OCT1 (IC50 = 36.8 μM); OCT2 (IC50 = 1852.6 μM)

Monocrotaline is a naturally occurring ligand that has strong anti-tumor action and dose-dependent cytotoxicity. Monocrotaline has been shown to have an IC50 of 24.966 µg/mL and a genotoxicity of 2 times IC50 in vitro when tested on HepG2 cells [2].

A rat model of hypertension can be created via animal modeling with monocrotaline. In rats, MCT results in pulmonary vascular syndrome, which is typified by cor pulmonale, pulmonary hypertension (PH), and proliferative pulmonary vasculitis [3]. Monocrotaline-induced animal models have the advantage of closely resembling several important aspects of human pulmonary arterial hypertension (PAH) in preclinical models, such as vascular remodeling, smooth muscle cell proliferation, endothelial dysfunction, inflammatory cytokine upregulation, and right ventricular failure. [4]. The administration of monocrotaline resulted in alterations to several pathways linked to the pathogenesis of peripheral hemorrhage (PH), such as the stimulation of glycolysis, elevations in markers of proliferation, disturbance of carnitine homeostasis, elevations in biomarkers of inflammation and fibrosis, and glutathione production. decrease[5]. Rats given a single dosage of monocrotaline (60 mg/kg i.p.) have considerably higher pulmonary artery pressure, as well as increased right ventricular hypertrophy and pulmonary artery structural remodeling. Then, astragaloside IV (ASIV) was given for 21 days at doses of 10 and 30 mg/kg/d. By enhancing pulmonary arterial remodeling and inflammation, ASIV can prevent pulmonary hypertension [7]. A rat model of pulmonary arterial hypertension (PAH) is induced by monocrotaline (60 mg/kg; ip; single dose) after 3–4 weeks [7]. In a rat model of left lung resection, monocrotaline (60 mg/kg; i.p.; single dose) exhibited significant antitumor efficacy along with dose-dependent cytotoxicity [9]. 1 N HCl was used to dissolve the monocrotaline, which was then diluted with sterile saline and brought to pH 7.4 using 1 N NaOH [7].

Current study systematically investigated the interaction of two alkaloids, anisodine and monocrotaline, with organic cation transporter OCT1, 2, 3, MATE1 and MATE2-K by using in vitro stably transfected HEK293 cells. Both anisodine and monocrotaline inhibited the OCTs and MATE transporters. The lowest IC50 was 12.9 µmol·L-1 of anisodine on OCT1 and the highest was 1.8 mmol·L-1 of monocrotaline on OCT2. Anisodine was a substrate of OCT2 (Km = 13.3 ± 2.6 µmol·L-1 and Vmax = 286.8 ± 53.6 pmol/mg protein/min). Monocrotaline was determined to be a substrate of both OCT1 (Km = 109.1 ± 17.8 µmol·L-1, Vmax = 576.5 ± 87.5 pmol/mg protein/min) and OCT2 (Km = 64.7 ± 14.8 µmol·L-1, Vmax = 180.7 ± 22.0 pmol/mg protein/min), other than OCT3 and MATE transporters. The results indicated that OCT2 may be important for renal elimination of anisodine and OCT1 was responsible for monocrotaline uptake into liver. However neither MATE1 nor MATE2-K could facilitate transcellular transport of anisodine and monocrotaline. Accumulation of these drugs in the organs with high OCT1 expression (liver) and OCT2 expression (kidney) may be expected[8].

Cell Viability Assay[2]
Cell Types: HepG2 cells
Tested Concentrations: 25, 50, 100 and 200 µg/mL
Incubation Duration: 48 h
Experimental Results: Induced apoptosis rate was dose-dependent.

