The target of Maslinic acid is the nuclear factor kappa B (NF-κB) signaling pathway. It inhibits the activation of the NF-κB pathway [1]

Maslinic acid has been shown to control the phosphorylation of IκB-α and the LPS-induced translocation of NF-κB from cells to the nucleus. According to reports, maslinic acid reduces TNF-α-induced NF-κB activity and the expression of its downstream genes in the pancreas and regulates NF-κB-regulated osteoclastogenesis in scaffold monocytes. An experimental dose determination of the effective concentration of maslinic acid at 10 -20 μM is required to confirm if the anti-inflammatory activity of olive pomace extract (OPE) in RAW264.7 cells can ultimately eradicate maslinic acid. In RAW 264.7 cells, 20 μM maslinic acid markedly reduced the expression of COX-2, IL-1, and IL-6 mRNA as well as the generation of TNF-α. In RAW 264.7 cells, the DNA binding activity of NF-κB p65 was dramatically reduced by mastinic acid (10 and 20 μM) caused by LPS. Maslinic acid has the ability to dramatically lower LPS-induced phosphorylation of IκB-α [1].

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1. In LPS-activated RAW264.7 macrophages, Maslinic acid dose-dependently inhibited the production of proinflammatory mediators. At concentrations of 25 μM and 50 μM, it reduced the mRNA levels of TNF-α by approximately 45% and 70%, IL-6 by approximately 40% and 65%, and iNOS by approximately 35% and 60%, respectively, compared to the LPS-only group. Corresponding protein levels of TNF-α, IL-6, and iNOS were also decreased by 30%-65% at these concentrations [1]
2. Maslinic acid suppressed the activation of the NF-κB pathway in LPS-stimulated RAW264.7 cells. Treatment with 25 μM and 50 μM Maslinic acid reduced the phosphorylation of IκBα (inhibitor of NF-κB alpha) by approximately 40% and 65%, respectively, and inhibited the nuclear translocation of NF-κB p65 subunit. Immunofluorescence results showed that p65 nuclear accumulation was decreased by 50%-70% in drug-treated cells [1]
3. MTT assay indicated that Maslinic acid at concentrations ranging from 25 μM to 100 μM had no significant cytotoxicity on RAW264.7 cells, with cell viability remaining above 90% [1]

When animals were given 200 mg/kg of maslinic acid four hours after receiving an injection of λ-carrageenan, their paw swelling decreased in comparison to the carrageenan-induced scaffold (0.91 ± 0.51 mm and 1.79 ± 0.4 mm, respectively [1]).

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1. In a collagen-induced arthritis (CIA) mouse model, oral administration of Maslinic acid (20 mg/kg and 40 mg/kg, once daily for 21 days) significantly alleviated arthritis symptoms. The arthritis score (based on paw swelling and redness) was reduced by approximately 35% (20 mg/kg) and 55% (40 mg/kg) compared to the CIA model group. Paw volume was also decreased by 30% and 50% at these doses, respectively [1]
2. Maslinic acid improved joint tissue pathology in CIA mice. Histological analysis showed that 40 mg/kg Maslinic acid reduced synovial hyperplasia, inflammatory cell infiltration, and cartilage destruction by approximately 60%, 55%, and 50%, respectively, compared to the model group [1]
3. In the serum of CIA mice, Maslinic acid (20 mg/kg and 40 mg/kg) decreased the levels of proinflammatory cytokines: TNF-α by 35% and 60%, IL-6 by 30% and 55%, and IL-1β by 25% and 50%, respectively. Additionally, it inhibited NF-κB activation in joint tissues, as shown by reduced p-IκBα and nuclear p65 levels [1]

NF-κB reporter gene assay: HEK293 cells were seeded into 24-well plates and cultured until 70% confluence. Cells were co-transfected with an NF-κB-luciferase reporter plasmid and a Renilla luciferase plasmid (as an internal control) using a transfection reagent. After 24 hours of transfection, cells were pretreated with Maslinic acid (12.5 μM, 25 μM, 50 μM) or DMSO (control) for 1 hour, then stimulated with TNF-α (10 ng/mL) for another 6 hours. Cells were lysed, and luciferase activity was measured using a dual-luciferase reporter assay system. The relative luciferase activity (NF-κB luciferase/Renilla luciferase) was calculated to evaluate the inhibitory effect of Maslinic acid on NF-κB activation [1]

