Methanol Reserpine

The first three extracts are combined and washed with 60 parts by volume of 2 N sodium carbonate solution and then with 60 parts by volume of distilled water. These washing solutions are saved and used for the yvashing of the 4th and final ethylene chloride extract. The combined ethylene chloride extracts are dried over sodium sulfate, filtered and evaporated in vacuo to a constant weight of a tan, frothy solid. One part by weight of this residue is dissolved in 1.5 parts by volume of warm methanol and the solution cooled to 5°C for 18 hours, whereby crystallization of a mixture containing principally reserpine sets in. After filtering this mixture and yvashing it with cool methanol, the filtrate is freed of Solvent in vacuo. 

Reserpine is extracted from powdered tablets with water saturated with ethyl acetate, after addition of a solution of propiophenone in ethyl acetate saturated with water (as internal standard). After centrifugation, the ethyl acetate layer is analysed by HPLC on a 10 pm Lichrosorb RP-8 column using methanol/0.05M sodium phosphate monobasic mixture (1 1) as the mobile phase. The flow rate is adjusted to 2 ml/minute. Detection is carried out under UV at 25U run. (Fig. 9). 

Reserpine is isolated from its plant producers by using a nonaqueous solvent process, using, for example, boiling methanol extraction of the African root Rauwolfia vomitoria. Naturally, these extractions are carried out under countercurrent methods. The methanol extract is concentrated and acidified with 15% acetic acid and then treated with petroleum naphtha to remove impurities. Extraction is made using ethylene dichloride. The solvent is neutralized with dilute sodium carbonate, evaporated to drive off the ethylene dichloride, and further evaporated to crystallize the crude reserpine crystals that are then crystallized. 

A specific cleavage reaction of NH2-terminal tyrosyl dipeptides occurred upon their treatment with DIB in methanolic potassium hydroxide. Tyrosine itself was converted into 4-hydroxybenzyl cyanide (52%) under these conditions [69], A related cleavage of tryptophan and several derivatives of it, including dipeptides, led to the formation of 3-methoxyindole [70]. Reserpine and also 2,3-dimethylindole reacted in the same way in the presence of alcohols, affording 3-alkoxyindolenines... 

For laboratory purposes, methanol is the usual solvent, and procedures based on its use have been described in detail (21, 39, 81). Of great practical usefulness for the separation of the weakly basic fraction is the solubility of certain alkaloidal acetates in chloroform, e.g., reserpine, ajmalicine, and aricine, whereas other acetates are insoluble in this solvent, e.g., ajmaline, yohimbine, and a-yohimbine. Since the anhyd-ronium alkaloids are extremely strong bases, they can only be extracted into an organic solvent in their tertiary base form at pH 11. For industrial purposes, the best process extracts water-moistened Rauwolfia root with hydrocarbons such as benzene, toluene, or xylene. In this procedure, only the weak bases are extracted. No complicated separation processes are involved, and reserpine is obtained in high yields (122). 

When 7-aeyl-7 -reserpine derivatives (XV) are refluxed in dilute methanol with a few drops of acetic acid, conversion into the corresponding oxindole (XIX) with concomitant formation of the five-membered spiro-ring C takes place. Under these conditions, also, carbon... 

Reserpine Bulk HPLC treated) Silica ide—methanol (1.5. 100) Methylene chloride permanganate 254 nm 1LC [1.8] HPLC [8.611. [ 1... 

Because of the difficulties associated with the Delphinium classification, the diterpene alkaloid content was assessed as an aid to determining the chemical taxonomic diversity of the toxic larkspur species [59]. Plant samples were collected from 18 different locations in five western states in the US. The crude methanolic extracts were analyzed for diterpene alkaloids using FI-ESI-MS. The data from the individual ESI mass spectra were statistically analyzed using canonical discriminant analysis and analysis of variance. In brief, the sample (100 mg) was extracted by mechanical shaking at room temperature with methanol (5mL) for 16h. Reserpine (500 p.g) was added as an internal reference standard and the sample mixed for 5 min and then centrifuged. An aliquot (30 jxL) of the supernatant was then diluted with of 1 1 methanol/1 % acetic acid (1.0 mL) and an aliquot (20 jjlL)... 

Settimj et al.6 described a gas chromatographic method for the estimation of reserpine and rescinnamine involving alkaline hydrolysis of the alkaloids and subsequent esterification of the acids formed by means of diazomethane. Reserpine gave quantitatively 3,4,5-trimethoxyben-zoic acid methylester, whereas the trars-3,4,5-trimethoxycinnamic acid methylester, which should be expected for rescinnamine, was partly isomerized to the cis-trimethoxycinnamic acid methylester or formed an adduct with a molecule of methanol, yielding 3-methoxy-3-(3,4,5-tr1-methoxyphenyl) propionic acid methylester. 

