Chiral HPLC analysis

Absolute configurations of the isoxazolidines obtained in the nitrone cydoaddition reactions described in Schemes 7.21 and 7.22 were determined to be 3S,41 ,5S structure by comparison of the optical rotations as well as retention times in a chiral HPLC analysis with those of the authentic samples. Selection of the si face at C/ position of 3-crotonoyl-2-oxazolidinone in nitrone cydoadditions was the same as that observed in the Diels-Alder reactions of cyclopentadiene with 3-croto-noyl-2-oxazolidinone in the presence of the J ,J -DBF0X/Ph-Ni(C104)2-3H20 complex (Scheme 7.7), and this indicates that the s-cis conformation of the dipolaro-phile has participated in the reaction. 

After removal of the solvent, the residue was eluted through a short silica gel column to remove the catalyst (elution with hexane ethyl acetate = 1 2). The eluent was concentrated in vacuo to give the product 2 (99 % yield) and the diastereoselectivity was determined by HPLC analysis (99 %). The enantios-electivity of the product was determined by lH NMR analysis with chiral shift reagent (+)-Eu(dppm) in CDCI3 and by chiral HPLC analysis (Chir-alcel-OD). 

LT048 Moinuddin, S. G., S. Hishiyama, M. H. Cho, L. B. Davin, and N. G. Lewis. Synthesis and chiral HPLC analysis of the dibenzyltetrahydrofuran lignans, larreatricins, 8 -epi-larreatricins, 3,3 -didemethoxyverrucosins and meso-3,3 -didemethoxynectandrin B in the creosote bush (Larrea tridentata) evidence for regiospecific control of coupling. Org Biomol Chem 2003 1(13) 2307-2313. 

Batch conditions are used with a ratio enzyme/substrate of 2 (weight), during 1 to 24 hours at 25 °C. Progress of the reaction is measured by proton NMR with an internal standard, and enantiomeric excess is obtained through chiral HPLC analysis of the product. 

To a solution of racemic 4-phenyl-3-butyn-2-ol (14.6 mg, 0.10 mmol) in chlorobenzene (1.0 mL) was added bicyclohexyl (16.6 mg, 0.10 mmol) as internal standard for GC analysis. An aliquot (50 iL) of this solution was taken out of the flask as a zero point and passed through silica gel (8 2hexane-EtOAc eluent) before analysis by GC. The Ru complex 1 (2.0 mg, 2.0 pmol) was added to the remaining solution and the mixture was stirred in air at room temperature for 17 h with irradiation by fluorescent light (100 V, 25 W). To remove the complex the mixture was filtered through silica gel (8 2 hexane-EtOAc eluent) and analyzed by GC (65.3% conversion). The filtrate was then concentrated and chromatographed on silica gel (8 2hexane-EtOAc). Chiral HPLC analysis (Chiralcel OD-H, 15 1 hexane-i-PrOH) of the purified alcohol showed >99.5 % ee. 

Determine the enantiomeric excess by chiral HPLC analysis Chiracel OD, heptane isopropanol, 90 10 flow 0.6 ml/min retention time 12.6 (major) and 15.3 (minor enantiomer) min. 

S)-Tetrahydro-l -methyl-3,3-diphenyl-1 H,3H-pyrrolo[1,2-c][1,3,2]oxazabo-role was prepared from (S)-proline in two steps according to the literature procedure4 and purified by bulb-to-bulb distillation (170°C, 0.2 mm). The enantiomeric purity of the intermediate, (S)-a,a-diphenyl-2-pyrrolidinemethanol, was determined to be 99% ee by chiral HPLC analysis of its corresponding N-p-toluenesulfonamide derivative (DIACEL Chiralcel OD column hexane/ethanol, 92/8 1.0 mLVmin Rt(S) 8.6 min Rt(R) 12.8 min). The checkers used the crystalline p-methyloxazaborolidineborane complex (1.84 g, 6.34 mmol) as the catalyst. 

Enantiomeric excess is determined by chiral HPLC analysis (DIACEL Chiralcel OJ column hexane/ethanol, 70/30 1.0 mLVmin Rt S-isomer (8.8 min) Rt R-isomer (17.9 min). 

Analysis of Reagent Purity by H NMR and X-ray analyses of its (/ )-a-methoxy-a-(trifluoromethyl)phenylacetic acid [(/ )-MTPA] derivative chiral HPLC analysis supercritical fluid chromatography (SFC). ... 

Isolated yields. Enantiomeric excesses of the allylated product were determined by chiral HPLC analysis. Absolute stereochemistry determined to be (JR) by CD spectrum of 2-methyl-2-propyl-4-methoxyindanone. 

Yields are for isolated and purified materials. ) ee s were determined by chiral HPLC analysis using a Chiralcel OD column. Enantiomeric ratios above 10 1 are rounded off to the nearest integer. The absolute stereochemistry of the product was established by hydrolysis to the known carboxylic acid and comparison of the sign of its rotation. 

