 13 C, and 1 H-1 H COSY NMR data of 4 were similar to those of 1 in the presence of the 2,4-dihydroxybenzoic acid moiety and a 1,2,4-triol group, in the aliphatic chain. A methylene group at δ H 4.48 and δ H 3.82 and a ketone carbon (δ C 212.5) in 4 instead of the enol group indicated that 4 had a keto group. Notably, compound 4 had an oxymethine proton signal at δ H 4.95 unlike previous compounds 1-3, suggesting that 4 had an ester group. An HMBC correlation of the oxymethine proton δ H 4.95 with the carboxylic carbon (C-1) at δ C 171.9 in addition to two oxymethine carbons (C-14 and C-17) indicated the presence of the 12-membered lactone ring. Additionally, HMBCs of the methylene protons H 2 -8 (δ H 4.48 and 3.82) with C-2 (δ C 107.5), C-6 (δ C 113.9), and C-9 (δ C 212.5) confirmed the presence of the keto group at C-9.
Compound 5 had the molecular formula C 18 H 23 ClO 7 according to (+)HRESIMS data and it exhibited a similar UV pattern to 4, characteristic for RAL-type compounds. Moreover, this was supported by an HMBC of H-17 (δ H 5.43) in a long chain with the carboxylic carbon C-1 (δ C 172.5). Chlorination at C-6 of the β-resorcylic acid moiety was assigned based on its HMBCs of H-4 and H 2 -8 with C-6 along with typical isotopic mass fragmentation. Compound 5 was structurally similar to the known compound nordinonediol (6), except for the presence of chlorine at C-6 of the RAL moiety.
Compound 7 was a yellowish amorphous solid with a molecular formula of C 23 H 28 O 12 based on (+)HRESIMS data. Additionally, we determined 7 to be a RAL-type compound based on its characteristic UV absorption, which was also supported by NMR resonances for an oxymethine group (δ H 5.38; δ C 72.3) representative of a lactone. Unlike previous RAL-type compounds 5 and 6, 7 had a conjugated double bond, which was confirmed by proton resonances and their coupling constants (δ H 6.08 ( Table 2). HMBCs of the olefinic protons at δ H 6.08 and 7.59 with the ketone at C-9 allowed the connection of the conjugated double bond with the ketone C-9. Two oxymethines that appeared in a relatively higher field (δ H 3.36 (m, H-14), δ C 56.6 (C-14); 3.08 (dt, J = 8.5, 3.0 Hz, H-15), 56.9 (C-15)) than the diol group were indicative of epoxide functionality. Moreover, the degrees of unsaturation suggested by the molecular formula were supportive of the epoxide ring. The conjugated double bond was then connected to the epoxide group according to 1 H-1 H COSY and HMBC correlations of the epoxide protons, H-14 and H-15 with C-13. The geometries of the double bonds (C-10/C-11, C-11/C-12, and C-12/C-13) were determined to be E, Z, and Z configurations, respectively, based on their coupling constants. Additionally, 1 H and 13 .2) suggested the presence of a pentose, which was confirmed to be ribose by comparing its NMR resonances with those of the references [28]. We confirmed the location of the ribose by an HMBC from the anomeric proton H-1 (δ H 5.65) with C-5 (δ C 156.5). The ribose moiety was in an α configuration based on the coupling constant of the anomeric proton ( 3 J HH = 4.5 Hz) [28].

