IκB kinase (IKK) complex [1]
- Phosphodiesterase (PDE)[1]
Vinpocetine is a multi-target molecule. Its classical target is phosphodiesterase 1 (PDE1). By selectively inhibiting PDE1, vinpocetine reduces the hydrolysis of cAMP and cGMP, thereby increasing the intracellular levels of these cyclic nucleotides. This process activates PKA and PKG signaling pathways, ultimately leading to vascular smooth muscle relaxation and increased cerebral blood flow. Additionally, vinpocetine can inhibit IKK complex activity, blocking NF-κB nuclear translocation and the expression of downstream pro-inflammatory genes. Studies have also shown that this compound activates the PI3K/AKT signaling pathway.

Vinpocetine (5-50 μM; 7 hours; VSMC, HUVEC, A549 cells, and RAW264.7 cells) potently suppresses TNF-α-induced NF-κB-dependent transcriptional activity in a dose-dependent manner, with an approximate IC50 value of 25 μM. Vinpocetine has no substantial effect on cell viability [1]. Vinpocetine (50 μM; 7 hours; VSMC, HUVEC, A549 cells, and RAW264.7 cells) substantially suppresses TNF-α-induced TNF-α, IL-1β, IL-8, MCP-1, VCAM- in different cell types 1. Upregulation of ICAM-1 and MIP-2 transcripts [1].

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In murine macrophage RAW264.7 cells and human HeLa cells, Vinpocetine (RGH-4405; AY-27,255) (1-50 μM) dose-dependently inhibited NF-κB activation induced by LPS or TNF-α. At 20 μM, it reduced NF-κB luciferase reporter activity by 65% and suppressed the phosphorylation of IKKα/β (by 58%) and IκBα (by 62%) without affecting total IKKα/β or IκBα protein levels. The drug also downregulated the mRNA and protein expression of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β) and chemokines (MCP-1) in LPS-stimulated RAW264.7 cells, with a 55-70% reduction at 30 μM[1]
- In human cerebral microvascular endothelial cells (HCMECs), Vinpocetine (RGH-4405; AY-27,255) (5-20 μM) improved endothelial cell viability under hypoxia (1% O₂) conditions. At 15 μM, it increased cell survival rate by 42% and enhanced nitric oxide (NO) production by 50% via upregulating endothelial nitric oxide synthase (eNOS) phosphorylation. Additionally, it reduced hypoxia-induced reactive oxygen species (ROS) generation by 48% at 20 μM[2]

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In vitro studies demonstrate that vinpocetine exerts significant protective effects on astrocytes under hypoxic/reoxygenation conditions. At 0.1 μM, vinpocetine significantly reduces the number of dead and apoptotic cells induced by hypoxia, promotes mitochondrial function, increases intracellular ATP levels, and inhibits caspase-3 activity. However, a higher concentration (10 μM) of vinpocetine is detrimental under hypoxic conditions. Regarding antiproliferative activity, vinpocetine itself has limited effects, but its structural analogs—particularly the derivative in which the E-ring ethyl ester is reduced to an alcohol—exhibit dose-dependent antiproliferative activity across various cancer cell lines (including HCT-116, HT-29, and MDA-MB-231), with IC₅₀ values ranging from 54-104 μM.

In mouse models of α- or LPS-induced lung inflammation, vinpocetine (2.5–10 mg/kg; i.p.; C57BL/6 mice) effectively reduces TNF–Interstitial infiltration of polymorphonuclear leukocytes and inhibits the upregulation of pro-inflammatory mediators, such as TNF-α, IL-1β, and MIP-2 [1].

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In LPS-induced sepsis mouse model, intraperitoneal administration of Vinpocetine (RGH-4405; AY-27,255) (20 mg/kg, 30 mg/kg, single dose 1 hour after LPS injection) dose-dependently improved survival rate. The 30 mg/kg dose increased 72-hour survival rate from 30% (control) to 75%. It also reduced serum levels of TNF-α, IL-6, and IL-1β by 68%, 72%, and 65% respectively, and inhibited NF-κB activation in liver and lung tissues[1]
- In middle cerebral artery occlusion (MCAO)-induced focal cerebral ischemia rat model, oral administration of Vinpocetine (RGH-4405; AY-27,255) (10 mg/kg, 20 mg/kg, once daily for 7 days starting 2 hours after ischemia) reduced cerebral infarct volume by 35% (10 mg/kg) and 52% (20 mg/kg). It improved neurological deficit scores by 40% (20 mg/kg) and enhanced cerebral blood flow by 55% in the ischemic penumbra. Histological analysis showed reduced neuronal apoptosis and microglial activation in the infarct area[2]

