Natural alkaloid

Harmaline and its derivatives were identified as anti-MDR agents against various highly resistant and Pakistani MDR clinical isolates of E. coli. These compounds may serve as the leads for further studies towards the development of treatment against the infections caused by MDR E. coli.[1]

---

Experiments were conducted on Control (Salin) and Experiment (Harmaline) groups, generating a dataset for developing predictive models. Because the dataset has a limited number of samples, we utilized models that are effective with small datasets. Among different groups of regression models (linear, ensemble, and tree models), the ensemble models, specifically the LGB method, can achieve better performance. The results demonstrate accurate prediction of first spike latency, with an average mean squared error of 0.0002 and mean absolute error of 0.01 in 10-fold cross-validation. The research suggests the potential of machine learning in forecasting the first spike latency, allowing reliable estimation without the need for extensive animal testing. This intelligent predictive system facilitates efficient analysis of first spike latency changes in both healthy and unhealthy brain cells, streamlining experimentation and providing more detailed insights into the captured signals.[2]

Depression is a mental disorder characterised by persistent low mood, anhedonia and cognitive impairment that affects an estimated 3.8% of the world's population, including 5% of adults. Peganum harmala L. (P. harmala) is a medicinal plant and has been reported to be effective against Alzheimer's disease, Parkinson's disease and depression. The present study was aimed to evaluate the behavioral and pharmacological effects of P. harmala seed extract in rats exposed to chronic unpredictable mild stress (CUMS) in vivo and to investigate the mechanism of action. CUMS-exposed rats were treated with P. harmala extract (75 and 150 mg/kg, i.p.) for 2 weeks. HPLC analysis was used to determine the concentration of Harmaline and harmine alkaloids in the extract. Heavy metal analysis in seeds was performed by ICP-MS. Our results showed that P. harmala at the dose of 150 mg/kg significantly reduced the depressive-like behaviors in CUMS-exposed rats, as evidenced by increased sucrose consumption in the sucrose preference test (SPT), decreased immobility time in the forced swim test (FST) and plasma corticosterone levels, increased the time spent in open arms in the elevated plus maze (EPM), and improved memory and learning in the passive avoidance test (PAT). In addition, P. harmala decreased monoamine oxidase-A (MAO-A) levels, and increased serotonin (5-HT), dopamine (DA), and noradrenaline (NA) levels in the brains of rats exposed to CUMS. P. harmala decreased the expression of the pro-inflammatory transcription factor nuclear factor-κB (NF-κB), and increased the antioxidant nuclear factor erythroid 2-related factor 2 (Nrf2) in rat brain. Furthermore, P. harmala improved brain-derived neurotrophic factor (BDNF) and tropomyosin receptor kinase B (TrkB) protein expression in rat brain. In conclusion, P. harmala at a dose of 150 mg/kg is more effective in preventing depressive-like behavior in CUMS-exposed rats by improving neurotransmitter levels, reducing oxidative stress, suppressing neuroinflammation and activating the BDNF/TrkB pathway, all of which are important in the pathogenesis of depression.[2]

The psychotropic beta-carboline alkaloids, showing high affinity for 5-hydroxytryptamine, dopamine, benzodiazepine, and imidazoline receptors and the stimulation of locus coeruleus neurons, are formed endogenously from tryptophan-derived indolealkylamines through the Pictet-Spengler condensation with aldehydes in both plants and mammals. Cytochromes P450 1A1 (18.5), 1A2 (20), and 2D6 (100) catalyzed the O-demethylation of harmaline, and CYP1A1 (98.5), CYP1A2 (35), CYP2C9 (16), CYP2C19 (30), and CYP2D6 (115) catalyzed that of harmine (relative activities). The dehydrogenation/aromatization of harmaline to harmine was not carried out by aromatase (CYP19), CYP1A2, CYP2C9, CYP2D6, CYP3A4, pooled recombinant cytochromes P450, or human liver microsomes (HLMs). Kinetic parameters were calculated for the O-demethylations mediated by each isozyme and by pooled HLMs. K(cat) (min(-1)) and Ku (uM) values for harmaline were: CYP1A1, 10.8 and 11.8; CYP1A2, 12.3 and 13.3; CYP2C9, 5.3 and 175; CYP2C19, 10.3 and 160; and CYP2D6, 39.9 and 1.4. Values for harmine were: CYP1A1, 45.2 and 52.2; CYP1A2, 9.2 and 14.7; CYP2C9, 11.9 and 117; CYP2C19, 21.4 and 121; and CYP2D6, 29.7 and 7.4. Inhibition studies using monoclonal antibodies confirmed that CYP1A2 and CYP2D6 were the major isozymes contributing to both harmaline (20% and 50%, respectively) and harmine (20% and 30%) O-demethylations in pooled HLMs. The turnover numbers for CYP2D6 are among the highest ever reported for a CYP2D6 substrate. Finally, CYP2D6-transgenic mice were found to have increased harmaline and harmine O-demethylase activities as compared with wild-type mice. These findings suggest a role for polymorphic CYP2D6 in the pharmacology and toxicology of harmine and harmaline.PMID:12649384 ; Yu AM et al; J Pharmacol Exp Ther 305 (1): 315-22 (2003)