Astragaloside IV blocks monocrotaline‑induced pulmonary arterial hypertension by improving inflammation and pulmonary artery remodeling[7]
Male Sprague-Dawley rats, 8 weeks old weighing 200-230 g, were obtained from the Animal Center of Qiqihar Medical University. The protocol for the present study was approved by the Qiqihar Medical University Institutional Review Board (no. QMU-AECC-2018-27). The rats were housed in a temperature- and humidity-controlled environment with 12-h light/dark cycles. Food and water were available ad libitum. The experiments conformed to the National Institutes of Health guidelines concerning the care and use of laboratory animals, and all animal procedures were approved by the Animal Care and Use Committee of the Qiqihar Medical University. The rats were randomly assigned to 4 groups (8 rats per group) as follows: The control group, the monocrotaline (MCT) group, the MCT + 10 mg/kg/dahy ASIV (ASIV10) group, and the MCT + 30 mg/kg/day ASIV (ASIV30) group. To establish MCT-induced PAH, the rats were administered a single intraperitoneal injection of MCT (60 mg/kg), while the control group received the same volume of saline. MCT was dissolved in 1 N HCl, diluted in sterile saline and adjusted to pH 7.4 with 1 N NaOH. ASIV was initially dissolved in DMSO as a stock solution and further diluted in saline immediately prior to use; the final DMSO concentration was 0.5%. Within hours of the MCT injection, there were signs of pulmonary vascular endothelial damage, but without an increase in pulmonary artery pressure. By 2 weeks, pulmonary artery pressure began to increase, as previously described. At 2 days following the MCT administration, ASIV or the vehicle (0.5% DMSO in saline) were administered intraperitoneally once a day for 21 days.

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A total of 36 male specific-pathogen free Sprague-Dawley rats (age, 6–8 weeks; weight, 300–350 g) were kept in a conventional room at 22±2°C, a relative humidity of 55±10% and a 12-h light/dark cycle. Rats had access to food and water ad libitum. The rats were divided into the following three equal groups at random: Control group, where rats received no treatment; model group, where rats underwent a left pneumonectomy plus subcutaneous injection of 60 mg/kg monocrotaline (MCT), a natural ligand exhibiting dose-dependent cytotoxicity with potent antineoplastic activity, at 7 days following the procedure; PTX group, where rats underwent the same procedure as those in the model group plus administration of 2 mg/kg PTX via the caudal vein (21) daily for 1 week at 3 weeks following injection of MCT[9].

Absorption, Distribution and Excretion
Following subcutaneous injection of monoclotaline in rats, 50-70% of the dose is present in the urine as unchanged monoclotaline… The highest concentrations of monoclotaline (or its metabolites) are found in the liver, kidneys, and stomach. The pyrrolizidine alkaloid monoclotaline has been shown to cause liver necrosis and pulmonary hypertension in rats. To better understand its mechanism of action, tissue distribution and covalent binding studies were performed at 4 and 24 hours after subcutaneous injection of 14C-labeled monoclotaline (60 mg/kg, 200 μCi/kg). At 4 hours, the equivalent concentrations of monoclotaline in erythrocytes, liver, kidneys, lungs, and plasma were 85, 74, 67, 36, and 8 nmol/g tissue, respectively, while the covalently bound concentrations of monoclotaline in these tissues were 125, 132, 39, 64, and 44 pmol/mg protein, respectively. The 24-hour tissue distribution concentrations of monoclotalpine equivalents were 49, 25, 9, 10, and 2 nmol/g tissue in erythrocytes, liver, kidneys, lungs, and plasma, respectively, while the covalently bound concentrations of monoclotalpine in liver, kidneys, and lungs were 74, 28, and 55 pmol/mg protein, respectively. We also investigated the kinetics of ¹⁴C monoclotalpine (60 mg/kg, 10 μCi/kg, intravenously), showing rapid elimination of the radioactive material, with approximately 90% of the injected radioactive material recovered in urine and bile within 7 hours. The plasma radioactivity level decreased from 113 nmol/g monoclotalpine equivalent to 11 nmol/g at 7 hours, while the erythrocyte radioactivity level decreased from 144 nmol/g to only 81 nmol/g at the same time point. The significant retention of monoclotalpine equivalents in erythrocytes suggests that erythrocytes may act as carriers of metabolites from the liver to other organs, including the lungs, and may play a role in pulmonary toxicity.