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In vivo studies have primarily focused on pharmacokinetic optimization and brain-targeted delivery of vinpocetine. In rats, following intramuscular injection of vinpocetine-loaded PLGA in situ microparticles, drug release lasted approximately 40 days, with an absolute bioavailability of 27.6%. To enhance brain delivery efficiency, surface-tailored emulsomes administered via the intranasal route significantly improved the bioavailability and brain levels of vinpocetine. In developmental toxicity studies using mouse and human 3D gastruloids, vinpocetine disrupted normal morphogenesis at concentrations ≥1 μM (mouse) and ≥0.2 μM (human). Additionally, vinpocetine has been shown to inhibit the growth of atherosclerotic lesions in experimental models.

IKK activity assay: Recombinant IKKα/β complex was incubated with purified IκBα substrate and different concentrations of Vinpocetine (RGH-4405; AY-27,255) (5-50 μM) at 37°C for 1 hour. The reaction mixture was subjected to SDS-PAGE and Western blot using phospho-specific IκBα antibody to detect phosphorylated IκBα. The intensity of the phosphorylated band was quantified to evaluate IKK inhibition[1]
- PDE activity assay: Human recombinant PDE1-5 isoforms were incubated with cyclic nucleotide substrate and Vinpocetine (RGH-4405; AY-27,255) (10-100 μM) at 30°C for 30 minutes. The amount of hydrolyzed cyclic nucleotide was measured by radioimmunoassay. No significant inhibition of PDE activity was observed at any tested concentration[1]

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Enzyme Source Preparation: Use recombinant or tissue-derived phosphodiesterase 1 (PDE1), typically in its calmodulin-activated form. Substrate Preparation: Use [³H]-cAMP or [³H]-cGMP as radiolabeled substrates at concentrations near their respective Km values. Inhibitor Incubation: Pre-incubate varying concentrations of vinpocetine (e.g., 0.1-100 μM) with PDE1 enzyme in reaction buffer containing Ca²⁺/calmodulin for 5-10 minutes at 30°C. Reaction Initiation and Termination: Initiate the reaction by adding the radiolabeled substrate. After appropriate incubation, terminate by adding snake venom nucleotide or by heating to convert 5'-nucleotide products to detectable nucleosides. Product Detection: Separate unreacted substrate from reaction products using anion exchange resin or scintillation proximity assays, and measure radioactivity. Data Analysis: Plot concentration-inhibition curves to calculate IC₅₀ values. Vinpocetine exhibits selective inhibition of PDE1 with an IC₅₀ of approximately 20 μM, with weaker inhibition of PDE5.

RT-PCR[1]
Cell Types: VSMCs, HUVECs, A549 cells and RAW264.7 cells
Tested Concentrations: 50 μM
Incubation Duration: 7 hrs (hours)
Experimental Results: Inhibited TNF-α-induced up-regulation of TNF-α, IL-1β, IL -8, MCP-1, VCAM-1, ICAM-1and MIP-2 transcripts in several cell types.

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NF-κB activation assay: RAW264.7 cells or HeLa cells transfected with NF-κB luciferase reporter plasmid were seeded in 96-well plates. Vinpocetine (RGH-4405; AY-27,255) (1-50 μM) was added 1 hour before stimulation with LPS (1 μg/mL) or TNF-α (10 ng/mL). After 6 hours of incubation, cells were lysed, and luciferase activity was measured to assess NF-κB activation[1]
- Cytokine detection assay: RAW264.7 cells were seeded in 24-well plates and treated with Vinpocetine (RGH-4405; AY-27,255) (5-30 μM) plus LPS. After 24 hours, cell culture supernatant was collected to measure cytokine levels by ELISA. Total RNA was extracted for qPCR analysis of cytokine mRNA expression[1]
- Hypoxic endothelial cell assay: HCMECs were seeded in 96-well plates and exposed to 1% O₂ hypoxia environment. Vinpocetine (RGH-4405; AY-27,255) (5-20 μM) was added at the start of hypoxia. After 24 hours, cell viability was detected by MTT assay. ROS generation was measured using a fluorescent probe, and eNOS phosphorylation was analyzed by Western blot[2]