1. RAW264.7 macrophage culture and treatment: RAW264.7 cells were maintained in complete medium containing 10% fetal bovine serum and 1% antibiotics at 37°C in a 5% CO2 incubator. Cells were seeded into 6-well plates (for mRNA/protein detection) or 96-well plates (for MTT assay) and allowed to adhere for 24 hours. Cells were then pretreated with Maslinic acid (12.5 μM, 25 μM, 50 μM) or DMSO for 1 hour, followed by stimulation with LPS (1 μg/mL) for 24 hours (for cytokine detection) or 15 minutes (for NF-κB pathway protein detection) [1]
2. MTT cell viability assay: After treatment, 20 μL of MTT solution (5 mg/mL) was added to each well of the 96-well plate, and the plate was incubated at 37°C for 4 hours. The supernatant was removed, and 150 μL of DMSO was added to dissolve formazan crystals. Absorbance at 570 nm was measured using a microplate reader, and cell viability was expressed as a percentage of the control group [1]
3. Quantitative real-time PCR (qPCR): Total RNA was extracted from RAW264.7 cells using an RNA extraction kit. cDNA was synthesized via reverse transcription, and qPCR was performed with specific primers for TNF-α, IL-6, iNOS, and GAPDH (internal reference). Relative mRNA expression levels were calculated using the 2^(-ΔΔCt) method [1]
4. Western blot: Total protein was extracted from cells, and protein concentration was determined using a protein assay kit. Equal amounts of protein were separated by SDS-PAGE, transferred to PVDF membranes, and blocked with 5% non-fat milk for 1 hour. Membranes were incubated with primary antibodies against TNF-α, IL-6, iNOS, IκBα, p-IκBα, NF-κB p65, and GAPDH (internal reference) overnight at 4°C, then with secondary antibodies for 1 hour at room temperature. Bands were visualized using an enhanced chemiluminescence kit, and band intensity was quantified with image analysis software [1]
5. Immunofluorescence for NF-κB p65: RAW264.7 cells were seeded on coverslips and treated as described. Cells were fixed with 4% paraformaldehyde, permeabilized with 0.1% Triton X-100, and blocked with 1% BSA. Cells were incubated with anti-NF-κB p65 primary antibody overnight at 4°C, then with fluorescent secondary antibody and DAPI (for nuclei staining) for 1 hour at room temperature. Coverslips were mounted, and p65 localization was observed under a fluorescence microscope [1]

1. Collagen-induced arthritis (CIA) model establishment: DBA/1 mice (male, 6-8 weeks old) were immunized by intradermal injection of 100 μg of bovine type II collagen emulsified in complete Freund's adjuvant at the base of the tail. A booster injection of 50 μg of type II collagen in incomplete Freund's adjuvant was given on day 21 after the first immunization [1]
2. Grouping and drug administration: Mice were divided into four groups (n=6 per group): normal control group (no immunization, no drug), CIA model group (immunized, oral administration of 0.5% carboxymethyl cellulose sodium (CMC-Na)), Maslinic acid low-dose group (immunized, oral administration of 20 mg/kg Maslinic acid dissolved in 0.5% CMC-Na), and Maslinic acid high-dose group (immunized, oral administration of 40 mg/kg Maslinic acid dissolved in 0.5% CMC-Na). Drug administration started on day 22 after the first immunization and continued once daily for 21 days [1]
3. Sample collection and analysis: During the experiment, paw volume was measured using a plethysmometer, and arthritis score was evaluated every 3 days. On day 43 (end of treatment), mice were anesthetized, and blood was collected from the heart to separate serum for cytokine detection. Hind paw joints were removed, fixed in 4% paraformaldehyde, decalcified, embedded in paraffin, sectioned, and stained with hematoxylin and eosin (H&E) for histological analysis. Joint tissues were also used for Western blot to detect NF-κB pathway-related proteins [1]

1. In vivo toxicity: In CIA mice treated with maslinic acid (20 mg/kg and 40 mg/kg, orally, for 21 days), no significant changes in body weight were observed compared with the normal control group. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), and creatinine (Cr) levels were within the normal range, indicating that no significant hepatotoxicity or nephrotoxicity was observed [1]. 2. In vitro toxicity: MTT assay showed that maslinic acid at concentrations up to 100 μM had no significant cytotoxicity to RAW264.7 cells, and cell viability remained above 90% [1].

