In human umbilical vein endothelial cells (HUVECs), Vulpinic acid at doses below 20 µM was found to be noncytotoxic, with maximum cell viability observed at 15 µM. The IC50 of hydrogen peroxide (H2O2) in HUVECs was found to be 215 µM. Pretreatment with 15 µM Vulpinic acid for 24 hours significantly increased cell viability by 41% (p < 0.001) in HUVECs subsequently treated with 215 µM H2O2 for 24 hours. [1]
Vulpinic acid at 15 µM reduced H2O2-induced reactive oxygen species (ROS) production significantly (p < 0.05) to control levels, and alone decreased ROS levels below the control group. [1]
In HUVECs, treatment with 215 µM H2O2 caused actin fragmentation, cell shrinkage, and decreased actin fluorescence intensity, while pretreatment with Vulpinic acid protected cells from these damages, resulting in homogeneous, filamentous actin distribution and normal cell morphology. [1]
Tie2 immunoreactivity (immunohistochemically scored on a 0-5 scale) was decreased in H2O2-treated groups (score 2) compared to control (score 4.5). Pretreatment with Vulpinic acid increased Tie2 immunoreactivity (score 3.5) compared to the H2O2-alone group. [1]
Vulpinic acid showed antibacterial activity against certain aerobic and anaerobic microorganisms. It inhibited clinical isolates of Staphylococcus aureus (MSSA, MRSA susceptible and resistant to mupirocin), Enterococcus faecalis, and Enterococcus faecium at concentrations ranging from 4 to >32 µg/mL, though it was generally less active than usnic acid. It did not inhibit Gram-negative rods (e.g., E. coli, P. aeruginosa) or fungi (e.g., C. albicans) at concentrations ≤ 32 µg/mL. [2]
MICs of Vulpinic acid for Bacteroides and Clostridium species ranged from 16 to >16 µg/mL. [2]

The cytotoxic and protective effects of Vulpinic acid were evaluated using the MTT (methyl thiazolyl diphenyl tetrazolium bromide) assay. HUVECs were seeded at 1x10^5 cells per well in 96-well plates and incubated for 24 hours for adhesion. Cells were then incubated with various concentrations of Vulpinic acid (5-300 µM) for 24 hours. The media was removed, and cells were treated with the IC50 dose of H2O2 (215 µM) for 24 hours. Post-incubation, MTT solution (0.5 mg/mL) was added for 2 hours, then removed, and the formazan salts were dissolved in DMSO. Optical density was read at 570 nm. [1]
Intracellular ROS levels were measured using the DCF-DA method. HUVECs were plated in 96-well plates, incubated for 24 hours, then washed with PBS. 100 µL of DCFH-DA/media solution was added and incubated for 60 minutes. After washing, cells were treated as follows: control (no treatment), Vulpinic acid alone (15 µM for 24h), H2O2 alone (215 µM for 24h), or Vulpinic acid pretreatment (15 µM for 24h) followed by H2O2 (215 µM for 24h). Fluorescence was measured at 480 nm (excitation) and 530 nm (emission). [1]
F-actin filament staining was performed using TRITC-phalloidin. HUVECs on coverslips were treated with Vulpinic acid (15 µM for 24h) and/or H2O2 (215 µM for 24h). Cells were fixed with 3.7% paraformaldehyde, permeabilized with 0.2% Triton X-100, and stained with 5 µg/mL TRITC-phalloidin for 60 minutes, then with DAPI (1 mg/mL) for 30 minutes before examination under a confocal microscope. [1]
Tie2 protein expression was analyzed via immunocytochemistry. HUVECs on coverslips were treated and fixed as above. Endogenous peroxidase was blocked with 3% H2O2, then cells were incubated with Tie2 primary antibody overnight at 4°C, followed by a biotinylated secondary antibody and streptavidin-peroxidase complex. Immunoreactivity was visualized using chromogenic 3-amino-9-ethylcarbazole, and nuclei were counterstained with hematoxylin. A semi-quantitative scoring system (0-5 scale based on percentage of positive cells) was used. [1]
The antimicrobial activity of Vulpinic acid was tested against aerobic and anaerobic bacteria using a standardized microdilution method. An inoculum of 5x10^5 CFU/mL was used in cation-adjusted Mueller-Hinton broth (for aerobes) or Schaedler broth (for anaerobes). Vulpinic acid was dissolved in tetrahydrofuran (0.2% by volume) as a solvent mediator. The maximum achievable concentration in broth was 32 µg/mL due to solubility limitations. The MIC was determined. [2]
The minimum bactericidal concentration (MBC) of Vulpinic acid was determined for enterococci and staphylococci with an MIC of 4 µg/mL for usnic acid. The MBC was defined as the lowest concentration that reduced the original inoculum by ≥99.9%. Vulpinic acid did not show bactericidal activity at the concentrations tested. [2]