Establishment of Stereochemistry
Colletogloeic acids A-C (1-3) have several stereogenic centers in their chain. We employed comprehensive spectroscopic analysis and various chemical reactions to establish their relative and absolute stereochemistries. The relative configurations of the consecutive stereogenic centers in the aliphatic chain were initially determined by measuring the NMR data of their acetonide derivatives. Colletogloeic acid A (1) was derivatized with 2,2-dimethoxy propane to obtain the acetonide derivative 1a. By interpreting HMBC data of 1a together with 1D NMR, 1,2-diol at C-14 and C-15 was confirmed to be acetonylated. The 13 C resonances for the two methyl groups of the acetonide derivative, which appeared at δ C 29.1 and 26.3, indicated a syn configuration of the dihydroxyl groups at C-14 and C-15 [29,30]. At this point, the relative stereochemistry of the hydroxyl group at C-17 remains unknown. To establish the relative stereochemistry at C-15 and C-17 in the acyclic chain, we employed J-based configuration analysis (JBCA). Long-range heteronuclear coupling constants ( 3 J CH and 2 J CH values) were acquired using a hetero-half-filtered TOCSY experiment [31]. Distinguishable methylene protons at C-16 allowed consecutive JBCA from C-15 to C-17 through C-16. Small 3 J H-15, C-17 (1.7 Hz), small 2 J C-15, H-16a (0.6 Hz), and large 2 J C-15, H-16b (6.3 Hz) indicated that the hydroxyl group at C-15 and one of the methylene protons at C-16 in the lower field (H-16a) were in an anti orientation (Figure 3b), whereas small 3 J H-17, C-15 (2.9 Hz), large 2 J C-17, H-16a (6.6 Hz), and small 2 J C-17, H-16b (0.7 Hz) suggested that the hydroxyl groups at C-17 and H-16a were in the same orientation ( Figure 3c). Thus, the hydroxyl groups at C-15 and C-17 were determined as having opposite orientations. This result is consistent with the assignment predicted by the application of Kishi's universal NMR database [32]. The relative stereochemistry of the diol groups at C-14 and C-15 established by JBCA was in agreement with that of the chemical reaction for its acetonide derivative. The small values of 2 J C-14, H-15 (0.8 Hz) and 2 J C-15, H-14 (1.8 Hz) indicated a threo configuration of both hydroxyl groups at C-14 and C-15 (Figure 3a), which agreed with that obtained from the acetonide reaction.
To determine the absolute configurations of C-14, C-15, and C-17 in 1, we attempted a modified Mosher's method [33]. Acetonide product 1a was treated with Rand S-MTPA-Cl to obtain the Sand R-MTPA derivatives 1b and 1c, respectively. The interpretation of 1 H NMR and 1 H-1 H COSY NMR of these MTPA ester derivatives allowed the assignment of the absolute configuration of C-17. A positive ∆δ S-R value for H 3 -18 and negative ∆δ S-R values for H-15 and H 2 -16 indicated that C-17 had an R configuration, subsequently establishing both S configurations for C-14 and C-15. The relative configurations of the consecutive stereogenic centers in compound 2 were established by JBCA together with the acetonide reaction. Colletogloeic acid B (2) was derivatized with 2,2-dimethoxy propane to obtain the acetonide derivative 2a. HMBC data together with 1D NMR for 2a indicated that two 1,3acetonide groups were attached at two positions; one at between C-12 and C-14, the other at between C-15 and C-17 ( Figure 4C). Both 1,3-diol groups corresponding to C-12/C-14 and C-15/C-17 were identified as a syn configuration based on the 13 C resonances for methyl groups of the acetonide derivatives at δ C 30.4/20.23 and δ C 30.5/20.17, respectively [34]. We then established the relative configuration of diols at C-14 and C-15 by JBCA, and the vicinal coupling constants for H-14 and H-15 (5.5 Hz) were obtained via decoupling experiments. Small values of 3 J C-13, H-15 (2.24 Hz), 2 J C-14, H-15 (0.4 Hz), and 2 J C-15, H-14 (3.2 Hz) indicated a syn relationship between H-14 and H-15 (Figure 3d), suggesting that all the hydroxyl groups in the side chain of 3 had syn configurations. The absolute stereochemistry of 3 could not be established since its Rand S-MTPA esters were decomposed. Compounds 4, 5, and 7 with a 1,2,4-triol system in the aliphatic chain were proposed to have the same stereochemistry as 1 based on the NMR shifts, coupling constants, and their synthetic origin.
Mar. Drugs 2022, 20, x 7 of 15 NMR database [32]. The relative stereochemistry of the diol groups at C-14 and C-15 established by JBCA was in agreement with that of the chemical reaction for its acetonide derivative. The small values of 2 JC-14, H-15 (0.8 Hz) and 2 JC-15, H-14 (1.8 Hz) indicated a threo configuration of both hydroxyl groups at C-14 and C-15 (Figure 3a), which agreed with that obtained from the acetonide reaction. To determine the absolute configurations of C-14, C-15, and C-17 in 1, we attempted a modified Mosher's method [33]. Acetonide product 1a was treated with R-and S-MTPA-Cl to obtain the S-and R-MTPA derivatives 1b and 1c, respectively. The interpretation of 1 H NMR and 1 H-1 H COSY NMR of these MTPA ester derivatives allowed the assignment of the absolute configuration of C-17. A positive ∆δS-R value for H3-18 and negative ∆δS-R values for H-15 and H2-16 indicated that C-17 had an R configuration, subsequently establishing both S configurations for C-14 and C-15. The relative configurations of the consecutive stereogenic centers in compound 2 were established by JBCA together with the acetonide reaction. Colletogloeic acid B (2) was derivatized with 2,2-dimethoxy propane to obtain the acetonide derivative 2a. HMBC data together with 1D NMR for 2a indicated that two 1,3-acetonide groups were attached at two positions; one at between C-12 and C-14, the other at between C-15 and C-17 ( Figure 4C). Both 1,3-diol groups corresponding to C-12/C-14 and C-15/C-17 were identified as a syn configuration based on the 13 C resonances for methyl groups of the acetonide derivatives at δC 30.4/20.23 and δC 30.5/20.17, respectively [34].  (Figure 3d), suggesting that all the hydroxyl groups in the side chain of 3 had syn configurations. The absolute stereochemistry of 3 could not be established since its R-and S-MTPA esters were decomposed. Compounds 4, 5, and 7 with a 1,2,4-triol system in the aliphatic chain were proposed to have the same stereochemistry as 1 based on the NMR shifts, coupling constants, and their synthetic origin.

