Physostigmine analysis

Analysis of ACh in blood and CSF poses two obstacles—low concentrations (<20 pmol/ml) and very fast hydrolysis by esterase (Tsai, 2000). An acetylcholinesterase inhibitor such as physostigmine maybe added to artificial CSF used for the collection of microdialysates to help overcome this problem (Kato et al., 1996). 

Frolich and colleagues (1998) analyzed ACh in human CSF by different methods, which included thermospray/mass spectroscopy, HPLC/mass spectroscopy, HPLC-EC Pt electrode and gas chromatogra-phy/mass spectroscopy (GC/MS). An SPE extraction was used for cleanup and concentration. Samples were run with and without the IMER to rule out any interference by physostigmine, a cholinesterase inhibitor, in the HPLC-EC assay. HPLC-EC and GC-MS gave data correlations with similar sensitivities, but the HPLC-EC values were 39% lower. Analysis using thermospray/mass spectroscopy and HPLC/ mass spectroscopy did not provide adequate sensitivity and the data obtained were inconsistent. 

Lawrence, G.D. Yatim, N. Extraction of physostigmine from biologic fluids and analysis by liquid chromatography with electrochemical detection. J. Pharmacol. Meth. 1990, 24, 137-143. 

Physostigmine salicylate in 0.5 % aqueous solution was determined by Teare and Borst by freeze-drying of samples of 0.2 ml, and conversion of the compound into its trimethyl silyl derivative by dissolving the residue in 5 yl of dry pyridine and 10 yl of N,0-bis(trimethyl -silyl)acetamide. The solution was allowed to stand for 1 h before injecting 1.2 yl into the gas chromatograph. A 3.8 % SE-30 on Diatoport S column and a temperature of 145°C was used for the analysis. The physostigmine THS and the salicylic acid TMS derivatives were eluted separately. Routine analysis over several days gave a precision of 11.5 %. 

Tymes07 reported a collaborative study of the analysis of physostigmine in pharmaceutical preparations using octadecyl stationary phases with the mobile phase acetonitrile - 0.05 M... 

Fig. 8.7. HPLC analysis pilocarpine, physostigmine, its degradation products and preservatives in pharmaceutical preparations4 ...

Sporidesmins, the series of sulfur-containing physostigmine-like metabolites, were isolated from Sporidesmium bakeri A (57), B (58) and C (59), a fungus that causes facial eczema and liver damage in farm animals, e.g. New Zealand sheeps [108-110]. Later studies identified sporidesmins D (60) [111], E (61) [112], F (62) [111], G (63) [113,114], H (64) [115]) and J 65) [115] from P. chartarum. Protonated [M+H]+ and deprotonated [M-H] ions were observed in positive and negative ion ESI modes, respectively [116]. In a further paper, complexation of sporidesmin A, with metals, was used for its analysis [117]. 

Arundinine (127), was obtained from the epigeal parts, and the strueture was established by spectroscopic methods as well as by X-ray analysis. The moleeule is constituted from the union of two tryptamine moieties, a physostigmine-type unit and another tryptamine unit. The two units are linked by an ether bridge from C(3a) of the former to the aromatic C(5 ) of the other unit (79). A number of related bisindoles have been subsequently isolated from the root extract of the same plant, viz., arundamine (128) (80,81), arundacine (129) (82), arundanine (130) (83), arundavine (131) (84), arundaphine (132) (85), and arundarine (133) (86). The structures of 128,131, and 132 have also been confirmed by X-ray analysis, while the NMR spectrum of arundacine (129) showed the existence of two equilibrating con-formers due to restricted rotation about the amide C-N bond. 

Figure 3.13 Analysis of physostigmine using the ESA Coulochem [PGEs, guard cell (positioned between column outlet and analytical cell) +0.8 V, Ej —0.2 V, E2 + 0.1 V vs Pd]. Column 150x4.6 (i.d.) mm Spherisorb S3W silica Eluent acetonitrile-methanol-aq. ammonium nitrate buffer (0.1 mol pH 9.5) (45 + 45 + 10) Flow-rate 0.75 mL min Injection 50 jiL methanol containing 1 ng physostigmine (1).

R. Whelpton, Analysis of plasma physostigmine concentrations by liquid chromatography, J. Chromatogr., 1983, 272, 216-220. 

S. Knapp, M.L. Wardlow and L.J. Thai, Sensitive analysis of plasma physostigmine levels using dual-cell electrochemistry in the redox mode, J. Chromatogr., 1990, 526, 97-107. 

In 1990, we reported [ 166] that CPT-11 inhibits acetylcholinesterase activity. In the paper, the rate of thiocholine (TCh) production from acetylthio-choline iodide (ATChl) by acetylcholinesterase was analyzed with Line-weaver-Burk plots (Figure 2.3). This kinetic analysis indicated that CPT-11 noncompetitively inhibited acetylcholinesterase. In this case, the apparent Michaelis constant (Km) and inhibition constant (Ki) ranged from 63 to 68 yUM and from 0.221 to 0.300 /iM, respectively (Table 2.3). The same analysis revealed that the Ki value of physostigmine (a potent acetylcholinesterase... 

Asymmetric intramolecular carbopalladation is an effective reaction for producing enantioenriched polycycles. Most examples are intramolecular Heck reactions. One seminal example from natural product synthesis is the 5-exo carbopalladation of eneamide 201 to oxindole 202 (after acid treatment) from a total synthesis of ( )-physostigmine 203 (Scheme 31).The reaction occurs in 84% yield with 95% ee, which is remarkably efficient for the construction of a quaternary center. Reaction conditions that favor the neutral manifold of the Heck reaction are employed. Examination of the scope of the oxindole synthesis and mechanistic analysis have appeared. " Group selective reactions are also powerful reactions in carbopalladation asymmetric synthesis.From a synthesis of (+)-vemolepin 206, alkenyl triflate 204 is... 

Morales-Rios, M.S., Santos-Sanchez, N.F., and Joseph-Nathan, P. (2002) Efficient formal total synthesis of physostigmine and physovenine conformational analysis of key intermediates. J. Nat. Prod., 65, 136-141. 

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