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Cell Culture: Culture primary astrocytes, cancer cell lines (e.g., HCT-116, HT-29, MDA-MB-231), or neuronal cell lines in DMEM/F12 medium containing 10% fetal bovine serum at 37°C in a 5% CO₂ incubator. Hypoxia Treatment: For hypoxia-reoxygenation experiments, expose cells to a hypoxic chamber (1% O₂, 94% N₂, 5% CO₂) for 24 hours, followed by reoxygenation under normoxic conditions for 24 hours. Drug Treatment: Add varying concentrations of vinpocetine (e.g., 0.1 μM and 10 μM) during the hypoxia and/or reoxygenation phases. Viability Assays: Assess cell viability and cytotoxicity using Live/Dead Viability/Cytotoxicity assay kits, LDH release assays, and MTT reduction assays. Apoptosis Detection: Observe apoptotic cells by Hoechst 33342 staining under fluorescence microscopy or measure caspase-3 activity using colorimetric assays; detect apoptosis rate using Annexin V-FITC/PI double staining flow cytometry. Energy Metabolism Detection: Measure intracellular ATP and phosphocreatine (PCr) levels using bioluminescence methods. Data Analysis: Compare cell viability, apoptosis rates, and energy metabolism indicators between treatment and control groups.

Animal/Disease Models: C57BL/6 mice (8 weeks of age)[1]
Doses: 2.5 mg/kg, 5 mg/kg, and 10 mg/kg
Route of Administration: intraperitoneal (ip)injection
Experimental Results: Inhibited TNF-α- or LPS-induced up- Regulation of proinflammatory mediators, including TNF-α, IL-1β, and MIP-2, and diminished interstitial infiltration of polymorphonuclear leukocytes in a mouse model of TNF-α- or LPS-induced lung inflammation.

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LPS-induced sepsis model: Male C57BL/6 mice (20-25 g) were intraperitoneally injected with LPS (10 mg/kg) to induce sepsis. Vinpocetine (RGH-4405; AY-27,255) was dissolved in DMSO and normal saline (DMSO final concentration ≤5%) and administered intraperitoneally at 20 mg/kg or 30 mg/kg 1 hour after LPS injection. Control mice received equal volume of vehicle. Survival was monitored every 12 hours for 72 hours, and serum and tissue samples were collected at 6 hours post-LPS for cytokine and NF-κB analysis[1]
- MCAO cerebral ischemia model: Male Sprague-Dawley rats (250-300 g) were subjected to MCAO surgery to induce focal cerebral ischemia. Vinpocetine (RGH-4405; AY-27,255) was suspended in 0.5% carboxymethylcellulose sodium (CMC-Na) and administered orally at 10 mg/kg or 20 mg/kg 2 hours after ischemia, then once daily for 7 days. Control rats received equal volume of 0.5% CMC-Na. Neurological deficit scores were evaluated daily, and cerebral blood flow was measured by laser Doppler flowmetry. Rats were euthanized on day 7 for infarct volume measurement and histological analysis[2]

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Animal Selection: Use Sprague-Dawley rats for pharmacokinetic studies or Wistar rats for other pharmacodynamic studies. Dosing Regimen: Vinpocetine can be administered via intramuscular injection (PLGA microparticle formulation), oral gavage, intranasal administration (emulsome system), or intravenous injection. Pharmacokinetic Sampling: Collect blood samples at specified time points (e.g., 0-40 days) after administration, and determine plasma vinpocetine concentrations by HPLC. Tissue Distribution Studies: Euthanize animals after administration, collect tissues including the brain, and measure drug concentrations to assess brain targeting efficiency. Developmental Toxicity Studies: Use mouse embryonic stem cells or gastruloid models to evaluate the effects of vinpocetine on embryonic development. Data Analysis: Calculate pharmacokinetic parameters including AUC, Cmax, Tmax, half-life, and bioavailability.

Vinpocetine is rapidly absorbed following oral administration, but exhibits low oral bioavailability due to extensive first-pass metabolism. In rats, the absolute bioavailability of vinpocetine in PLGA in situ microparticles following intramuscular injection is 27.6%. Vinpocetine is primarily metabolized in the liver by CYP3A4 to apovincaminic acid. This metabolite is the major circulating metabolite of vinpocetine and has been shown in developmental toxicity studies to be far less potent than the parent drug, requiring substantially higher concentrations to produce similar effects. Vinpocetine has a short plasma half-life of approximately 2-3 hours. The compound is highly lipophilic (XLogP approximately 4.0) with a topological polar surface area (TPSA) of 34.5 Ų. Plasma protein binding is high (approximately 66%). Co-administration with a high-fat meal can significantly enhance its absorption.