[1]. Anti-inflammatory and anti-arthritic effects of pentacyclic triterpenoids maslinic acid through NF-κB inactivation. Mol Nutr Food Res. 2016 Feb;60(2):399-409.

Maslinic acid is a pentacyclic triterpenoid compound with the structure oleanolic-12-ene, substituted with hydroxyl groups at positions 2 and 3 and a carboxyl group at position 28 (2α,3β stereoisomer). It was isolated from olive (Olea europaea) and Canary Island sage (Salvia canariensis) and possesses anti-inflammatory, antioxidant, and antitumor activities. It is a plant metabolite with multiple functions including antioxidant, antitumor, and anti-inflammatory effects. It is a pentacyclic triterpenoid compound and also a dihydroxy monocarboxylic acid. It is derived from the hydride of oleanolic acid. It has been reported that maslinic acid is present in Salvia miltiorrhiza, Clematis chinensis, and other organisms with relevant data. See also: whole plant (part) of Centaurium erythraea.

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Mechanism of Action
Maslinic acid is a pentacyclic triterpenoid found in the waxy protective layer of olive (Olea europaea L.) leaves and fruits, and is a promising drug for the prevention of colon cancer. Researchers have previously demonstrated that maslinic acid can significantly inhibit the proliferation of colon cancer cells and activate mitochondrial apoptosis. This study aimed to investigate the apoptotic molecular mechanism of this compound. Researchers used HT29 adenocarcinoma cells. Changes in genotoxicity were analyzed by single-cell gel electrophoresis (comet assay). Cell cycle determination was performed using flow cytometry. Finally, changes in protein expression were detected by Western blotting. Statistical comparisons were performed using the Student's t-test. HT29 cells treated with maslinic acid showed significantly increased genotoxicity after 72 hours of treatment, significant cell cycle arrest in the G0/G1 phase, and a peak in sub-G0/G1 apoptosis after 96 hours… The antitumor activity of maslinic acid may exert its effect through p53-mediated apoptosis, the mechanism of which is by acting on key signaling pathway components that lead to increased p53 activity and inducing other factors involved in the apoptosis pathway. Researchers discovered that maslinic acid activates the expression of c-Jun N-terminal kinase (JNK) in HT29 cells, thereby inducing p53 expression. Treatment of tumor cells with maslinic acid increased the expression of Bid and Bax, suppressed Bcl-2 expression, released cytochrome c, and increased the expression of caspase-9, -3, and -7. Furthermore, maslinic acid induced delayed activation of caspase-8, thereby enhancing initial mitochondrial apoptosis signaling. All these results indicate that maslinic acid induces apoptosis in human HT29 colon cancer cells by activating p53 via the JNK-Bid-mediated mitochondrial apoptosis pathway…
Maslinic acid (2-α,3-β-dihydroxyoleanolic-12-en-28-ic acid) is a natural triterpenoid compound derived from olive. This compound inhibits oxidative stress and the production of pro-inflammatory cytokines in vitro. This study investigated the anti-inflammatory effects of maslinic acid in the central nervous system using lipopolysaccharide (LPS)-stimulated rat astrocyte cultures. Western blot analysis and quantitative real-time PCR were employed to evaluate different proteins involved in the nuclear factor-κB (NF-κB) signaling pathway. The results showed that maslinic acid exerted a significant anti-inflammatory effect by inhibiting the production of nitric oxide and tumor necrosis factor-α (TNF-α). Western blot analysis revealed that maslinic acid attenuated, in a concentration-dependent manner, LPS-induced translocation of the NF-κB p65 subunit to the nucleus and inhibited LPS-induced phosphorylation of IκBα. Furthermore, maslinic acid significantly inhibited the expression of cyclooxygenase 2 (COX-2) and inducible nitric oxide synthase (iNOS) at both the protein and mRNA levels. These results suggest that maslinic acid may alleviate neuroinflammation by inhibiting the NF-κB signaling pathway in cultured cortical astrocytes.
The receptor activator NF-κB ligand (RANKL) activates the NF-κB and MAPK/AP-1 (AP-1) signaling pathways, which are crucial for osteoclast activity. Targeting the NF-κB and MAPK/AP-1 signaling pathways to regulate osteoclast activity has been a promising strategy for treating osteoclast-related diseases. This study investigated the effects of maslinic acid (MA), a pentacyclic triterpenoid acid widely found in edible plants, on RANKL-induced osteoclastogenesis, osteoclast function, and signaling pathways using in vitro and in vivo experimental systems. In mouse bone marrow mononuclear cells (BMM) and RAW264.7 cells, MA inhibited RANKL-induced osteoclastogenesis in a dose-dependent manner within a non-growth-inhibiting concentration range and reduced the expression of osteoclastogenesis-related marker genes, including TRACP, MMP9, c-Src, CTR, and cathepsin K. Specifically, MA inhibited osteoclastogenesis and actin ring formation in the early stages. In ovariectomized mice, administration of maslinic acid (MA) prevented ovariectomy-induced bone loss by inhibiting osteoclast activity. At the molecular level, MA eliminated MAPK phosphorylation and AP-1 activity, inhibited IκBα phosphorylation and degradation, and suppressed NF-κB/p65 phosphorylation, nuclear translocation, and DNA binding activity by downregulating RANK expression and blocking the interaction between RANK and TRAF6. These data collectively indicate that MA inhibits RANKL-induced osteoclastogenesis through the NF-κB and MAPK/AP-1 signaling pathways, and that MA holds promise as an effective drug for treating osteoclast-related diseases such as osteoporosis.