Background: The present study aimed to elucidate the potential anticancer activity and mechanism of P. harmala's alkaloid extract, harmine (HAR), and harmaline (HAL) in HCT-116 colorectal cancer cells. Methods and results: P. harmala's alkaloid was extracted from harmala seeds. HCT-116 cells were treated with P. harmala's alkaloid extract, HAR and HAL. Cytotoxicity was determined by MTT assay, apoptotic activity detected via flow cytometry and acridine orange (AO)/ethidium bromide (EB) dual staining, and cell cycle distribution analyzed with flow cytometry. The mRNA expression of Bcl-2-associated X protein (Bax) and glycogen synthase kinase-3 beta (GSK3β) was measured by real-time PCR. Furthermore, the expression of Bax, Bcl-2, GSK3β and p53 proteins, were determined by western blotting. The findings indicated that, P. harmala's alkaloids extract, HAR and HAL were significantly cytotoxic toward HCT116 cells after 24 and 48 h of treatment. We showed that P. harmala's alkaloid extract induce apoptosis and cell cycle arrest at G2 phase in the HCT116 cell line. Downregulation of GSK3β and Bcl-2 and upregulation of Bax and p53 were observed. Conclusion: The findings of this study indicate that the P. harmala's alkaloid extract has anticancer activity and may be further investigated to develop future anticancer chemotherapeutic agents. [4] Mol Biol Rep. 2024 Jun 13;51(1):732. doi: 10.1007/s11033-024-09655-7. https://pubmed.ncbi.nlm.nih.gov/38872006/

Absorption, Distribution and Excretion
Halamine is a known inhibitor of monoamine oxidase type A (MAO) and has been found in the adult brains of various animals. Studies have shown that fetuses born to mother rats injected with halamine 2–4 hours before cesarean section exhibit elevated levels of dopamine and serotonin (5-HT). Similar stimulatory effects were observed with the norepinephrine metabolite 3-methoxy-4-hydroxyphenylethylene glycol (MHPG), but without a significant effect on norepinephrine itself. Dopamine metabolite 3,4-dihydroxyphenylacetic acid (DOPAC) and 5-HT metabolite 5-hydroxyindoleacetic acid (5-HIAA) were both decreased under the same treatment. These results suggest that halamine or one of its metabolites may cross the placental barrier and affect the fetal brain system, not only as an inhibitor of type A monoamine oxidase (i.e., relatively 5-HT specific), but also as a stimulator of aldosterone reductase or catechol-O-methyltransferase (COMT), or as a drug that inhibits the binding, efflux, or turnover of biogenic amine metabolites (such as MHPG).

---

Metabolites
Psychoactive β-carboline alkaloids have high affinity for serotonin, dopamine, benzodiazepines, and imidazoline receptors and can stimulate locus coeruleus neurons. These alkaloids are endogenously formed in both plants and mammals from tryptophan-derived indolealkylamines and aldehydes via the Pictet-Spengler condensation reaction. Cytochrome P450 1A1 (18.5), 1A2 (20), and 2D6 (100) catalyzed the O-demethylation of halamine, while CYP1A1 (98.5), CYP1A2 (35), CYP2C9 (16), CYP2C19 (30), and CYP2D6 (115) catalyzed the O-demethylation of halamine (relative activity). Aromatases (CYP19), CYP1A2, CYP2C9, CYP2D6, CYP3A4, a mixture of recombinant cytochrome P450, or human liver microsomes (HLM) could not catalyze the dehydrogenation/aromatization of halamine to halamine. Kinetic parameters of the O-demethylation reactions mediated by each isoenzyme and recombinant HLM were calculated. Halmin's K(cat) (min(-1)) and Ku(uM) values were: CYP1A1, 10.8 and 11.8; CYP1A2, 12.3 and 13.3; CYP2C9, 5.3 and 175; CYP2C19, 10.3 and 160; and CYP2D6, 39.9 and 1.4. Halmin's K(cat) and Ku(uM) values were: CYP1A1, 45.2 and 52.2; CYP1A2, 9.2 and 14.7; CYP2C9, 11.9 and 117; CYP2C19, 21.4 and 121; and CYP2D6, 29.7 and 7.4. Inhibition studies using monoclonal antibodies confirmed that CYP1A2 and CYP2D6 are the major isoenzymes for halamine (20% and 50%, respectively) and halamine (20% and 30%, respectively) O-demethylation in mixed human liver microsomes (HLM). The turnover rate of CYP2D6 is among the highest reported for CYP2D6 substrates to date. Furthermore, CYP2D6 transgenic mice showed increased halamine and halamine O-demethylase activities compared to wild-type mice. These findings suggest that polymorphic CYP2D6 plays an important role in the pharmacology and toxicology of halamine. Known metabolites of halamine include halmorol.

