In mast cells, Orientin inhibits the FcεRI-mediated signaling pathway, reducing phosphorylation of Lyn, Syk, Akt, and NF-κB p65 [1].
In spinal nerve ligation (SNL) rats, Orientin inhibits the Toll-like receptor 4 (TLR4)/NF-κB signaling pathway [2].

On mBMMC and RPMC cells, the allergy receptor RBL-2H3 is inhibited by orientin (1 µM) [1].

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Orientin (0.01 - 100 μM) did not cause cytotoxicity in RBL-2H3, mouse bone marrow-derived mast cells (mBMMCs), and rat peritoneal mast cells (RPMCs) up to 100 μM [1].
Orientin (0.01 - 1 μM) significantly suppressed histamine levels in a concentration-dependent manner in RBL-2H3, mBMMCs, and RPMCs compared to DNP-HSA-stimulated cells [1].
Orientin (0.01 - 1 μM) significantly suppressed β-hexosaminidase levels in a concentration-dependent manner in RBL-2H3, mBMMCs, and RPMCs compared to DNP-HSA-stimulated cells [1].
Orientin (0.01 - 1 μM) inhibited the increased intracellular calcium levels in a concentration-dependent manner in DNP-HSA-stimulated RBL-2H3, mBMMCs, and RPMCs [1].
Orientin (0.01 - 1 μM) markedly inhibited the gene expression (mRNA) of TNF-α and IL-4 in DNP-HSA-stimulated RBL-2H3 cells in a concentration-dependent manner [1].
Orientin (0.01 - 1 μM) markedly inhibited the secretion of pro-inflammatory cytokines (TNF-α and IL-4) in DNP-HSA-stimulated RBL-2H3 cells in a concentration-dependent manner [1].
In RBL-2H3 cells, Orientin inhibited the phosphorylation of FcεRI-mediated signaling proteins: Lyn, Syk, Akt, and NF-κB p65 compared to DNP-HSA-stimulated cells [1].
In an SNL rat model, Orientin (10, 20, 40 mg/kg) suppressed the levels of pro-inflammatory cytokines IL-1β, TNF-α, and IL-6 and increased the level of anti-inflammatory cytokine IL-10 in the spinal cord [2].
In SNL rats, Orientin (10, 20, 40 mg/kg) down-regulated malondialdehyde (MDA) levels and up-regulated superoxide dismutase (SOD) and glutathione (GSH) levels in the spinal cord [2].
In SNL rats, Orientin (10, 20, 40 mg/kg) inhibited the activation of microglia and astrocytes in the spinal dorsal horn, as shown by reduced OX42 and GFAP immunofluorescence [2].
In SNL rats, Orientin (10, 20, 40 mg/kg) inhibited the TLR4/NF-κB signaling pathway in the spinal cord in a dose-dependent manner [2].

In a PCA mouse model, one intravenous dosage of orientin (0.1–10 mg/kg) decreases allergy responses [1]. Orientin injections, 10–40 mg/kg, intraperitoneally once daily for 12 days) Within SNL

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Oral administration of Orientin (0.1 - 10 mg/kg) suppressed IgE-mediated passive cutaneous anaphylaxis (PCA) reactions in ICR mice in a dose-dependent manner, evidenced by reduced Evan's blue pigmentation and ear swelling [1].
In a rat spinal nerve ligation (SNL) model, intraperitoneal injection of Orientin (10, 20, 40 mg/kg) for 12 days alleviated mechanical and thermal allodynia, as shown by increased paw mechanical withdrawal threshold (PWT) and paw thermal withdrawal latency (PWL) [2].
In SNL rats, Orientin (10, 20, 40 mg/kg) suppressed pro-inflammatory cytokines (IL-1β, TNF-α, IL-6), increased anti-inflammatory cytokine IL-10, reduced oxidative stress (MDA), increased antioxidant enzymes (SOD, GSH), and inhibited microglial and astrocyte activation in the spinal cord [2].
In SNL rats, the neuroprotective effect of Orientin (10, 20, 40 mg/kg) was mediated by inhibition of the TLR4/NF-κB signaling pathway [2].

Cell viability assay [1]
Cell Types: RBL-2H3, mBMMCs, and RPMCs
Tested Concentrations: 0.01–100 µM
Incubation Duration: 8 h
Experimental Results: Significant inhibition of histamine and β-hexosamine in RBL-2H3, mBMMCs, and RPMC cells enzyme levels. Inhibits intracellular calcium levels in RBL-2H3, mBMMC and RPMC cells. Dramatically inhibits the expression and secretion of pro-inflammatory cytokines in RBL-2H3 cells. Inhibits phosphorylation in RBL-2H3 cells.