Antifungal Activity
We evaluated the antifungal activities of the isolated polyketides by measuring their MIC values based on EUCAST guidelines ( Figure 5). Among the isolated compounds, 8 and 9 showed potent in vitro antifungal activities against C. neoformans H99, with MIC values (12.5 µM) almost equivalent to those of amphotericin B (AMB) and fluconazole (FCZ) (12.5 µM), which are clinically used for the treatment of systemic cryptococcosis. By contrast, only compound 9 showed weak antifungal activities (MIC: 200 µM) against C. albicans SC5314, which was much less potent than AMB (12.5 µM) and FCZ (25 µM)  (1b and 1c) for the acetonide derivative (1a) and its ∆δ S−R values observed in 1 H NMR (CD 3 OD). (C) Synthesis of 1,2−diol−acetonide for 2 and its 13 C shifts in CD 3 OD (2a).

Antifungal Activity
We evaluated the antifungal activities of the isolated polyketides by measuring their MIC values based on EUCAST guidelines ( Figure 5). Among the isolated compounds, 8 and 9 showed potent in vitro antifungal activities against C. neoformans H99, with MIC values (12.5 µM) almost equivalent to those of amphotericin B (AMB) and fluconazole (FCZ) (12.5 µM), which are clinically used for the treatment of systemic cryptococcosis. By contrast, only compound 9 showed weak antifungal activities (MIC: 200 µM) against C. albicans SC5314, which was much less potent than AMB (12.5 µM) and FCZ (25 µM) used as positive controls. Collectively, these results indicated that compounds 8 and 9 displayed potent in vitro antifungal activity.

Fungal Strain
The fungal strain JS419 was isolated from S. japonica Makino, which was collected from a swamp in Suncheon, South Korea, in September 2011. The tissues of this plant were cut into small pieces (0.5 × 0.5 cm) and rinsed sequentially for 1 min with 2% sodium hypochlorite, 70% ethanol, and sterilized distilled water to remove external microorganisms. After air drying, the sterilized plant tissues were incubated on malt extract agar (MEA) medium (4 g yeast extract, 10 g malt extract, 4 g potato dextrose broth, and 18 g agar per 1 L sterilized distilled water) with 50 ppm kanamycin, 50 ppm chloramphenicol, and 50 ppm Rose Bengal at 22 °C for 7 days. The actively growing fungus was transferred onto potato dextrose agar (PDA) medium (24 g potato dextrose broth and 18 g agar per 1 L Figure 5. Antifungal activities of compounds 8 and 9. The heat map of EUCAST MIC test results for compounds 8 and 9 against C. albicans SC5314 and C. neoformans H99. Fungal cells were prepared, as described, and incubated at 35 • C for 2 days in 96-well microtiter plates containing MOPSbuffered RPMI-1640 medium with two-fold-diluted natural compounds. Amphotericin B (AMB) and fluconazole (FCZ) were used as positive controls.