[1]. Harmaline and its Derivatives Against the Infectious Multi-Drug Resistant Escherichia coli. Med Chem. 2017;13(5):465-476.
[2]. Peganum harmala L. seed extract attenuates anxiety and depression in rats by reducing neuroinflammation and restoring the BDNF/TrkB signaling pathway and monoamines after exposure to chronic unpredictable mild stress. Metab Brain Dis . 2024 Aug 22. doi: 10.1007/s11011-024-01416-6.
[3]. Comparison of Regression Methods to Predict the First Spike Latency in Response to an External Stimulus in Intracellular Recordings for Cerebellar Cells. Stud Health Technol Inform . 2024 Aug 22:316:796-800.
[4]. Cytotoxicity of alkaloids isolated from Peganum harmala seeds on HCT116 human colon cancer cells. Mol Biol Rep. 2024 Jun 13;51(1):732. doi: 10.1007/s11033-024-09655-7. https://pubmed.ncbi.nlm.nih.gov/38872006/

Harmaline is a Halman alkaloid in which the Halman skeleton is substituted with a methoxy group at the C-7 position and the 3,4 bond is reduced. It has a nocturnal effect. It is derived from the hydride of Halman. Harmaline has been reported in Passionflower, Daphnia magna, and other organisms with relevant data. It is a β-carboline alkaloid isolated from the seeds of Elaeagnus pungens. Mechanism of Action: Three psychoactive components extracted from Elaeagnus pungens seeds—Harmaline, Harmaline, and halmorol—all exhibited vasodilatory activity in isolated rat thoracic aortic specimens pre-constricted with phenylephrine or potassium chloride, with the vasodilatory potency order being: Harmaline > Harmaline > halmorol. The vasodilatory effects of Harmaline and halmorol (but not halmorol) can be ablated by endothelial removal or by pretreatment with the nitric oxide synthase inhibitor Nω-nitro-L-arginine methyl ester. In cultured rat aortic endothelial cells, halamine and halmalin (but not halmalol) increased NO release, and this process was dependent on the presence of extracellular Ca2+. In endothelial-desorbed vascular preparations, pretreatment with halamine, halmalin, or halmalol (3–30 μM) inhibited phenylephrine-induced contraction in a non-competitive manner. Receptor binding assays showed that all three compounds interacted with cardiac α1-adrenergic receptors with similar affinity (Ki values approximately 31–36 μM), but only halamine weakly interacted with the cardiac 1,4-dihydropyridine binding site of L-type Ca2+ channels (Ki value 408 μM). Therefore, the current results suggest that the vasodilatory effects of halamine and halmalin are attributed to their action on endothelial cell-mediated NO release and their action on vascular smooth muscle to inhibit contraction induced by receptor-coupled and voltage-dependent Ca2+ channel activation. The vasodilatory effect of halmalol was endothelial cell-independent.

---

Therapeutic Use
/EXPL THER/ Oxidative modification of low-density lipoprotein (LDL) particles is closely related to the development of atherosclerosis. Antioxidants that can prevent LDL oxidation may help alleviate atherosclerosis. We investigated the protective effects of P. harmala extract (P extract) and two major alkaloids (harmin and harine) from its seeds against copper sulfate-induced LDL oxidation. The extract (P-extract) and its compounds showed inhibitory effects by determining the production of malondialdehyde (MDA) and conjugated dienes, as well as the hysteresis time. Furthermore, harmin and harine reduced the rate of vitamin E consumption and exhibited significant free radical scavenging capacity (DPPH). However, harmin showed significantly higher antioxidant capacity than harine in scavenging or preventing free radicals and inhibiting the aggregation of the LDL protein moiety (apolipoprotein B). These results suggest that P. harmala compounds may be an important source for inhibiting copper-induced LDL oxidative modification.

C13H14N2O

214.26

214.11

C, 72.87; H, 6.59; N, 13.07; O, 7.47

304-21-2

304-21-2

3564

Orthorhombic bipyramidal prisms, tablets from methanol, rhombic octahedra from ethanol

1.3±0.1 g/cm3

426.4±45.0 °C at 760 mmHg

232-234 °C(lit.)

211.7±28.7 °C

0.0±1.0 mmHg at 25°C

1.647

0.66

1

2

1

16

302

0

CC1=NCCC2=C1NC3=C2C=CC(=C3)OC

RERZNCLIYCABFS-UHFFFAOYSA-N

InChI=1S/C13H14N2O/c1-8-13-11(5-6-14-8)10-4-3-9(16-2)7-12(10)15-13/h3-4,7,15H,5-6H2,1-2H3

7-methoxy-1-methyl-4,9-dihydro-3H-pyrido[3,4-b]indole

Harmaline; 304-21-2; Dihydroharmine; Harmidine; Armalin; 3,4-Dihydroharmine; Harmalol methyl ether; O-Methylharmalol;

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

---

Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

View More

Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

---

Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

View More

Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

---

Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

---

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