- Tacrine hydrochloride hydrate targets acetylcholinesterase (AChE) with a Ki value of 0.045 μM and butyrylcholinesterase (BChE) with a Ki value of 0.37 μM [1]
The primary targets of tacrine hydrochloride hydrate are acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). As a reversible inhibitor, tacrine binds to the reversible inhibition sites of both enzymes, preventing acetylcholine hydrolysis, thereby increasing acetylcholine levels in the synaptic cleft and enhancing cholinergic neurotransmission. Kinetic analysis indicates that tacrine acts as a mixed-type inhibitor (exhibiting both competitive and non-competitive components) for both enzymes, with higher affinity for AChE (Ki = 0.045 μM) than for BChE (Ki = 0.37 μM). Additionally, studies have revealed that tacrine possesses multi-target activity, including acting as a histamine N-methyltransferase inhibitor, affecting serotonin and norepinephrine uptake, and serving as a low-affinity antagonist of the N-methyl-D-aspartate (NMDA) receptor.

In a suspended concentration manner, tacrine hydrochloride (aqueous complex) (12.5 to 37.5 nM) inhibits both human blood butylacetylcholinesterase and venom acetatecholinesterase. AChE from snake venom has an IC50 of 31 nM, while BChE from humans has an IC50 of 25.6 nM[1].

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- Tacrine hydrochloride hydrate potently inhibited the activity of human erythrocyte AChE and equine serum BChE in a concentration-dependent manner. Kinetic analysis revealed it acted as a mixed-type inhibitor for both enzymes, with higher affinity for AChE (Ki = 0.045 μM) than BChE (Ki = 0.37 μM). The inhibition curves showed concentration-response relationships, with IC₅₀ values consistent with the Ki values, confirming its specific inhibitory effect on cholinesterases [1]

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In vitro studies demonstrate that tacrine hydrochloride hydrate exhibits potent inhibitory activity against both AChE and BChE. The half-maximal inhibitory concentrations (IC₅₀) are 31 nM for snake venom AChE and 25.6 nM for human serum BChE. In concentration-dependent experiments, tacrine (12.5-37.5 nM) inhibits both enzymes in a concentration-related manner. Kinetic analysis reveals mixed-type inhibition for both enzymes, with inhibition constants (Ki) of 13 nM for snake venom AChE and 12 nM for human serum BChE calculated by Lineweaver-Burk plots. Furthermore, tacrine exhibits activities affecting acetylcholine synthesis and release, as well as modulating muscarinic acetylcholine receptors.

During reacquisition, administration of tacrine hydrochloride (hydrate) can also change the absolute levels of cocaine self-regulation. Body weight was regained by roughly 0.5% four days following intravenous tacrine hydrochloride hydrate therapy. The osmotic pump conduit method of tacrine hydrochloride (hydrate) does not change the linear or repeated cocaine-induced locomotor activity. During recovery, there was no discernible primary effect or reaction from tacrine hydrochloride (hydrate) therapy on active lever responses. Conditioned place preference was used to measure phosphatase levels, and post hoc comparisons revealed that self-prepared cocaine had much lower levels than phosphoric acid treated with saline [2].

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- In male Wistar rats, Tacrine hydrochloride hydrate exhibited enduring effects on cocaine-reinforced behavior: (1) Conditioned-place preference (CPP) assay: Cocaine (15 mg/kg, i.p.) induced significant CPP. Co-administration of Tacrine hydrochloride hydrate (1, 3, 10 mg/kg, i.p.) during CPP training dose-dependently attenuated cocaine-induced CPP, with the 10 mg/kg dose showing the most potent effect. (2) Temporal separation experiment: Administering Tacrine hydrochloride hydrate (10 mg/kg, i.p.) 24 hours after cocaine training still reduced CPP expression, indicating a long-lasting effect independent of immediate drug reward association. (3) Reinstatement assay: After extinguishing cocaine-induced CPP, priming with cocaine (7.5 mg/kg, i.p.) reinstated CPP. Pretreatment with Tacrine hydrochloride hydrate (10 mg/kg, i.p.) significantly blocked this reinstatement, suggesting it modulates cocaine's motivational properties [2]

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In vivo studies demonstrate that tacrine hydrochloride hydrate modulates both cognitive behavior and drug reward behavior in animal models. In male Wistar rats, tacrine dose-dependently attenuates cocaine-induced conditioned place preference (CPP), with the 10 mg/kg dose showing the most potent effect. Tacrine also blocks cocaine-induced reinstatement of CPP, suggesting modulation of cocaine's motivational properties. In cognitive research, tacrine serves as a core pharmacophore for developing novel multi-target-directed ligands, with tacrine-based hybrid molecules showing potential for improving cognitive function in animal models. However, significant interindividual variation in tacrine's efficacy exists, which is thought to influence both therapeutic response and incidence of adverse effects in patients.

