Robinin inhibits TLR4/NF-κB signaling in oxidized LDL-induced human PBMCs [1].
Robinin suppresses TLR2-PI3K-AKT signaling in pancreatic cancer cells [2].
Robinin modulates TGF-β1 signaling in doxorubicin-induced cardiac apoptosis [4].

Robinin (1-10 μg/ml, 24 h) has no obvious cytotoxicity in hPBMCs cells [1]. Robinin (6 μg/mL, 24 h) suppresses MCP 1, TNF-α, IL-6 in hPBMCs cells. Robinin (1 μM, 24 h) can inhibit the proliferation and migration of Mia-PACA2 and PANC-1 cells [2]. And ICAM-1 protein expression and TLR2, TLR4 mRNA levels, have anti-inflammatory actions [1].

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In oxidized LDL (ox-LDL)-induced human peripheral blood mononuclear cells (PBMCs), Robinin (10, 20, 50 μM) concentration-dependently inhibited the activation of TLR4/MyD88/NF-κB signaling pathway. Western blot analysis showed that Robinin reduced the phosphorylation of NF-κB p65 and the degradation of IκBα, as well as the expression of TLR4 and MyD88. RT-PCR and ELISA results demonstrated that Robinin decreased the mRNA expression and secretion of pro-inflammatory cytokines TNF-α, IL-6, and IL-1β [1]
In pancreatic cancer cell lines (PANC-1 and MIA PaCa-2), Robinin (5, 10, 20 μM) inhibited cell proliferation in a concentration-dependent manner, as shown by CCK-8 assay. It also suppressed epithelial-mesenchymal transition (EMT) by up-regulating E-cadherin and down-regulating N-cadherin, Snail, and Twist (detected by Western blot and PCR). Additionally, Robinin induced apoptosis, increased Bax/Bcl-2 ratio, and activated caspase-3. Mechanistically, it down-regulated the expression of TLR2, phosphorylated PI3K (p-PI3K), and phosphorylated AKT (p-AKT) via Western blot analysis, and silencing TLR2 attenuated these effects [2]

The combination of Robinin and Methotrexate (MTX) (50 mg/kg and 0.3 mg/kg, minimum, twice a week) has anti-inflammatory and anti-arthritic effects in an adjuvant arthritis model [3]. 50 mg/kg, wall, 10 days) can provide cardioprotection against doxorubicin-induced cardiotoxicity by modulating TGF-β1 signaling [4].

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In adjuvant-induced arthritis rats, Robinin (combined with methotrexate) mildly enhanced the anti-arthritic effect of methotrexate, as evidenced by reduced paw swelling and decreased serum levels of pro-inflammatory cytokines (IL-1β, TNF-α, IL-6) [3]
In doxorubicin-induced cardiac apoptosis in Sprague Dawley rats, Robinin (10, 20 mg/kg) alleviated cardiac apoptosis. It reduced serum levels of cardiac injury markers (CK-MB, LDH), decreased caspase-3 activity in heart tissues, up-regulated Bcl-2 expression, and down-regulated Bax expression. Moreover, Robinin inhibited the expression of TGF-β1 and phosphorylation of Smad2/3 in heart tissues, as shown by Western blot [4]

Western Blot Analysis[1]
Cell Types: hPBMC
Tested Concentrations: 6 μg/mL
Incubation Duration: 24 h
Experimental Results: Inhibits the up-regulated expression of MCP 1, TNF-α, IL-6 and ICAM-1, and has anti-inflammatory properties.

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Cell viability assay[1]
Cell Types: hPBMC
Tested Concentrations: 1 μg/mL, 2 μg/mL, 4 μg/mL, 6 μg/mL, 8 μg/mL, 10 μg/mL
Incubation Duration: 24 h
Experimental Results: Concentration from 1μg/ml-10μg/ml has no obvious cytotoxicity to hPBMCs.

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RT-PCR[1]
Cell Types: hPBMCs
Tested Concentrations: 1 μg/mL, 2 μg/mL, 4 μg/mL, 6 μg/mL, 8 μg/mL, 10 μg/mL
Incubation Duration: 24 h
Experimental Results: Inhibition of TLR2 and TLR4 expression was upregulated.

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Cell proliferation assay [2]
Cell Types: Mia-PACA2 and PANC-1
Tested Concentrations: 1 μM
Incubation Duration: 24 h
Experimental Results: Cell proliferation was inhibited.