Fungal Strain
The fungal strain JS419 was isolated from S. japonica Makino, which was collected from a swamp in Suncheon, South Korea, in September 2011. The tissues of this plant were cut into small pieces (0.5 × 0.5 cm) and rinsed sequentially for 1 min with 2% sodium hypochlorite, 70% ethanol, and sterilized distilled water to remove external microorganisms. After air drying, the sterilized plant tissues were incubated on malt extract agar (MEA) medium (4 g yeast extract, 10 g malt extract, 4 g potato dextrose broth, and 18 g agar per 1 L sterilized distilled water) with 50 ppm kanamycin, 50 ppm chloramphenicol, and Mar. Drugs 2022, 20, 195 9 of 15 50 ppm Rose Bengal at 22 • C for 7 days. The actively growing fungus was transferred onto potato dextrose agar (PDA) medium (24 g potato dextrose broth and 18 g agar per 1 L sterilized distilled water). The fungal strain was identified as C. gloeosporioides based on the internal transcribed spacer sequences by one of the authors (S. K.) and was deposited as a 20% glycerol stock in a liquid nitrogen tank at the Wildlife Genetic Resources Bank of the National Institute of Biological Resources (Incheon, Korea).

Cultivation and Extraction of the Fungal Strain
The JS419 strain was cultured on a solid PDA medium for 7 days at room temperature. Agar plugs were cut into small pieces (0.5 × 0.5 cm) under aseptic conditions and inoculated on a solid rice medium (60 g rice, 90 mL distilled water per 500 mL Erlenmeyer flask). After incubation at room temperature for 30 days, 200 mL of ethyl acetate was added to each culture flask (50 × 500 mL Erlenmeyer flasks) and placed for 1 day prior to extraction. They were then filtered to separate the supernatants from the solid mycelia, after which the supernatants were evaporated under reduced pressure at 35 • C to obtain the extract (110 g).

Conclusions
In this study, we isolated nine β-resorcylic acid derivatives, represented by RALs, from cultures of C. gloeosporioides JS0419. While most RALs reported to date feature a 14-membered macrocyclic ring fused to β-resorcylic acid, compounds 1-3 harbored β-resorcylic acid with a long unesterified aliphatic side chain and compound 4 had a 12-membered macrocyclic ring fused to the β-resorcylic acid in this chemical investigation. Although several ring-opened β-resorcylic acids have been reported to date [36,37], they mostly had non-oxidized C-9 in the long aliphatic chain. Given the rarity of this observation in polyketides, we did not exclude the possibility of artifacts. For subsequent analyses, we did not use acids or alkalis throughout the isolation and purification procedures. We carefully analyzed the HPLC chromatograms of the initial extracts of this fungal strain, which identified a fair amount of resorcylic acid with a free chain together with RAL-type compounds. Compound 4 was lactonized with the ester linkage between C-15 and C-1 to form a 12-membered lactone ring. Although many RALs had 12-membered rings, to our knowledge, this is the first report of RAL with not methyl but propyl group at C-15. Although many RAL-type compounds have been evaluated for antifungal activities against several plant pathogenic fungi, to our knowledge, there is no report on their activity against C. albicans and C. neoformans causing fatal candidiasis and cryptococcosis, respectively. The known compounds, radicicol, and monocillin I, exhibit potent in vitro antifungal activities, suggesting that an epoxide group, non-sugar moiety, and lactonization of the long polyketide chain could be important for antifungal activities. Optimization of the polyketides through structure-activity relationship studies may further increase their antifungal activity and allow the examination of their in vivo efficacy for treating systemic fungal infections.