- Cholinesterase activity inhibition assay: Human erythrocyte AChE and equine serum BChE were purified and prepared into reaction systems with appropriate buffers. Different concentrations of Tacrine hydrochloride hydrate were added to the systems, followed by the addition of specific substrates (acetylthiocholine for AChE, butyrylthiocholine for BChE). The reaction was incubated at 37°C for a fixed time, and the formation of thiocholine (product) was detected by a colorimetric method using a chromogenic reagent. The enzyme activity was calculated based on absorbance values, and the inhibition rate was determined relative to the control group. Kinetic parameters (Ki values) were derived from Lineweaver-Burk plots to identify the inhibition type [1]

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Enzyme Source Preparation: Purify human erythrocyte AChE and horse serum BChE, or use snake venom-derived AChE. Substrate Preparation: Use the Ellman method with acetylthiocholine iodide (ATChI) as substrate, co-prepared with DTNB in phosphate buffer (pH 7.4). Inhibitor Incubation: Pre-incubate varying concentrations of tacrine hydrochloride hydrate (e.g., 0-100 nM) with the enzyme in buffer at 37°C. Reaction Initiation and Detection: Initiate the reaction by adding substrate, continuously monitor absorbance change at 412 nm, and record the reaction rate. Kinetic Analysis: Plot competitive inhibition curves using Lineweaver-Burk double reciprocal plots to calculate IC₅₀ values (AChE: 31 nM, BChE: 25.6 nM) and inhibition constant Ki.

Cell Culture: Culture neuronal cell lines (e.g., SN56 cells) or hepatocyte cell lines (e.g., HepG2) in DMEM medium containing 10% fetal bovine serum at 37°C in a 5% CO₂ incubator. Drug Treatment: After seeding cells in culture plates, treat with varying concentrations of tacrine hydrochloride (e.g., 0.1-100 μM) and incubate for 24-72 hours. Viability Assay: Measure cell viability using MTT or CCK-8 assays to calculate IC₅₀ values. Cholinesterase Activity Assay: Lyse cells and measure intracellular AChE and BChE activity using the Ellman method to assess enzyme inhibition by the drug. Calcium Channel Assay: Measure the inhibitory effect of tacrine on depolarization-induced calcium influx through L-type calcium channels in SN56 neuronal cells. Neurotransmitter Uptake Assay: Measure the effect of tacrine on serotonin and norepinephrine uptake.

- Cocaine-induced CPP model in Wistar rats: Male Wistar rats were randomly divided into control, cocaine-only, and Tacrine hydrochloride hydrate + cocaine groups (n=8–10 per group). The CPP apparatus consisted of two distinct compartments with different visual and tactile cues. (1) Training phase: Rats in the cocaine group received cocaine (15 mg/kg, i.p.) before being placed in the drug-paired compartment for 30 minutes, while the control group received saline. The Tacrine hydrochloride hydrate groups were administered Tacrine hydrochloride hydrate (1, 3, 10 mg/kg, i.p.) 30 minutes before cocaine injection, followed by placement in the drug-paired compartment. This training was repeated for 4 days. (2) Testing phase: 24 hours after the last training, rats were allowed free access to both compartments for 15 minutes, and the time spent in each compartment was recorded to calculate CPP scores. (3) Temporal separation experiment: Rats were trained with cocaine for 4 days, then administered Tacrine hydrochloride hydrate (10 mg/kg, i.p.) 24 hours after the last training, followed by the CPP test 24 hours later. (4) Reinstatement experiment: After CPP extinction (daily saline injections in both compartments for 5 days), rats were pretreated with Tacrine hydrochloride hydrate (10 mg/kg, i.p.) 30 minutes before cocaine priming (7.5 mg/kg, i.p.), and the CPP test was conducted 24 hours later [2]

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Animal Selection: Use male Wistar rats or ICR/C57BL/6 mice as experimental animals. Dosing Regimen: Tacrine can be administered via oral gavage, intraperitoneal injection (e.g., 10 mg/kg), or intravenous injection. Behavioral Assessment: Cognitive Function: Assess learning and memory capacity using Morris water maze, Y-maze, or passive avoidance tests. Drug Addiction: Use conditioned place preference (CPP) assays to evaluate the modulatory effects of tacrine on cocaine-induced CPP acquisition, expression, and reinstatement. Toxicological Assessment: Collect blood samples for liver function biochemical tests; after euthanasia, collect liver, kidney, brain, and other tissues for histopathological examination. Data Analysis: Compare cognitive behavior, drug addiction behavior, and biochemical parameters between treatment and control groups.