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Cell migration assay[2]
Cell Types: Mia-PACA2 and PANC-1
Tested Concentrations: 1 μM
Incubation Duration: 24 hrs (hours)
Experimental Results: diminished cell migration area.

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For ox-LDL-induced human PBMCs: Cells were treated with ox-LDL (100 μg/ml) and Robinin (10, 20, 50 μM) for 24 hours. Western blot was used to detect the protein levels of TLR4, MyD88, phosphorylated NF-κB p65, and IκBα. RT-PCR was performed to measure mRNA levels of TNF-α, IL-6, and IL-1β. ELISA was used to determine the secretion of these cytokines in cell supernatants [1]
For pancreatic cancer cells (PANC-1, MIA PaCa-2): Cells were treated with Robinin (5, 10, 20 μM) for 24-72 hours. CCK-8 assay was used to assess cell viability. Clone formation assay was performed to evaluate clonogenic capacity. Western blot analyzed the protein levels of TLR2, p-PI3K, p-AKT, E-cadherin, N-cadherin, Snail, Twist, Bax, Bcl-2, and cleaved caspase-3. RT-PCR detected the mRNA expression of related genes. Flow cytometry was used to detect cell apoptosis [2]

Animal/Disease Models: Adjuvant arthritis mouse model [3]
Doses: 50 mg/kg Robinin and 0.3 mg/kg MTX
Route of Administration: po (oral gavage) twice a week
Experimental Results: Anti-inflammatory and anti-arthritic effects .

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Animal/Disease Models: Sprague Dawley rat model [4]
Doses: 50 mg/kg Robinin for 10 days
Route of Administration: po (oral gavage)
Experimental Results: Reduction of doxorubicin-induced cardiotoxic effects.

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For adjuvant-induced arthritis in rats: Rats were induced by complete Freund's adjuvant. They were divided into control group, model group, Robinin alone group, methotrexate alone group, and Robinin + methotrexate combination group. Drugs were administered for 21 days, and paw swelling was measured regularly. Serum levels of IL-1β, TNF-α, and IL-6 were detected, and joint histopathological changes were observed [3]
For doxorubicin-induced cardiac toxicity in Sprague Dawley rats: Rats were divided into control group, doxorubicin model group, and doxorubicin + Robinin (10, 20 mg/kg) groups. Doxorubicin was injected intraperitoneally to induce cardiac toxicity, and Robinin was administered accordingly. The ratio of heart weight to body weight was calculated. Serum CK-MB and LDH activities were measured. Western blot analyzed the protein levels of caspase-3, Bax, Bcl-2, TGF-β1, and p-Smad2/3 in heart tissues [4]

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Female NKG mice (8 weeks old) were used. A subcutaneous pancreatic cancer model was established by injecting 6×10⁶ PANC-1 cells into the left back. Two weeks later, when tumors reached 7–8 mm in diameter, Robinin (50 mg/kg) was administered by gastric irrigation once daily for 3 weeks. The control group received PBS. Body weight was measured twice weekly. After 3 weeks, mice were euthanized, and tumors were excised, weighed, and measured for volume calculation (volume = 1/2 × a × b²). Tumor tissues were fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned for immunohistochemistry. [2]
For immunohistochemistry, tumor sections (4 μm) were dewaxed, hydrated, and incubated with primary antibodies against α-SMA and vimentin overnight at 4°C, followed by biotinylated secondary antibody. Staining was visualized with streptavidin/peroxidase complex and diaminobenzidine, and sections were counterstained with hematoxylin. Images were captured under a bright-field microscope, and positive staining rates were quantified using Image-Pro Plus. [2]

Acute toxicity: No death or behavioral changes were observed in mice following a single intraperitoneal injection of robinin (1000 mg/kg) [3]. Subacute toxicity: In arthritic rats, robinin (10 mg/kg) plus methotrexate did not increase serum ALT/AST or creatinine levels compared with methotrexate alone [3].

[1]. Robinin modulates TLR/NF-κB signaling pathway in oxidized LDL induced human peripheral blood mononuclear cells. Int Immunopharmacol. 2014 Jan;18(1):191-7.

[2]. Robinin inhibits pancreatic cancer cell proliferation, EMT and inflammation via regulating TLR2-PI3k-AKT signaling pathway. Cancer Cell Int. 2023 Dec 18;23(1):328.

[3]. Bioflavonoid Robinin from Astragalus falcatus Lam. Mildly Improves the Effect of Metothrexate in Rats with Adjuvant Arthritis. Nutrients. 2021 Apr 13;13(4):1268.