In HUVECs, Vulpinic acid was found to be noncytotoxic at doses below 20 µM. At 300 µM, cell viability decreased to 37%. The most effective non-cytotoxic protective dose was 15 µM. [1]

[1]. Vulpinic acid, a lichen metabolite, emerges as a potential drug candidate in the therapy of oxidative stress-related diseases, such as atherosclerosis. Hum Exp Toxicol. 2019 Jun;38(6):675-684.

[2]. In vitro activities of the lichen secondary metabolites vulpinic acid, (+)-usnic acid, and (-)-usnic acid against aerobic and anaerobic microorganisms. Antimicrob Agents Chemother. 1995 Nov;39(11):2541-3.

Fox acid is a butenolactone. It has been reported to be found in the Colombian whitefly, Candelilla simonii, and other organisms with relevant data.

---

Vulpinic acid is a secondary metabolite of lichens, which are symbiotic associations of fungi and algae. [1]
Oxidative stress, induced by reactive oxygen species (ROS) like H2O2, is a basis for many diseases including atherosclerosis, diabetes, cancer, and neurodegenerative diseases. Endothelial damage due to oxidative stress is a key initial step in atherosclerosis development. [1]
The study suggests that Vulpinic acid decreases H2O2-induced oxidative stress and related damage in HUVECs, indicating it may be a potential drug candidate for treating atherosclerosis and other oxidative stress-related diseases. [1]
Lichens have a history of use in traditional medicine, and their secondary metabolites, including usnic acid and Vulpinic acid, were identified as active compounds in the 1950s. [2]
The lack of solubility of Vulpinic acid in water-based nutrient broths limits its testing to a maximum concentration of 32 µg/mL. [2]
Due to its lipophilic properties, higher concentrations of Vulpinic acid might be achievable in ointments, suggesting a possible topical application for eradicating nosocomial S. aureus isolates, though the lack of bactericidal activity may limit its use. [2]

C19H14O5

322.31146

322.084

C, 70.80; H, 4.38; O, 24.82

521-52-8

54690323

Light yellow to yellow a crystalline solid

1.375g/cm3

568.8ºC at 760mmHg

146-148ºC

210.2ºC

1.656

3.096

1

5

4

24

575

0

COC(=O)/C(=C\1/C(=C(C(=O)O1)c2ccccc2)O)/c3ccccc3

OMZRMXULWNMRAE-BMRADRMJSA-N

InChI=1S/C19H14O5/c1-23-18(21)15(13-10-6-3-7-11-13)17-16(20)14(19(22)24-17)12-8-4-2-5-9-12/h2-11,20H,1H3/b17-15+

methyl (2E)-2-(3-hydroxy-5-oxo-4-phenylfuran-2-ylidene)-2-phenylacetate

Vulpinic acid; NSC 5897

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~25 mg/mL (~77.57 mM)

Solubility in Formulation 1: ≥ 1.25 mg/mL (3.88 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 12.5 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

---

Solubility in Formulation 2: ≥ 1.25 mg/mL (3.88 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 12.5 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

---

 (Please use freshly prepared in vivo formulations for optimal results.)