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Cell viability was measured using an MTT assay. Cells (RBL-2H3, mBMMCs, RPMCs) were treated with Orientin (0.01 - 100 μM) for 8 hours, then incubated with MTT reagent for 2 hours. The formed formazan crystals were dissolved in dimethylsulfoxide, and absorbance was measured at 570 nm [1].
For mast cell degranulation, cells (RBL-2H3, mBMMCs, RPMCs) were sensitized with anti-DNP IgE (50 ng/mL), pre-treated with or without Orientin (0.01 - 1 μM) or dexamethasone for 1 hour, and then stimulated with DNP-HSA (100 ng/mL). Histamine levels in the media were measured using o-phthaladialdehyde (OPA) with fluorescence at 360 nm excitation and 440 nm emission. β-hexosaminidase levels were measured using a substrate buffer incubated for 1 hour at 37°C, and absorbance was read at 405 nm [1].
For intracellular calcium measurement, sensitized cells were pre-incubated with Fluo-3/AM for 1 hour, pre-treated with or without Orientin for 1 hour, and then stimulated with DNP-HSA. Fluorescence intensity was measured at 485 nm excitation and 520 nm emission [1].
For gene expression analysis, RNA was purified from RBL-2H3 cells. qPCR was used to measure mRNA levels of TNF-α and IL-4. Primer sequences were provided for β-actin, TNF-α, and IL-4 [1].
For cytokine secretion, an ELISA kit was used following the manufacturer's instructions. Absorbance was measured at 450 nm [1].
For Western blot, RBL-2H3 cells were sensitized, pre-treated with or without Orientin for 1 hour, and then stimulated with DNP-HSA for 5 minutes (Lyn and Syk), 30 minutes (Akt), or 1 hour (IκBα and NF-κB p65). Proteins were extracted, electrophoresed on 10% SDS-PAGE, transferred to nitrocellulose membranes, and incubated with specific primary antibodies followed by HRP-conjugated secondary antibodies. Immunodetection used an enhanced chemiluminescence kit [1].
For immune fluorescent staining in SNL rats, spinal cord tissues were fixed, sectioned (4 μm), blocked, and incubated with GFAP primary antibody (pre-conjugated with Alexa Fluor 488) or OX42 anti-rabbit antibody overnight at 4°C. After washing, OX42 sections were incubated with donkey IgG Alexa Fluor 488 secondary antibody for 20 minutes at 37°C. Fluorescent images were captured and immuno-intensity processed [2].
For biochemical parameter investigation in SNL rats, L5 spinal cord tissues were collected and assayed for malondialdehyde (MDA), glutathione (GSH), and superoxide dismutase (SOD) levels using lipid peroxidation assay kits. Absorbance was read at 535 nm (MDA), 412 nm (GSH), and 560 nm (SOD) [2].
For cytokine measurement in SNL rats, ELISA kits were used to probe TNF-α, IL-6, IL-1β, and IL-10 levels in spinal cord homogenates [2].
For Western blot in SNL rats, proteins were extracted from the right part of the spinal cord, quantified by BCA kit, separated by SDS-PAGE, transferred to nitrocellulose membranes, and probed with primary antibodies followed by secondary antibodies. Histone-H3 was used as a nuclear protein control and β-actin as a total protein control [2].

Animal/Disease Models: SNL rat model [2]
Doses: 10, 20, 40 mg/kg
Route of Administration: intraperitoneal (ip) injection; one time/day for 12 days
Experimental Results: Alleviates the down-regulation of PWL and promotes behavior in the SNL rat model recover. Reduces levels of IL-6, IL-1β and TNF-α. Increases levels of anti-inflammatory IL-10. Reduce MDA levels.

