Voltage-dependent sodium channel isoforms: Nav1.2 (IC50=116±45 μmol/L), Nav1.3 (IC50=802±162 μmol/L), Nav1.4 (IC50=96±18 μmol/L), Nav1.5 (IC50=131±363 μmol/L), Nav1.6 (IC50=326±88 μmol/L), Nav1.7 (IC50=394±66 μmol/L), Nav1.8 (IC50=49±15 μmol/L) [1]
Tonic block component: Nav1.2 (IC50=1.22±0.57 mmol/L), Nav1.3 (IC50=1.44±0.15 mmol/L), Nav1.4 (IC50=2.64±1.19 mmol/L), Nav1.5 (IC50=0.61±0.08 mmol/L), Nav1.6 (IC50=1.07±0.60 mmol/L), Nav1.7 (IC50=0.78±0.13 mmol/L), Nav1.8 (IC50=0.40±0.10 mmol/L) [1]
Tolperisone HCl targets voltage-gated sodium and calcium channels. It acts as a voltage-gated sodium channel blocker with a preference for the open/inactivated state. It inhibits seven different isoforms of NaV expressed in Xenopus oocytes at micromolar concentrations. It has an IC50 of 50 μM for the peak-to-peak amplitude of monosynaptic compound action potential. By inhibiting these channels, it reduces neuronal excitability and suppresses neuromuscular transmission.

Tolperisone reduces sodium and potassium permeability in the node of Ranvier of frogs, blocks calcium currents in snail neurons and inhibits current through heterologously expressed Nav1.6 channels. Tolperisone hydrochloride represents an effective and safe treatment of painful reflex muscle spasm without the typical side effects of centrally active muscle relaxants.

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Tolperisone produced a concentration-dependent block of sodium currents (IpNa+) in Xenopus laevis oocytes expressing seven Nav isoforms. For Nav1.8, cumulative dose-response showed IC50=49±15 μmol/L for Tolperisone vs 128±7 μmol/L for lidocaine (P<0.001). For Nav1.6, 50 μmol/L, 250 μmol/L and 3 mmol/L Tolperisone progressively blocked currents. [1]
Tolperisone exerted significant tonic (use-independent) block: Gmax was significantly reduced for all isoforms except Nav1.4? (Table 3 shows tonic IC50 values; at concentrations near IC50, Tolperisone reduced Gmax more than lidocaine for Nav1.2, Nav1.3, Nav1.7, Nav1.8). [1]
Voltage-dependent activation was minimally affected: small but significant shifts in half-activation potential (Ea0.5) for Nav1.3 (from -10.6±1.4 to -6.1±0.4 mV, P<0.01) and Nav1.8 (from -17.9±1.7 to -13.1±2.1 mV, P<0.05) under Tolperisone. Slope (ka) changed for Nav1.3 (3.80±0.17 to 4.64±0.28, P<0.05) and Nav1.8 (5.63±0.13 to 6.07±0.14, P<0.05). [1]
Steady-state inactivation: Tolperisone shifted half-inactivation potential (Ei0.5) to more negative values for Nav1.2 (-46.8±0.9 to -51.7±1.0 mV, P<0.01), Nav1.3 (-26.1±1.0 to -30.8±1.9 mV, P<0.05), Nav1.4 (-41.1±2.15 to -52.6±3.4 mV, P<0.01), Nav1.7 (-62.6±0.6 to -68.0±1.3 mV, P<0.01). Slope (ki) increased for Nav1.3 (7.75±0.24 to 10.38±0.33, P<0.001) and Nav1.7 (7.47±0.20 to 8.97±0.22, P<0.001). [1]
Recovery from inactivation: Tolperisone did not significantly prolong the fast recovery time constant (τ1) for any isoform, unlike lidocaine which prolonged τ1 for Nav1.3, Nav1.5, Nav1.7. For slow recovery (τ2), Tolperisone significantly prolonged τ2 for Nav1.2 (0.75±0.86 to 3.31±0.48 s, P<0.01), Nav1.5 (1.85±0.26 to 4.24±0.36 s, P<0.05), Nav1.8 (0.62±0.07 to 1.60±0.12 s, P<0.001); shortened τ2 for Nav1.7 (8.86±0.24 to 5.85±0.33 s, P<0.01). The fraction of fast recovery (f1) was reduced by Tolperisone for Nav1.3 (0.67±0.02 to 0.50±0.02, P<0.01) and Nav1.5 (0.74±0.02 to 0.25±0.03, P<0.001). [1]
In vitro, Tolperisone HCl acts as an acute blocker of seven different isoforms of NaV expressed in Xenopus oocytes at micromolar concentrations. It is a voltage-gated sodium channel blocker with a preference for the open/inactivated state. It has an IC50 of 50 μM for the peak-to-peak amplitude of monosynaptic compound action potential. These in vitro studies confirm its mechanism as an ion channel blocker and its potential as a muscle relaxant.

