Activities of extract and constituents of Banisteriopsis caapi relevant to parkinsonism

This early cell-based study (2003) found that B. caapi stem extract may be useful in the treatment of Parkinson’s disease. This study was the first to find it stimulated dopamine release.

Abstract

“Dopamine deficiency is characteristic of Parkinson’s disease (PD) and treatments aim at elevating levels by administration of its precursor l-dihydroxyphenylalanine (l-DOPA), or inhibiting monoamine oxidases (MAOs), thus preventing its breakdown. Reports of improvements in PD patients treated with Banisteriopsis caapi extracts stimulated investigation of B. caapi stem extract and its two ingredients, harmine and harmaline for these activities. Tests for MAO inhibition using liver homogenate showed that extract and harmaline showed a concentration-dependent inhibition of MAO A (IC50 1.24 μg/ml and IC50 4.54 nM, respectively) but had little effect on MAO B activity. The extract at 2.5 mg/ml caused a highly significant increase in release of [3H]dopamine from rat striatal slices, as did 200 μM harmine and 6 μM harmaline. In both these experiments, the amount of harmine present could not account for the total activity of the extract. The ability of harmine and harmaline to stimulate dopamine release is a novel finding. These results give some basis to the reputed usefulness of B. caapi stem extract in the treatment of PD.”

Authors: M. J. Schwarz, P. J. Houghton, S. Rose, P. Jenner & A. D. Lees

Summary

Abstract

Parkinson’s disease is treated by elevating dopamine levels or inhibiting monoamine oxidases. Banisteriopsis caapi stem extract contains harmine and harmaline.

Harmaline and extract of B. caapi showed concentration-dependent inhibition of MAO A and little effect on MAO B activity. The extract and harmine increased the release of [3H]dopamine from rat striatal slices.

Banisteriopsis caapi has anti-Parkinson’s disease properties.

1. Introduction

Ayahuasca is a hallucinogenic drink made from Banisteriopsis caapi, which is also known as ‘hoasca’, ‘yage’ and ‘caapi’. Its hallucinatory properties have been known to many indigenous peoples for centuries.

Harmine, then called banisterine, was used in animals and patients with symptoms similar to Parkinson’s disease. Recently, it has been shown to be beneficial in treating patients with Parkinson’s disease.

2. Methods

2.1. Plant material and extraction

A dried stem of B. caapi was obtained from Dr. Marcos Serrano-Duenas, Neurology Service, Carlos Andrade Marin Hospital Quito, and was extracted with 500 ml distilled water for 1 h.

2.2. Thin-layer chromatographic analysis of the extract

The freeze-dried extract of the root was dissolved in methanol and applied to a TLC plate. The amount of harmine and harmaline present in the extract was determined by reflectance scanning using a Shimadzu TLC scanner at 500 nm.

2.3. Material for biological tests

Male Wistar rats (190 ± 10 g) were killed by stunning anddecapitation. The abdomen was opened and a liver samplewas dissected. The liver was homogenised in 20 volumes(5% wt/vol) of ice-cold sodium phosphate buffered 0.9%saline (PBS; 0.1 M, pH 7.4). Homogenates were stored at
70°C until assayed for MAO activity. On the day of theassay, samples were allowed to defrost on ice and weremixed by vortexing. Liver samples were thawed and thenfurther diluted 1:10 with ice-cold PBS and vortexed toproduce a final liver homogenate of 1:200 volumes (0.5%wt/vol).

2.4. MAO inhibition assay

2.4.1. In vitro assay

MAO activity was assessed by adding liver homogenate to a solution of [14C]5-HT or [14C]phenylethylamine (PEA), and incubating for 30 min at 37 °C. The assay was terminated by adding 2 M citric acid.

2.4.2. Extraction of deaminated metabolites

MAO A and B activities were determined by extracting the deaminated metabolites from 5-HT and PEA, respectively, and determining the radioactivity in the extracts by liquid scintillation spectroscopy.