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For the passive cutaneous anaphylaxis (PCA) model, male ICR mice (30-35 g, 6 weeks old) were sensitized intradermally in the ear with 0.5 μg/site of anti-DNP IgE or PBS. After 48 hours, mice received Orientin (0.1, 1, 10 mg/kg) or dexamethasone (10 mg/kg) per orally (p.o.). One hour later, mice were challenged intravenously (i.v.) with a mixture of DNP-HSA (1 mg/mouse) and 4% Evans blue (1:1). Mice were euthanized 30 minutes after i.v. injection. Ear thickness and Evans blue dye extravasation (absorbance) were measured [1].
For the spinal nerve ligation (SNL) model, male Sprague-Dawley (SD) rats (200-220 g) underwent surgery to ligate the right L5 spinal nerve with a 6-0 silk suture. The sham operation group underwent the same procedure without ligation. After SNL, Orientin was dissolved in 5% DMSO/saline and administered intraperitoneally (i.p.) once daily for 12 days at doses of 10 mg/kg (low), 20 mg/kg (medium), or 40 mg/kg (high). The model group received an equivalent volume of 5% DMSO/saline. The positive drug group received pregabalin (10 mg/kg i.v.) daily. Penicillin (0.2 million units i.v.) was given for 3 days post-surgery to prevent infection [2].
For behavioral assays in SNL rats, paw mechanical withdrawal threshold (PWT) was measured using calibrated nylon Von Frey filaments (0.4-26 g) applied to the ipsilateral hind paw. Paw thermal withdrawal latency (PWL) was measured using a thermal radiation instrument on a glass plate. Both tests were performed every 3 days from the start of Orientin administration to the end [2].

Orientin did not cause any cytotoxicity in RBL-2H3, mBMMCs, or RPMCs up to 100 μM [1].
No toxic signs such as death, vomiting, lacrimation, or diminished activity were observed in rats during 12 days of intraperitoneal administration of Orientin at doses up to 40 mg/kg [2].

[1]. Inhibitory effects of orientin in mast cell-mediated allergic inflammation. Pharmacol Rep. 2020 Jan 24.

[2]. Orientin and neuropathic pain in rats with spinal nerve ligation. 61Int Immunopharmacol. 2018 May;58:72-79.

Orientin is a C-glycoside compound, a product of luteolin with β-D-glucopyranoside substituted at the 8-position. It possesses antioxidant and metabolic activities. Orientin is a C-glycoside compound, a tetrahydroxyflavone, and a 3'-hydroxyflavone compound whose functions are related to luteolin. Orientin has been reported in Cecropia hololeuca, Gentiana algida, and other organisms with relevant data. See also: fenugreek seeds (partial); aerial parts of Cannabis sativa subsp. indica; acai berry pulp (partial).

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Orientin suppresses allergic inflammation by inhibiting FcεRI-mediated mast cell activation, reducing degranulation (histamine and β-hexosaminidase), pro-inflammatory cytokines (TNF-α, IL-4), and intracellular calcium levels [1].
Orientin exerts neuroprotective effects in a spinal nerve ligation model of neuropathic pain by reducing pro-inflammatory cytokines (IL-1β, TNF-α, IL-6), oxidative stress (MDA), and activating antioxidant enzymes (SOD, GSH), as well as inhibiting microglial and astrocyte activation via the TLR4/NF-κB pathway [2].
The anti-allergic effect of Orientin was demonstrated in RBL-2H3 cells, mouse bone marrow-derived mast cells, rat peritoneal mast cells, and an IgE-mediated PCA mouse model [1].
The neuroprotective effect of Orientin was demonstrated in a rat spinal nerve ligation model using behavioral assays (PWT, PWL), biochemical assays, immunofluorescence, and Western blot [2].

C21H20O11

448.3769

448.1

C, 56.25; H, 4.50; O, 39.25

28608-75-5

5281675

Light yellow to yellow solid powder

1.8±0.1 g/cm3

816.1±65.0 °C at 760 mmHg

260-285ºC

289.1±27.8 °C

0.0±3.1 mmHg at 25°C

1.767

1.58

8

11

3

32

729

5

O1[C@]([H])(C([H])([H])O[H])[C@]([H])([C@@]([H])([C@]([H])([C@]1([H])C1=C(C([H])=C(C2C(C([H])=C(C3C([H])=C([H])C(=C(C=3[H])O[H])O[H])OC1=2)=O)O[H])O[H])O[H])O[H])O[H]

PLAPMLGJVGLZOV-VPRICQMDSA-N

InChI=1S/C21H20O11/c22-6-14-17(28)18(29)19(30)21(32-14)16-11(26)4-10(25)15-12(27)5-13(31-20(15)16)7-1-2-8(23)9(24)3-7/h1-5,14,17-19,21-26,28-30H,6H2/t14-,17-,18+,19-,21+/m1/s1

2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-8-[(2S,3R,4R,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]chromen-4-one

Lutexin; Orientin

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 : ~31.25 mg/mL (~69.70 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (4.64 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 (4.64 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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 (Please use freshly prepared in vivo formulations for optimal results.)