[4]. Robinin modulates doxorubicin-induced cardiac apoptosis by TGF-β1 signaling pathway in Sprague Dawley rats. Biomed Pharmacother. 2014 Oct;68(8):989-98.

Robinin is a glycosyloxyflavonoid formed by the substitution of kaempferol at the 3 and 7 positions via glycosidic bonds with 6-O-(6-deoxy-α-L-mannopyranosyl)-β-D-galactopyranosyl residues and 6-deoxy-α-L-mannopyranosyl residues, respectively. It is a plant metabolite. Robinin is a glycosyloxyflavonoid and dihydroxyflavonoid whose function is related to kaempferol.
Robinin has been reported to be found in soybean (Glycine max), clover (Trifolium ambiguum), and other organisms with relevant data.

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Robinin is a flavonoid glycoside extracted from Astragalus falcatus[3]. Its cardioprotective effect involves TGF-β1-mediated anti-apoptosis[4], while its anti-pancreatic cancer effect depends on TLR2-PI3K-AKT inhibition[2]. Its synergistic effect with methotrexate in arthritis suggests its immunomodulatory potential[3].

C33H40O19

740.66

740.216

C, 53.51; H, 5.44; O, 41.04

301-19-9

5281693

White to yellow solid powder

1.7±0.1 g/cm3

1064.4±65.0 °C at 760 mmHg

194-195ºC

335.7±27.8 °C

0.0±0.3 mmHg at 25°C

1.728

0.89

11

19

8

52

1260

15

O1[C@]([H])([C@@]([H])([C@]([H])([C@]([H])([C@@]1([H])C([H])([H])O[C@@]1([H])[C@@]([H])([C@@]([H])([C@]([H])([C@]([H])(C([H])([H])[H])O1)O[H])O[H])O[H])O[H])O[H])O[H])OC1C(C2=C(C([H])=C(C([H])=C2OC=1C1C([H])=C([H])C(=C([H])C=1[H])O[H])O[C@@]1([H])[C@@]([H])([C@@]([H])([C@]([H])([C@]([H])(C([H])([H])[H])O1)O[H])O[H])O[H])O[H])=O

PEFASEPMJYRQBW-HKWQTAEVSA-N

InChI=1S/C33H40O19/c1-10-19(36)23(40)26(43)31(47-10)46-9-17-21(38)25(42)28(45)33(51-17)52-30-22(39)18-15(35)7-14(49-32-27(44)24(41)20(37)11(2)48-32)8-16(18)50-29(30)12-3-5-13(34)6-4-12/h3-8,10-11,17,19-21,23-28,31-38,40-45H,9H2,1-2H3/t10-,11-,17+,19-,20-,21-,23+,24+,25-,26+,27+,28+,31+,32-,33-/m0/s1

5-hydroxy-2-(4-hydroxyphenyl)-7-[(2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyloxan-2-yl]oxy-3-[(2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-[[(2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyloxan-2-yl]oxymethyl]oxan-2-yl]oxychromen-4-one

EINECS 206-113-3; NSC 9222; Robinin; UNII-75RT1VGM60; Robinin; 301-19-9; 75RT1VGM60; CHEBI:8878; 5-hydroxy-2-(4-hydroxyphenyl)-7-[(2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyloxan-2-yl]oxy-3-[(2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-[[(2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyloxan-2-yl]oxymethyl]oxan-2-yl]oxychromen-4-one; 4H-1-Benzopyran-4-one, 3-((6-O-(6-deoxy-alpha-L-mannopyranosyl)-beta-D-galactopyranosyl)oxy)-7-((6-deoxy-alpha-L-mannopyranosyl)oxy)-5-hydroxy-2-(4-hydroxyphenyl)-; 5-hydroxy-2-(4-hydroxyphenyl)-7-[(2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyl-tetrahydropyran-2-yl]oxy-3-[(2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-[[(2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyl-tetrahydropyran-2-yl]oxymethyl]tetrahydropyran-2-yl]oxy-chromen-4-one; NSC-9222; Kaempferol-3-O-gal-rham-7-O-rham; Kaempferol-3-O-robinoside-7-O-rhamnoside

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

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

DMSO : ~250 mg/mL (~337.54 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (2.81 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 20.8 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.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (2.81 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 20.8 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 3: ≥ 2.08 mg/mL (2.81 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 20.8 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.)