"Soxhlet exraction procedure

Still frozen A. rigidula was packed into Whatman single thickness cellulose extraction thimbles (ca 50g/thimble, 200 g total) and extracted continuously for 24 h with MeOH. The MeOH was removed and replaced with CHCl₃ and the extraction continued for an additional 24 h. A pilot study established that the extracts could be safely concentrated by rotary evaporation (water aspirator). The MeOH extract was concentrated and the residue dissolved in 100 ml of CHCl₃. This solution was extracted X 3 with 50 ml portions of 10% aq. HCl . The acid fractions were combined, the pH adjusted to ca 10.3 by addition of aq. NaOH, and the resulting solution was first extracted X 3 with 50 ml portions of CHCl₃, followed by extraction X 3 with 50 ml portions of EtOAc. These organic extracts were combined, dried with MgSO₄, filtered, concentrated under vacuum, and stored under argon prior to analysis by GC-MS. The CHCl3 fraction from the Soxhlet extraction was handled in an identical fashion to the MeOH extract with the exception that it was not first concentrated prior to aq. acid extraction. The MeOH and CHCl₃ extracts from the Soxhlet extraction were not combined.

Acid extraction of plant material

Acacia rigidula, 100 g, was placed into a 1 l Erlenmeyer flask and mixed with 500 ml of 10% aq. acid (HCl and HOAc were both used). The suspension was stirred under argon at 60° overnight. The darkened suspension was filtered through glass wool covered with a 2 cm bed of sand. The filtrate was extracted X 2 with 100 mL portions of EtOAc followed by X 3 extraction with 100 ml portions of CHCl₃. The pH of the filtrate was adjusted to ca 10 by addition of NaOH pellets. This solution was extracted X 3 with 150 ml portions of CHCl₃, followed by extraction X 3 with 150 ml portions of EtOAc. The organic extracts were combined, dried over MgSO₄, filtered, concentrated under vacuum and stored under argon prior to analysis by GC-MS."

[The authors analysis by GC-MS revealed the following alkaloids in Acacia rigidula:

2-Cyclohexylethylamine; N-2-Cyclohexylethyl-N-methylamine; Phenethylamine; N-Methylphenethylamine; N,N-Dimethylphenethylamine; Amphetamine; Methamphetamine; N,N-Dimethyl-a -methylphenethylamine; p-Hydroxyamphetamine; p-Methoxyamphetamine; Tyramine; N-Methyltyramine; Hordenine (anhaline); Dopamine; N-Methyldopamine; N,N-Dimethyldopamine; 3-Methoxytyramine; N-Methyl-3-methoxytyramine; 3-Hydroxy-4-methoxyphenethylamine; N-Methyl-3-hydroxy-4-methoxyphenethylamine; 3,4-Dimethoxyphenethylamine, N-Methyl-3,4-dimethoxyphenethylamine; 3,4,5-Trihydroxyphenethylamine; N-Methyl-3,4,5-Trihydroxyphenethylamine; Mescaline; N-Methylmescaline; Trichocereine; 3,5-Dimethoxytyramine; 3,4-Dimethoxy-5-hydroxy-b -phenethylamine; b -Methoxy-3,4-dihydroxy-5-methoxy-b -phenethylamine; 3,4-Dimethoxy-a -methyl-5-hydroxy-b -phenethylamine; Tryptamine; N-Methyltryptamine; N,N-Dimethyltryptamine; Nicotine; Nornicotine; Anhalamine; Anhaladine (N-methylanhalamine); Anhalonidine; Peyophorine; Pipecolamide; p-Hydroxypipecolamide; 1,4-Benzezediamine and 4-Methyl-2-pyridinamine.