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Metabolism/Metabolites
Studies on monoclotaline have confirmed that the mixed-function oxidase system of rat liver microsomal components can generate pyrrole metabolites. Dehydromonoclonaline (monoclonaline pyrrole) is highly cytotoxic, causing lung, heart, vascular, and liver damage similar to that of the parent alkaloid. It is a highly reactive alkylating agent that, upon intracellular generation, immediately reacts with cellular components to generate soluble or conjugated secondary metabolites, or hydrolyzes to generate the dehydroamino alcohol—dehydrorhelotrin.
In male rat liver microsomes pretreated with phenobarbital, all 13 tested alkaloids were metabolized to N-oxides and pyrroles. These two metabolic pathways appear to be independent. The ratio of N-oxides to pyrrole metabolites varied depending on the type of ester: the highest ratio was found in open-ring diester alkaloids, while the lowest ratios were found in 12-membered macrocyclic diesters and monoesters. Monoclotaline was one of the tested compounds.
This study compared the metabolism of the pyrrolizidine alkaloid (14)C monoclostaline using rat and guinea pig liver microsomes. …In guinea pigs, esterase hydrolysis accounted for 92% of the metabolism; however, rats did not show esterase activity. This result may explain the resistance of guinea pigs to the toxicity of pyrrolizidine alkaloids. Dehydropyrrole was found to be the main pyrrole metabolite in guinea pigs, although colorimetric analysis showed the presence of multiple pyrrole fractions in rat microsomal incubation solutions.
This report shows that the Ehrlich reagent-positive metabolites of monoclostaline and senna were excreted in the urine of male rats as (+/-)-6,7-dihydro-7-hydroxy-1-hydroxymethyl-5H-pyrrolazine N-acetylcysteine conjugates…This finding suggests that the active metabolites of pyrrolizidine alkaloids produced in the liver can survive in the aqueous environment of the circulatory system in the form of glutathione conjugates or thiouric acid. For more complete metabolite/metabolite data on monoclonal clotazone (7 metabolites), please visit the HSDB record page.

Interactions
SKF 525a and metheprone both protected aged rats from monoclotalpine-induced right ventricular and pulmonary hypertrophy. Mixed-function oxidase inhibitors were more effective than thiol substitution in mitigating monoclotalpine toxicity. Protective effects were weaker in young rats. Dietary supplementation with ethoxyquinoline protected mice from the lethality of monoclotalpine and acute hepatotoxicity (as determined by plasma alanine aminotransferase and aspartate aminotransferase levels). Dietary supplementation with cysteine (1%) also protected mice from the lethality of this alkaloid, but not from acute hepatotoxicity. Except for ethoxyquinoline, other feed additives did not increase hepatic glutathione levels. Dietary supplementation with either ethoxyquinoline or cysteine significantly increased glutathione S-transferase activity using dichloronitrobenzene as a substrate. Dietary supplementation with ethoxyquinoline increased hepatic cytochrome P-450 levels and promoted the in vitro conversion of monoclotalpine to pyrrole metabolites via hepatic microsomes. Although ethoxyquinoline did not reduce the in vivo activation of monoclotalpine, it still protected mice from the lethality and hepatotoxicity of monoclotalpine. Therefore, its mechanism of action is likely due to increased hepatic glutathione levels, thereby enhancing the detoxification process. Dietary supplementation with 0.25% and 0.75% butylated hydroxyanisole (BHA) protected juvenile female mice from acute monoclotalpine toxicity. This protective effect was associated with decreased levels of pyrrole metabolites in the liver, decreased activity of hepatic aminopyrine demethylase, and a reduced rate of in vitro microsomal conversion of monoclotalpine to pyrrole metabolites. BHA also increased hepatic sulfhydryl levels and cytoplasmic glutathione S-transferase activity. Dietary supplementation with cysteine (1%) was less protective against monoclotalpine toxicity than BHA. The LD50 values of monoclotalpine in the control and cysteine groups were 259 mg/kg and 335 mg/kg, respectively.

