Reference: K Genest. A Direct Densitometric method on Thin-Layer Plates for the Determination of Lysergic Acid Amide, Isolysergic Acid Amide and Clavine Alkaloids in Morning Glory Seeds.
IntroductionThe discovery by Hofmann
1 that the ancient magic drugs "Ololiuqui" (
Rivea corymbosa) and "Badoh Negro" (
Ipomoea violacea) contain psychotomimetic substances prompted other workers to investigate ornamental varieties of morning glories which belong to the genus
Ipomoea and
Convolvulus. Taber et al.
2 found lysergic acid and clavine alkaloids in the seeds, embryo, leaf and stem in several horticultural varieties of morning glories. Similar results referring to the total alkaloidal content and the chromatographic identification of several ergoline alkaloids were published by Gröger
3, Beyerman et al
4 and Alexander
5. Pharmacologically the most important of the alkaloids in morning glory seeds are lysergic acid amide (LAA) and its epimer isolysergic acid amide (isoLAA). Solms
6 found that LAA is a psychotomimetic with strong sedative action. Information in the literature on the quantitative content of the individual alkaloids is limited to Hofmann's report on two large samples of
Rivea corymbosa and
Ipomoea violacea and on the analysis of Taber et al. of one sample each of Ololiuqui and Pearly Gates. The latter authors gave only relative percentage values of several alkaloidal components after elution from thin-layer chromatograms.
It is the purpose of this paper to present a method for the determination of lysergic acid amide, isolysergic acid amide and clavine alkaloids in morning glory seeds by direct densitometry on thin layer plates.
Materials and methodsThin-layer chromatography (TLC) - Desaga equipment according to Stahl on 20x20 cm plates and 250 µ layers was used. The standard alkaloids were dissolved in ethanol or chloroform to contain 1 µg per µL. Morning glory extracts were prepared as described under "Total alkaloids". For qualitative TLC about 5 µg total alkaloids per spot were applied. The layer material used was MN-Silica Gel-HR' and Aluminium Oxide GF. After spotting the thin-layer plates were chromatographed until the solvent front reached 15 cm. The following systems were used:
(1) Layer: Aluminium Oxide GF; Solvent: chloroform:EtOH (96:4)
3(2) Layer: MN-Silica Gel-HR'; Solvent: acetone:piperidine (9:1)
(3) Layer: MN-Silica Gel-HR'; Solvent: acetone:ethylpiperidine (9:1)
(4) Layer: Aluminium Oxide GF; Solvent: acetone:ethylpiperidine (9:1)
(5) Layer: MN-Silica Gel-HR'; Solvent: acetone:EtOAc:DMF (5:5:1)
3After chromatography the plates were examined under 3660 Å and 2537 Å UV-light and sprayed with a solution of p-dimethylaminobenzaldehyde (DMBA).
Total alkaloids - Powdered morning glory seeds (1.0 g) were defatted by refluxing with petroleum ether (bp 30-60°) transferred into a Erlenmeyer flask (250 mL) and wetted with 5 mL of 10% aluminium hydroxide. Fifty mL of peroxide-free ether was added and the mixture was shaken mechanically for 30 min. The ether layer was then extracted with 5, 2.5 and 2 mL of 0.1 N sulfuric acid. The acid extracts were collected in a sparatory funnel. The extraction of the seed-ammonium hydroxide mixture was repeated with fresh portions of ether until judged to be complete by the absence of a grey-blue fluorescence in the acidic layer when irradiated with 3660 Å UV light. The combined acid extracts were made alkaline with ammonia and the solution was extracted with 20, 10 and three times 5 mL portions of chloroform. The chloroform extracts were collected in a 50 mL volumetric flask and filled up to the mark with chloroform. For the determination of total alkaloids two 5 mL aliquots were removed and evaporated to dryness in a stream of nitrogen. One mL of 0.1 N sulfuric acid and 2 mL of Van Urk reagent were added to the residue. After 30 min the absorbance at 550 mµ minus the baseline absorbance at 720 mµ were measured in a Beckman DK-2 spectrophotometer.
Van Urk reagent: 0.125 g DMBA was dissolved in 65% sulfuric acid (100 mL) to which 0.1 mL 5% ferric chloride was added.
[...]
Results[...]
Alkaloidal content of morning glory seeds
Horticultural variety | Botanical name | TA (%) | Origin | Remarks |
[/red]
Flying Saucers | Ipomoea violacea L. | 0.057a | California | |
Heavenly Blue | Ipomoea violacea L. | 0.023 | Rhodesia | |
Major Tall | Ipomoea violacea L. and Ipomoea purpurea L. Roth | 0.026 | likely California | seed mixture |
Pearly Gates | Ipomoea violacea L | 0.032 | California | |
Wedding Bells | Ipomoea violacea L. | 0.030 | California | |
Ololiuqui | Rivea corymbosa L | 0.045 | Cuba | |
Scarlet O'Hara | Ipomoea Nil L Roth | 0.001 | California | |
Darling | Ipomoea Nil L Roth | 0.001 | California | |
Royal Marine | Ipomoea Nil L Roth | 0.001 | England | |
Double Rose Marie | Ipomoea Nil L Roth | 0.000 | California | SFb |
Tall Mixed | Ipomoea purpurea L Roth | 0.000 | Rhodesia | SF |
Candy Pink | Ipomoea Nil L Roth | 0.000 | California | |
Crimson Rambler | Ipomoea purpurea L Roth | 0.000 | The Netherlands | |
Everblooming | Convolvulus tricolor L | 0.000 | likely Denmark | |
Moon Flower | Ipomoea alba L | 0.000 | Likely California | |
Fine Mix | Convolvulus tricolor L | 0.000 | Denmark | SF |
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[a] Calculated for MW 262
Slight fluorescence in acidic extract.
[...]
[1] A Hofmann. Planta Med 9 (1961) 354
[2] WA Taber, LC Vining, RA Heacock. Phytochemistry 2 (1963) 65
[3] D Gröger. Flora (Jena) 153 (1963) 373
[4] HC Beyerman, A Van de Linde, GJ Henning. Chem Weekblad 59 (1963) 508
[5] TG Alexander, personal communications
[6] H Solms. J Clin Explt Psychopath Quart Rev Psychiat Neurol 17 (1956) 429