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1. Maslinic acid is a pentacyclic triterpenoid compound, primarily isolated from olive peel extracts. Its anti-inflammatory and anti-arthritis effects are mainly achieved by inhibiting the NF-κB signaling pathway—specifically, by reducing IκBα phosphorylation levels and preventing NF-κB p65 nuclear translocation, thereby inhibiting the production of pro-inflammatory cytokines and mediators [1]. 2. Studies have shown that maslinic acid can alleviate collagen-induced arthritis in mice without significant toxicity, suggesting its potential for developing anti-inflammatory or anti-arthritis drugs [1].

C30H48O4

472.6997

472.355

4373-41-5

73659

White to off-white solid powder

1.1±0.1 g/cm3

570.0±50.0 °C at 760 mmHg

249 - 250 °C

312.6±26.6 °C

0.0±3.6 mmHg at 25°C

1.568

7.87

3

4

1

34

919

9

C[C@@]12CC[C@@H]3[C@@]([C@H]1CC=C4[C@]2(CC[C@@]5([C@H]4CC(CC5)(C)C)C(=O)O)C)(C[C@H]([C@@H](C3(C)C)O)O)C

MDZKJHQSJHYOHJ-LLICELPBSA-N

InChI=1S/C30H48O4/c1-25(2)12-14-30(24(33)34)15-13-28(6)18(19(30)16-25)8-9-22-27(5)17-20(31)23(32)26(3,4)21(27)10-11-29(22,28)7/h8,19-23,31-32H,9-17H2,1-7H3,(H,33,34)/t19-,20+,21-,22+,23-,27-,28+,29+,30-/m0/s1

(4aS,6aR,6aS,6bR,8aR,10R,11R,12aR,14bS)-10,11-dihydroxy-2,2,6a,6b,9,9,12a-heptamethyl-1,3,4,5,6,6a,7,8,8a,10,11,12,13,14b-tetradecahydropicene-4a-carboxylic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

DMSO : ~100 mg/mL (~211.55 mM)

Solubility in Formulation 1: 2.5 mg/mL (5.29 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (5.29 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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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