The equilibrium of the enzyme acylation reaction can be shifted towards the synthesis of the amide by precipitation of the acylated product formed (Fig. 6). The racemic ethyl 3-amino-5-(trimethylsilyl)-4-pentynoate 3 is an insoluble liquid, whereas the (R)-phenylacetamide 10 is an insoluble solid. The racemic ethyl 3-amino-5-(trimethylsilyl)-4-pentynoate 3 was added to dilute hydrochloric acid. The pH of the reaction medium was then adjusted to 6. Phenylacetic acid (2 equiv.) was added and the pH of the medium was readjusted to 6. Soluble PGA (50 units/100 mg of racemic amine) was added, and the reaction was stirred at room temperature. After completion of the reaction, the pH of the reaction mixture was adjusted to 4. Filtration of the reaction mixture gave (R)-amide 10 in quantitative yield. Chiral HPLC analysis of this isolated amide showed the absence of (S)-amide. The pH of the filtrate was raised to 8, and the filtrate was extracted with ethyl acetate to obtain (S)-amine 11 (yield 90%) (Fig. 6). The chiral HPLC analysis indicated an R S ratio of 2 98. 

In deacylation, as the enzyme cleaved the phenylacyl group, phenylacetic acid was formed, which lowered the pH of the reaction medium. Base was added to maintain the starting pH. (Note Use of ammonium hydroxide led to the formation of desilylated byproducts desilylation was eliminated when bicarbonates were used.) This approach was not required in the acylation reaction. At pH above 7.5 the (R)-and (S)-amines are practically insoluble in water. Organic solvents were used to extract the free amines from the aqueous reaction medium at pH 8.0. p-Fluoro-benzoyl, 1-naphthoyl, and phenylacetyl derivatives of the racemic amine were prepared and their behavior on the chiral HPLC column was studied. Based on ease of preparation and HPLC analysis, the 1-naphthoyl derivatives (Fig. 7) were preferred. Reversed phase HPLC analysis on a Vydac-C18 analytical column used a gradient of acetonitrile (0.1% triethylamine) in water (0.05% phosphoric acid) to quantify the total amide in the reaction mixture. Chiral HPLC analysis on (S,S) Whelk-O Chiral column used isopropanol hexane (30 70) as a solvent system to separate and quantify the (R)- and (S)-enantiomers. 

Aigbirhio, F., Pike, V.W., Francotte, E., Waters, S.L., Banfield, B., Jaeggi, K.A., Drake, A., 1992. 5-[l-(2,3-Diaminophenoxy)]-3 -(A -t-butylamino)propan-2 -ol simplified asymmetric synthesis with CD and chiral HPLC analysis. Tetrahedron Asymmetry 3, 539-554. Antoni, G., Ulin, J., Langstrom, B., 1989. Synthesis of the "C-labelled p-adrenergic receptor ligands atenolol, metoprolol and propranolol. Appl. Radial. Isot. 7, 561-564. 

Identified in a roasted Columbia arabica by Rathbone et al. (1989a,b) who suspected its presence in green coffee. The aqueous coffee extract was extracted with dichloromethane and, after TLC, the acid was analyzed by HPLC or the methyl ester by GC/MS. The level of the acid was 0.55-1.2 ppm in roasted coffee. In the second publication, the authors performed a chiral HPLC analysis of the ester and found the ( -enantiomer to be predominant (ca 80%). 

In order to clarify the absolute configuration of okaramines, we determined the absolute stereochemistry of the above-mentioned derivatives of cyclo (Trp-Trp) (20). Acid hydrolysis of 20 gave L-tryptophan, which was identified by comparison with standard D,L-tryptophan samples by chiral HPLC analysis. Thus, the absolute configuration at C-2 was proved to be S [21]. The absolute stereochemistries of 21, 22, 23, and 24 were defined by a CD comparison with cyclo (L-Trp-L-Trp) (20). Furthermore, acid hydrolysis of 23 afforded L-tryptophan through the loss of a reverse-prenyl side chain [26, 27]. Hydrolysis of 21, 22, and 24 also gave L-tryptophan. Accordingly, these results elucidated the absolute stereochemistries of 21, 22, 23, and 24 as those depicted [21]. 

The stereochemistries of okaramine C (4), okaramine J (11), okaramine K (12), okaramine L (13), and okaramine M (14), including the absolute configurations, were then established [21]. The absolute configuration at C-2 of 4, 11, 12, and 13 was determined to be S by chiral HPLC analysis of the acid hydrolysate of each of these compounds. NOESY and NOE difference experiments were carried out to define the stereochemistry of 11. Because NOEs were observed between H-2 and H-2 , between H-2 and H-8a, and between H-8a and 3a-OH, the absolute stereochemistry of 11 was determined. Based on NOESY and NOE difference data, the absolute stereochemistry of 11 was found to be identical to those of 4 and 13. The relative configurations at C-2, C-8a,... 

Thus we found that our purpose of further structural confirmation of 5 by total synthesis was completed, although the absolute value of the optical rotation of synthetic triacetate (6) was smaller than that of natural specimen of 6. The optical purity of the synthetic compound was examined by means of chiral HPLC analysis after conversion of the corresponding alcohol derived from 21 into tetraacetate (7), which had been obtained by ozonolysis of 5 [7], to reveal that the synthetic tetraacetate (7) obtained in this study was 60% ee. The optical purity of synthetic 6 was estimated to be parallel to this result. This result may be attributable to partial racemization during oxidation-reduction process to obtain the erythro-z coho (18). In our previous study [7], chiral HPLC had showed tihat no crucial racemization occurred since the erythro-zXcohoX (18) and its threo-Xsomtr were separated after conversion into monopivaloyl esters (10 and its isomer) by 4-times repeated silica gel chromatographies. 

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