Sams reported the determination of reserpine in plasma by ion-pair chromatography. To obtain high specifity and to increase the sensitivity reserpine - after extraction from plasma with benzene - was oxidized to 3-dehydroreserpine with vanadium pentoxide in concentrated phosphoric acid, to which 9 ml methanol were added. The fluorescent compound was subsequently analyzed on an octadecyl column with methanol - 0.001 M heptanesulfonate in water (65 35) as mobile phase. Reserpine and rescinnamine could not be distinguished by this method. 

In the total synthesis of reserpine. Woodward and collaborators (10) have reported that the quaternary iminium salt 18 was reduced with aqueous methanol ic sodium borohydride to methyl 0-acetyl isoreserpate ( S). This is the anticipated product whether the stereochemical sense of the reaction is subject to steric or thermodynamic control as pointed out by Woodward. It is also the expected one on the basis of stereoelectronic control. 

Figure 30. In-phase fundamental-harmonic a.c. polarographic peak current vs. concentration profiles for reserpine and methyltestosterone. System (A) CH3CN 2,4,6,8,10 x 10"5 M reserpine at DME, 2s drop life, 0.01 TEAPFB in CH3CN, (A) DMF 2,4,6,8,10 x lO" M reserpine at DME, 2s drop life, 0.01 M TEAPFB in DMT, (A) Methanol-water 0.01 M TBAOH 2,4,6,8,10 x lO" M reserpine at DME, 2s drop life, 0.01 M TBAOH in 60% methanol-40% water, (B) CH3CN 2,4,6,8,T0 x 10-5 M methyl-testosterone at DME, 2s drop life. 0.01 M TEAPFB in CH3CN, (B) DMF 2,4,6,8,10 x lO" M methyl testosterone at DME, 2s drop life, 0.01 M TEAPFB in DME, (B) Aqueous Base 2,4,6,8,10 x 10" methyl testosterone at DME,...

Fig. 2 MS signal (black) for a 10 )iM reserpine solution (50 % methanol in water with 0.1% acetic acid) as a function of the applied voltage/ electrospray operating regime (Adapted with permission from Ref. [5], copyright American Chemical Society), (b) MS signal (black) and spray current (cyan) for a 5 pM leucine enkephalin solution (33 % acetonitrile in water with 1 % trifluoroacetic acid) as a function of applied voltage (Adapted with permission from Ref. [6], copyright American Chemical Society)...

Reserpine therapeutically the most important Rauwolfia alkaloid. M, 608.9, m.p. 265 C, (oj -123° (CHQj). Hydrolysis of R. with alcoholic KOH produces reserpinic add (structurally similar to yohimbine), trihydroxybenzoic add and methanol. R. is found widely in Ae genus Rauwolfia and is responsible for the sedative properties of these plants. After a latent period, R. causes long lasting sedation, with decrease of blood pressure and decreased pulse rate. It is used as a powerful neuroleptic drug in psychiatry. 

In a flow injection analysis (FIA) SIM scan using a TWF for isolation step 2, 1-pL injections of reserpine, miz 609, were made into a 200-pL/min flow rate consisting of a solvent system of 1 % acetic acid in a 50 50 methanol water. A detection limit of 8 fmol (5 pg) with a signal-to-noise ratio of 7 was measured. 

The route employed to prepare indanone 51 involved the cycloaddition-hydrolysis-aldol sequence shown in Scheme 3.9. Accordingly, condensation of cyclopentenone 52 with ynamine 53 (84) afforded the bicyclic enamine 54 which was converted to indanones 51 and 55 by hydrolytic cyclobutane ring opening followed by intramolecular aldol condensation. Interestingly, treatment of 54 with aqueous formic acid yielded indanone 51 which has stereochemistry complementary to that at C(15) and C(20) in reserpine. In contrast, hydrolysis of this substance with aqueous hydrochloric acid afforded the trans-fused indanone 55. Subsequent to this work, the Ficini group found that esterification of 51 followed by photochemically induced addition of methanol afforded adduct 56 which has four of the reserpine stereocenters in place (23). While no further work on this problem has been reported, these preliminary investigations demonstrate a novel use of [2 -h 2] photocycloaddition chemistry in potential approaches to yohimbane alkaloid synthesis. 

---