In vivo, Tolperisone HCl acts as an antispastic agent that attenuates monosynaptic, disynaptic, and polysynaptic reflux responses to dorsal root stimulation. It is used for the treatment of spasticity, chronic pain, and other muscle tone disorders. It is a centrally acting muscle relaxant. Its efficacy in reducing muscle spasticity has been established in clinical settings. It is used in the treatment of neurological diseases.

Voltage-dependent sodium channel currents were recorded using the two-electrode voltage clamp technique in Xenopus laevis oocytes. Oocytes were superfused with ND96 solution (96 mmol/L NaCl, 2 mmol/L KCl, 1 mmol/L MgCl2, 1 mmol/L CaCl2, 5 mmol/L HEPES, pH 7.4 with NaOH) at room temperature (20°C). Agarose cushion electrodes were used. Voltage jumps and current recordings were performed via a TL-1-125 interface and PClamp 5.0 software. Cumulative dose-response: holding potential Eh1, constant frequency f, suprathreshold command potential Ecom1 of duration t1. Parameters varied per isoform. For example, Nav1.8: Eh1=-70 mV, f=0.1 Hz, Ecom1=+20 mV, t1=155 ms. [1]
Voltage-dependent activation protocol: membrane held at Eh2 for t2, then stepped to various Ecom3 in 5 mV increments for t3. For Nav1.6: Eh2=-120 mV, t2=6 s, Ecom3 from -60 to +40 mV, t3=182 ms. Currents fitted to equation IpNa+ = Gmax*(Ecom3 - Erev)*(1 - 1/(1+exp((Ecom3-Ea0.5)/ka))). [1]
Steady-state inactivation protocol: held at Eh4 for t4, then prepulse at Ecom5 for t5 (in 5 mV increments), then test pulse to Ecom6. For Nav1.6: Eh4=-120 mV, t4=6 s, t5=1 s, Ecom5 from -115 to +45 mV, Ecom6=-10 mV. Fitted to Boltzmann isotherm. [1]
Recovery from inactivation: held at Eh7 for t7, then inactivating prepulse at Ecom8 for t8, then recovery at -100 mV for increasing intervals (quasi-logarithmic scale), then test pulse to Ecom9. For Nav1.5: Eh7=-120 mV, t7=6 s, t8=3 s, Ecom8=0 mV, Ecom9=-10 mV. Recovery currents fitted to sum of two exponentials. [1]
Non-cellular assays for Tolperisone HCl typically involve measuring its inhibition of voltage-gated sodium or calcium channels. These assays may use purified channels or membrane preparations and measure the displacement of a labeled ligand or the inhibition of ion flux. Its effects on sodium channels can be assessed in oocyte expression systems. These experiments are essential for characterizing its channel blocking properties.

cRNA encoding Nav α-subunits (Nav1.2, Nav1.3, Nav1.4, Nav1.5, Nav1.6, Nav1.7, Nav1.8) and β1 subunit were injected into Xenopus laevis oocytes one day after follicular layer removal by mechanical dissociation. Injection amounts (ng/oocyte): Nav1.2: 0.05-0.25; Nav1.3: 10-15; Nav1.4: 0.25-1.5 (+5 ng β1); Nav1.5: 0.25-1.5; Nav1.6: 2.5; Nav1.7: 25-50; Nav1.8: 50 (+5 ng β1). Oocytes were incubated in NDE solution (ND96 with 2.5 mmol/L pyruvate, 0.1% antibiotics, 1.8 mmol/L CaCl2) at 19°C for 3-5 days prior to recording. [1]
Oocytes were obtained from Xenopus laevis frogs anesthetized with MS 222. Ovarian lobes were removed, placed in CS solution, and shaken at 0.2-0.5 Hz at room temperature (<22°C) to disaggregate follicular layer. Enzymatic disaggregation was stopped when ~50% of oocytes lost follicular layer; remaining follicular layers were removed mechanically by sucking into fire-polished Pasteur pipettes. Oocytes were washed with ND96Ca0 and then placed in NDE. [1]
In vitro cell-based assays for Tolperisone HCl are conducted using neuronal or muscle cells to assess its effects on ion channel activity and excitability. Patch-clamp electrophysiology is used to measure its effects on sodium and calcium currents. The compound's ability to inhibit action potential firing can be assessed. These experiments are crucial for confirming its mechanism of action at the cellular level.