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Non-human toxicity values
Rat intravenous LD50: 92 mg/kg
Mouse intraperitoneal LD50: 259 mg/kg
Rat oral LD50: 66 mg/kg
Mouse intravenous LD50: 261 mg/kg

[1]. The monocrotaline model of pulmonary hypertension in perspective. Am J Physiol Lung Cell Mol Physiol. 2012 Feb 15;302(4):L363-9.
[2]. Antineoplastic activity of monocrotaline against hepatocellular carcinoma. Anticancer Agents Med Chem. 2014;14(9):1237-48.
[3]. Mechanisms and pathology of monocrotaline pulmonary toxicity. Crit Rev Toxicol. 1992;22(5-6):307-25.
[4]. Exploring the monocrotaline animal model for the study of pulmonary arterial hypertension: A network approach. Pulm Pharmacol Ther. 2015 Dec;35:8-16.
[5]. Metabolic Changes Precede the Development of Pulmonary Hypertension in the Monocrotaline Exposed RatLung. PLoS One. 2016 Mar 3;11(3):e0150480.
[6]. Experimental animal models of pulmonary hypertension: Development and challenges. Animal Model Exp Med. 2022 Sep; 5(3):207-216.
[7]. Astragaloside IV blocks monocrotaline‑induced pulmonary arterial hypertension by improving inflammation and pulmonary artery remodeling. Int J Mol Med. 2021 Feb;47(2):595-606.
[8]. An in vitro study on interaction of anisodine and monocrotaline with organic cation transporters of the SLC22 and SLC47 families. Chin J Nat Med. 2019 Jul;17(7):490-497.
[9]. Effects of paclitaxel intervention on pulmonary vascular remodeling in rats with pulmonary hypertension. Exp Ther Med. 2019 Feb;17(2):1163-1170.

According to an independent committee of scientific and health experts, monoclotaline may be carcinogenic. Monoclotaline is a pyrrolizidine alkaloid. It has been reported in Crotalaria sessiliflora, Crotalaria retusa, and other organisms with relevant data. Monoclotaline is a pyrrolizidine alkaloid and a toxic plant component that can cause poisoning in livestock and humans through the ingestion of contaminated grains and other foods. This alkaloid can cause pulmonary hypertension, right ventricular hypertrophy, and pulmonary vascular disease. Oral magnesium supplementation significantly reduced the cardiopulmonary lesions. Mechanism of Action: The toxicology of monoclotaline is complex, and the mechanisms by which it causes lung injury, pulmonary hypertension, and right ventricular enlargement remain unclear. Monoclotaline is bioactivated in the liver to active electrophilic pyrroles, which then reach the lungs via the bloodstream, causing lung damage. When rats were intravenously injected with low doses of monoclotallin pyrrole, lung injury and pulmonary hypertension did not appear until several days later. Moderate reduction of platelet count before and after lung injury alleviated subsequent right ventricular enlargement, suggesting a weakened response to pulmonary hypertension induced by monoclotallin pyrrole. This observation prompted us to investigate the role of platelet-derived mediators in the cardiopulmonary response induced by monoclotallin pyrrole. Compared to the control group, stable thromboxane A2 (TxA2) analogues in rats treated with monoclotallin pyrrole resulted in a significant increase in right ventricular pressure, and TxB2 production from lung tissue isolated from rats treated with monoclotallin pyrrole was also higher than in the control group. However, in vivo experiments showed that the use of drugs that inhibit thromboxane synthesis or antagonize thromboxane action did not protect the body from damage caused by monoclotallin pyrrole. Serotonin, another vasoactive mediator released by platelets, caused excessive vasoconstriction in isolated rat lung tissue treated with monoclotallin pyrrole. Furthermore, monoclotalin pyrrole treatment impairs the clearance and inactivation of circulating serotonin in pulmonary vessels. However, in vivo experiments showed that the use of serotonin receptor antagonists did not alleviate the cardiopulmonary toxicity of monoclotalin pyrrole. These results suggest that neither TxA2 nor serotonin are the sole mediators of monoclotalin pyrrole-induced pulmonary toxicity. Therefore, the pathogenesis of pulmonary toxicity induced by monoclotalin pyrrole involving platelets remains a mystery. Monoclotalin can cause changes in arterial smooth muscle contraction response, altered smooth muscle Na/K-ATPase activity, platelet factor release, and reduced 5-hydroxytryptamine transport in vascular endothelial cells. This study investigated the effects of intraperitoneal injection of monoclotalin on the activity of hepatic epoxide hydrolase and aryl hydrocarbon hydroxylase in young male Long Evans rats. The results showed that monoclotalin failed to stimulate epoxide hydrolase activity, but instead reduced the activities of glutathione S-transferase, aminopyrine demethylase, and aryl hydrocarbon hydroxylase. In vitro experiments showed that, apart from a slight stimulation of epoxide hydrolase activity and a slight decrease in aminopyrine demethylase activity, the studied hepatic drug-metabolizing enzymes were unaffected. ...The active metabolite of monoclotamyl, dehydromonoclotamyl (DHM), can alkylate the N7 guanine of DNA, with a particular preference for 5'-GG and 5'-GA sequences. Furthermore, electrophoresis and electron microscopy confirmed that it can also generate piperidine-resistant and heat-resistant multiple DNA crosslinks. Based on these findings, we propose that DHM rapidly polymerizes to form a structure capable of crosslinking multiple DNA fragments.