Tacrine hydrochloride is rapidly and well absorbed after oral administration, with peak plasma concentrations (Cmax) achieved within 0.5 to 3 hours following a single 20-50 mg oral dose. However, its oral bioavailability is low (6-36%) with significant interindividual variation, primarily due to extensive first-pass metabolism. Tacrine exhibits wide tissue distribution with a large volume of distribution, showing high concentrations in kidney, liver, adrenal gland, and brain tissue in animal models. The elimination half-life is short: 1.5-2.5 hours after single oral and intravenous doses, and 2.9-3.6 hours after multiple oral doses. Tacrine is primarily metabolized in humans by hepatic CYP1A2 to 1-, 2-, 4-, and 7-hydroxylated metabolites, and excreted in urine as mono- and dihydroxylated metabolites and glucuronide conjugates.

Hepatotoxicity is the most clinically significant adverse effect of tacrine and the primary reason for its eventual withdrawal from the market. The mechanism of this toxicity is linked to its hepatic metabolism: tacrine is metabolized by CYP1A2 to protein-reactive metabolites (conversion of 7-hydroxy-tacrine to a quinone methide intermediate), which can deplete intracellular glutathione and covalently bind to proteins, leading to hepatocellular necrosis. In clinical studies, approximately 30-50% of patients developed elevated serum transaminases (ALT > 3× upper limit of normal), typically occurring within 4-8 weeks after treatment initiation. According to safety data sheets, tacrine hydrochloride is an acute toxic substance: toxic if inhaled (H331), harmful if swallowed (H302), causes skin irritation (H315), causes serious eye irritation (H319), and may cause respiratory irritation (H335). It has UN number UN2811, hazard class 6.1, and should be stored locked up, with waste disposal in accordance with hazardous chemical regulations. This compound is for research use only and is not intended for human use.

[1]. Inhibition of two different cholinesterases by tacrine. Chem Biol Interact. 2006 Aug 25;162(2):165-71.

[2]. Enduring effects of tacrine on cocaine-reinforced behavior: Analysis by conditioned-place preference, temporal separation from drug reward, and reinstatement. Pharmacol Res. 2015 Jul;97:40-7.

See also: tacrine hydrochloride (note moved to); tacrine (note moved to).

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- tacrine hydrochloride hydrate is a cholinesterase inhibitor that exerts its pharmacological effects by blocking acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), thereby increasing acetylcholine levels in the synaptic cleft [1]
- Its persistent effect on cocaine-enhancing behavior suggests that it may regulate the cholinergic system’s involvement in drug reward and addiction-related processes by interacting with central cholinergic receptors or downstream signaling pathways associated with motivation and reward [2]
- tacrine hydrochloride hydrate’s mixed inhibition of cholinesterase suggests that it binds to both the active and allosteric sites of the enzyme, resulting in its potent inhibitory activity [1]

C13H17CLN2O

252.739882230759

252.102

206658-92-6

Tacrine hydrochloride;1684-40-8; 321-64-2; 206658-92-6

6420002

White to off-white solid powder

3

3

0

17

229

0

C1CCC2=NC3=CC=CC=C3C(=C2C1)N.O.Cl

PXGRMZYJAOQPNZ-UHFFFAOYSA-N

InChI=1S/C13H14N2.ClH.H2O/c14-13-9-5-1-3-7-11(9)15-12-8-4-2-6-10(12)13;;/h1,3,5,7H,2,4,6,8H2,(H2,14,15);1H;1H2

1,2,3,4-tetrahydroacridin-9-amine;hydrate;hydrochloride

206658-92-6; RefChem:410547; DTXSID101092736; 1,2,3,4-Tetrahydro-9-acridinamine hydrochloride hydrate (1:?:?); 7149-50-0;

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)

H2O : ~100 mg/mL
DMSO : ~32 mg/mL

Solubility in Formulation 1: ≥ 2.5 mg/mL (Infinity 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 (Infinity 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 (Infinity 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: 100 mg/mL (Infinity 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.)