2.4.3. Drugs

The radioactive substrates 5-hydroxytryptamine-[sidechain-2-14C]creatinesulphate([14C]5-HT)andb-phenylethyl-amine-[side chain-ethyl-1-14C]hydrochloride ([14C]PEA)were obtained from the Radiochemical Centre, Amersham.All standard compounds were obtained from Sigma. Allother reagents were standard laboratory reagents of analyt-ical grade wherever possible.

2.4.4. Data and statistical analysis

MAO activity was calculated from radioactive dissolutions per minute to moles product formed per milligram tissue per 30 min using Prism 3 Software.

2.5. In vitro release of [³H]dopamine ([³H]DA) from ratstriatal slice

Male Wistar rats were killed by cervical dislocation and decapitation, their brains removed and immediately cooled on ice, and their striata were dissected and cut coronally into slices. The slices were incubated in Krebs containing [3H]DA for 30 min at 37 °C, and then superfused with Krebs throughout the experimental period.

2.6. The effect of B. caapi extract, harmine and harmalineon [3H]DA release from rat striatal slices

After superfusion with Krebs, B. caapi extract, harmine or harmaline was incorporated into the Krebs for 60 min to determine dopamine efflux. Data were analysed using two-way ANOVA.

3. Results

3.1. MAO inhibition

Clorgyline (10^-15to 10^-6M) produced a concentra-tion-related inhibition of MAO-A activity with an IC50of0.66 nM. Clorgyline produced some inhibition of MAO B with maximal inhibition of 25% at 10-6M, and an IC50of greater than 10-6M(Fig. 2a, Table 1). It is appropriate to quote an IC50value since clorgyline is known to produce a reversible inhibition at these concentrations.Selegiline (3^-10-9to 10^-6M) produced a concen-trated-related inhibition of MAO B activity with an IC₅₀of5.0 nM. Selegiline produced some inhibition of MAO-A at higher concentrations, with a maximal inhibition of 48% at10^-6M, and an IC₅₀of greater than 10^-6M(Fig. 2b,Table 1).

B. caapi extract also produced a concentration-related inhibition of MAO-A activity with an IC₅₀of 1.24^-10
6g/ml.B. caapi extract had little effect on MAO B activity, producing a maximal inhibition of 15% at between 10^-5 and 10^-6g/ml (Fig. 2c, Table 1). Harmine (3^-10-9to 10^-6M) produced a concentration-related inhibition of MAO-A activity with an IC₅₀of4.54^-10-9M. Harmine had little effect on MAO Bactivity, with maximal inhibition of 20% at 10^-6M(Fig.2d, Table 1).

3.2. Release of [³H]DA from rat striatal slices

The results of these experiments show that the B. caapi extract (2.5 mg/ml) and harmine (200 mM) increased [3H]DA release, but harmaline (58.3 mM) increased [3H]DA release more than harmine.

4. Discussion

4.1. Content of harmine in extract

The value of 19.2 mg harmine per gram lyophilized extract is comparable with values obtained in a previous study on several samples where the mean value was found to be 23.8 mg/g(McKenna et al., 1984).

4.2. MAO inhibition

The B. caapi extract and harmine inhibit MAO-A rather than MAO-B, i.e., they behave like clorgyline rather than selegiline. The activity of the extract is therefore due primarily to harmine, and may be due to a synergistic effect or the existence of a much more active compound.

4.3. Release of DA from striatal slices

B. caapi extract, harmine and harmaline increased [3H]DA release from rat striatal slices, but harmine was more potent than harmine. The ratio of harmine to harmaline in the extract is about 5:1, and the 2.5-mg/ml solution that gave a significant effect has concentrations of 20.1 and 5.83 mM, respectively, the middle concentration of the alkaloids used.

4.4. Overall conclusions

The ability of the extract to inhibit MAO A and stimulate release of DA from striatal cells would increase DA levels in the CNS and alleviate symptoms of Parkinson’s disease if the alkaloids were able to cross into the bloodstream from the intestine.