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Therapeutic Use
/Exptl Ther/ The antitumor effects of 22 pyrrolizidine alkaloids and their derivatives were investigated in mice with adenocarcinoma 755, L-1210 leukemia, or sarcoma 180, rats with Walker 256 carcinosarcoma (intramuscular or subcutaneous injection), and kb cancer cell cultures; each compound was also tested in mice with ascites, Ehrlich carcinoma, and hamsters with plasmacytoma 1. Monoclotaline (NSC-28693) showed significant activity against solid tumors in all three tumor types and p-1 according to CCNSC criteria (tumor volume reduction of 58% or more). Monoclotaline n-oxide did not show significant activity in any of the tested systems.
/Exptl Ther/ Monoclotaline from Crotalaria sessiliflora has been shown to be effective against human skin cancer and cervical cancer.

C16H23NO6

325.36

325.152

C, 59.07; H, 7.13; N, 4.31; O, 29.50

315-22-0

9415

Prisms from absolute alcohol
Colorless

1.4±0.1 g/cm3

537.3±50.0 °C at 760 mmHg

204ºC (dec.)(lit.)

278.7±30.1 °C

0.0±3.2 mmHg at 25°C

1.586

-0.37

2

7

0

23

575

5

O=C(O[C@]1([H])CCN2[C@]1([H])C(CO3)=CC2)[C@H](C)[C@@](C)(O)[C@@](C)(O)C3=O

QVCMHGGNRFRMAD-XFGHUUIASA-N

InChI=1S/C16H23NO6/c1-9-13(18)23-11-5-7-17-6-4-10(12(11)17)8-22-14(19)16(3,21)15(9,2)20/h4,9,11-12,20-21H,5-8H2,1-3H3/t9-,11+,12+,15+,16-/m0/s1

20-Norcrotalanan-11,15-dione, 14,19-dihydro-12,13-dihydroxy-, (13-alpha,14-alpha)- (9CI)

NSC 28693; NSC-28693; monocrotaline; Crotaline; 315-22-0; Monocrotalin; (-)-Monocrotaline; CHEBI:6980; Retronecine cyclic 2,3-dihydroxy-2,3,4-trimethylglutarate; (13-alpha,14-alpha)-14,19-Dihydro-12,13-dihydroxy-20-norcrotalanan-11,15-dione; NSC28693

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 (e.g. under nitrogen), avoid exposure to moisture and light.

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

1M HCl : 200 mg/mL (~614.70 mM)
DMSO : ~25 mg/mL (~76.84 mM)
H2O : ~2 mg/mL (~6.15 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (7.68 mM) (saturation unknown) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
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 (7.68 mM) (saturation unknown) in 5% DMSO + 95% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution.
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 (6.39 mM) 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 4: ≥ 2.08 mg/mL (6.39 mM) 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 5: ≥ 2.08 mg/mL (6.39 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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Solubility in Formulation 6: ≥ 0.5 mg/mL (1.54 mM)(saturation unknown) in 1% DMSO 99% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.

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Solubility in Formulation 7: 4.17 mg/mL (12.82 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication (<60°C).

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Solubility in Formulation 8: 21 mg/mL (64.54 mM) in 20% HP-β-CD in Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; Need ultrasonic and warming and heat to 53°C.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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