In vivo animal studies for Tolperisone HCl are typically conducted in animal models of spasticity or muscle hypertonia to evaluate its muscle relaxant efficacy. The compound is administered orally or intravenously, and its effects on muscle tone and reflexes are assessed. Behavioral tests may be performed to evaluate its effects on motor function. These studies are essential for validating its in vivo efficacy as a muscle relaxant.

Tolperisone HCl has a molecular weight of 281.82 g/mol. It is an orally active compound. It is well-absorbed after oral administration and is extensively metabolized in the liver. Its half-life and other pharmacokinetic parameters are well-characterized. It is a prescription drug for the treatment of spasticity and muscle disorders.

Tolperisone HCl has an established safety profile from its clinical use. Common side effects include muscle weakness, dizziness, and gastrointestinal disturbances. It can also cause allergic reactions. Comprehensive toxicological data are available from its clinical use. Tolperisone HCl is an approved muscle relaxant in some countries.

Eur J Pharmacol.2006 May 24;538(1-3):5-14;Pain.1996 Oct;67(2-3):417-25.

Tolperisone hydrochloride is an aromatic ketone compound. It is a centrally acting muscle relaxant used to treat symptoms of spasms and muscle cramps. (Excerpt from Martindale Pharmacopoeia, 30th edition, page 1211)

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Tolperisone (also known as AV 650, CAS# 3644-61-9) is a centrally acting muscle relaxant used for the treatment of chronic pain and acute non-specific low back pain. Its mechanism involves blocking voltage-dependent sodium channels via both tonic (use-independent) and use-dependent components. Unlike lidocaine, Tolperisone does not prolong the fast recovery from inactivation of Nav1.3, Nav1.5, and Nav1.7, suggesting differential interaction with the local anesthetic receptor site. The drug exhibits higher potency (lower IC50) than lidocaine on Nav1.6, Nav1.7, and Nav1.8. Tolperisone shifts steady-state inactivation to more negative potentials for several isoforms but has minimal effect on voltage-dependent activation. [1]
Tolperisone HCl (AV 650) is a centrally acting muscle relaxant that inhibits voltage-gated sodium and calcium channels. It is used for the treatment of spasticity and chronic pain. It attenuates monosynaptic, disynaptic, and polysynaptic reflexes. Tolperisone HCl is an approved drug in some countries for the treatment of muscle spasticity and related disorders.

C16H23NO.HCL

281.82

281.154

3644-61-9

Tolperisone-d10 hydrochloride;1185160-65-9

92965

White to off-white solid powder

373.9ºC at 760 mmHg

181-183°C

134.4ºC

4.039

1

2

4

19

262

0

Cl[H].O=C(C1C([H])=C([H])C(C([H])([H])[H])=C([H])C=1[H])C([H])(C([H])([H])[H])C([H])([H])N1C([H])([H])C([H])([H])C([H])([H])C([H])([H])C1([H])[H]

ZBUVYROEHQQAKL-UHFFFAOYSA-N

InChI=1S/C16H23NO.ClH/c1-13-6-8-15(9-7-13)16(18)14(2)12-17-10-4-3-5-11-17;/h6-9,14H,3-5,10-12H2,1-2H3;1H

2-Methyl-3-piperidino-1-p-tolylpropan-1-one hydrochloride

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, avoid exposure to moisture.

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: ≥ 2.5 mg/mL (8.87 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 25.0 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: ≥ 2.5 mg/mL (8.87 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 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 3: ≥ 2.5 mg/mL (8.87 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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Solubility in Formulation 4: 25 mg/mL (88.71 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.

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