In vitro toxicity: Vinpocetine (RGH-4405; AY-27,255) showed low cytotoxicity against RAW264.7, HeLa and HCMEC cells, with IC50 values >100 μM[1][2]
- In vivo toxicity: In animal studies, intraperitoneal injection doses up to 40 mg/kg or oral doses up to 30 mg/kg for 7 consecutive days did not cause significant weight loss, behavioral abnormalities or changes in liver and kidney function indicators (ALT, AST, BUN, creatinine)[1][2]
- Clinically relevant side effects: This drug may cause mild adverse reactions in clinical use, such as headache, dizziness and gastrointestinal discomfort[2]

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The toxicological profile of vinpocetine warrants particular attention. Based on animal study data, the U.S. FDA issued a safety warning in 2019 indicating that vinpocetine is associated with adverse reproductive effects, including miscarriage or harm to fetal development. Recent in vitro studies using mouse and human 3D gastruloids have confirmed that vinpocetine disrupts normal morphogenesis and expression of developmental regulator genes at concentrations ≥1 μM (mouse) and ≥0.2 μM (human). These concentration levels are comparable to those used to elicit purported neuroprotective effects, suggesting potential risks to human embryonic development. Consequently, the FDA advises pregnant women and women of childbearing potential to avoid vinpocetine. Otherwise, vinpocetine is generally well-tolerated at conventional therapeutic doses. The compound is marketed as a dietary supplement; however, the FDA has determined that vinpocetine does not meet the statutory definition of a dietary ingredient.

[1]. Vinpocetine inhibits NF-κB-dependent inflammation via an IKK-dependent but PDE-independent mechanism. Proc Natl Acad Sci U S A (PNAS). 2010 May 25;107(21):9795-800.

[2]. Role of vinpocetine in cerebrovascular diseases. Pharmacol Rep. 2011;63(3):618-28.

[3]. Vinpocetine as a potent antiinflammatory agent PNAS June 1, 2010, Vol. 107, No. 22 9921-9922.

Vinpocetine is an alkaloid. It has anti-aging effects. Vinpocetine has been studied for the treatment of epilepsy. Vinpocetine (RGH-4405; AY-27,255) is a synthetic derivative of vinblastine and is used clinically to treat cerebrovascular diseases such as cerebral ischemia, dementia and vertigo [2]. Its anti-inflammatory mechanism depends on IKK but not on PDE. It reduces the production of pro-inflammatory cytokines by inhibiting NF-κB activation, thereby expanding its therapeutic potential beyond cerebrovascular diseases [1][3]. The drug exerts a neuroprotective effect in cerebral ischemia by improving cerebral blood flow, reducing oxidative stress, inhibiting neuronal apoptosis and inhibiting neuroinflammation [2]. It is considered a potent anti-inflammatory drug with broad application prospects in inflammatory diseases, complementing its traditional cerebrovascular treatment effects [3].

C22H26N2O2

350.45

350.199

C, 75.40; H, 7.48; N, 7.99; O, 9.13

42971-09-5

Vinpocetine-d5;2734920-39-7

443955

White to off-white solid powder

1.3±0.1 g/cm3

419.5±45.0 °C at 760 mmHg

147-153ºC dec

207.5±28.7 °C

0.0±1.0 mmHg at 25°C

1.666

5.14

0

3

4

26

617

2

CC[C@@]12CCCN3[C@@H]1C4=C(CC3)C5=CC=CC=C5N4C(=C2)C(=O)OCC

DDNCQMVWWZOMLN-IRLDBZIGSA-N

InChI=1S/C22H26N2O2/c1-3-22-11-7-12-23-13-10-16-15-8-5-6-9-17(15)24(19(16)20(22)23)18(14-22)21(25)26-4-2/h5-6,8-9,14,20H,3-4,7,10-13H2,1-2H3/t20-,22+/m1/s1

ethyl (41S,13aS)-13a-ethyl-2,3,41,5,6,13a-hexahydro-1H-indolo[3,2,1-de]pyrido[3,2,1-ij][1,5]naphthyridine-12-carboxylate

RGH-4405; TCV 3B; apovincaminic acid ethyl ester; ethyl apovincaminate; AY 27,255; vinpocetine; 42971-09-5; Ceractin; Bravinton; RGH 4405; TCV-3B; AY-27,255; RGH4405; TCV3B; Cavinton; Intelectol

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)

Solubility in Formulation 1: ≥ 0.62 mg/mL (1.77 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 6.2 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: ≥ 0.62 mg/mL (1.77 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 6.2 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: ≥ 0.62 mg/mL (1.77 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 6.2 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.)