Alcohols, HPLC analysis

The phenylalanine-derived oxazolidinone featured here enjoys three practical advantages over the valine-derived oxazolidinone developed earlier in this laboratory. First, both the intermediate g-amino alcohol and the derived oxazolidinone are crystalline solids which can be purified conveniently by direct crystallization. Second, the oxazolidinone contains a UV chromophore which greatly facilitates TLC or HPLC analysis when it is employed as a chiral auxiliary. Finally, both enantiomers of phenylalanine are readily available, enabling stereocontrol in either sense simply by using the oxazolidinone derived from the appropriate enantiomer. 

Figure E3.4.3 Representative chromatograms. (A) Acetic acid standard (0.4 mg). (B) Methanol standard (2.0 mg). (C) Pectin sample. The start of each recording begins 12 min into HPLC analysis. Numbers above each peak refer to minutes elapsed after start of recording. The additional peaks in (C) are presumably due to other alcohols and acids released by alkaline treatment.

The second half of the 1990s saw an increase in the use of dialysis (as a liquid-liquid extraction procedure). Its main advantage is the possibility of operating in an automatic mode by coupling a dialysis unit with an automatic injector, as demonstrated not only in HPLC analysis (17) but also in flow-injection determinations of reducing sugars in wines (18) and alcoholic fermentation broths (19). 

RF McFeeters. Single injection HPLC analysis of acids, sugars, and alcohols in wine. Abstract of papers of the American Chemical Society, (22 August 1993), 206(1) 51, 1993. 

During HPLC analysis of aspartame, the most commonly used stationary phase is reverse-phase Cl 8. Two main types of mobile phase have been used an alcohol (methanol or isopropanol)... 

The solubility of NHDC in hot water, alcohol, aqueous alkali, acetonitrile, dimethyl sulfoxide, and alcohol/water mixture facilitates its selective extraction from food samples (20,91,94). It is extracted from jams, fruit juices, and dairy products with methanol (66,93) or acetone (95) and filtered or centrifuged. Chewing gum samples are dissolved in chloroform and extracted with water. The extract is centrifuged, and the clear supernatant is injected into the HPLC (95). If necessary, sample cleanup and concentration may be achieved by selective adsorption or desorption (20) on Sep-Pak Cl8 (96). Tomas-Barberan et al. (93) used Amberlite XAD-2 resin for purification of jam extract. Sugars, pectin, and other polar compounds were eluted with water, and NHDC was eluted with methanol. After concentration, the extract was further purified on a Sephadex LH-20 column prior to HPLC analysis. 

I McMurrough, JR Byrne. HPLC analysis of bittering substances, phenolic compounds, and various compounds of alcoholic beverages. In LML Nollet, ed. Food Analysis by HPLC. New York Marcel Dekker, 1992, pp 579-641. 

If fat-soluble vitamins (A, D and E) are added to a product, the first step in the analysis will often be hydrolysis. In fortified products this converts any esters, for example, vitamin A palmitate or vitamin E acetate, to their free alcohols. The product is then extracted with a hydrophobic solvent, such as hexane or diethyl ethyl or a mixture of these solvents, prior to HPLC analysis often using a reverse-phase (Ci8) column. 

These methods are not always applicable or convenient. A more general method used by Richard and Jencks utilizes HPLC analysis of carbocation formation in alcohol-water mixtures.22 As shown in Scheme 2 for an a-aryl ethyl cation, formation of the ether product from reaction of the carbocation with the alcohol depends on the rate constant for carbocation formation kll and the partition ratio between product formation and the back reaction to form the alcohol kROiiAii2o- This ratio may be determined from the ratio of products formed from reaction of the carbocation generated from a suitable solvolytic precursor such as an alkyl halide. 

The high-pressure cells and temperature control units are similar to the ones described by Betts and Bright (29). Samples for analysis were prepared by directly pipetting the appropriate amount of stock solution into the cell. To remove residual alcohol solvent, the optical cell was placed in a heated oven (60 °C) for several hrs. The cell was then removed from the oven, connected to the high-pressure pumping system (29), and a vacuum (50 pm Hg) maintained on the entire system for 10-15 minutes. The system was then charged with CF3H and pressurized to the desired value with the pump (Isco, model SFC-500). Typically, we performed experiments at 10 /xM PRODAN and there was no evidence for primary or secondary interfilter effects. HPLC analysis of PRODAN subjected to supercritical solvents showed no evidence of decomposition or additional components. 

Using similar reaction conditions with rapeseed oil, fatty acids were treated with various supercritical alcohols. From the HPLC analysis, it was shown that selective reactions could be obtained. Figure 5 presents the yields of alkyl esters of five fatty acids treated in various supercritical alcohols at 300°C. In the case of methanol, the reaction time for the complete conversion... 

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. 

Figure 5. HPLC analysis of human plasma from a marijuana smoker to which yill (A9-u-THC) has been added as internal standard. Mobile phase of 0.6% isopropyl alcohol in heptane used on a tandem alkylamine—alkylnitrile column with a flow rate of 60 mL/hr.

The oxidation reactions were performed in a glass batch reactor, equipped with magnetic stirrer (mechanic for L-sorbose oxidation), reflux condenser and thermometer. The reaction conditions are summarized in Table I. Before reaction the catalyst was pre-reduced in situ in a nitrogen atmosphere ( 20 min) with the alcohol reactant in 30-40 ml alkaline water (and dodecylbenzenesulfonic acid sodium salt detergent for water-insoluble reactants). The reactor worked in a mass transfer limited regime, controlled by the air flow rate (7.5-20 cm3min 1) and the mixing rate (1500-1800 min 1). The reactions were followed by GO or HPLC analysis. 

In Figure 5-41 the active steroid (triamcinolone acetonide) and preservative (benzyl alcohol) are determined from a steroid cream. The higher-molecular-weight components of the cream base are well separated from the analytes. The ability to elute all the components of a cream or ointment in an SMGPC analysis gives this approach an important advantage over competing separation techniques that require sample preparation prior to the HPLC analysis. 

Four luminescent cyclometallated iridium(III) diimine complexes [Ir(ppy-spacer-biotin)2(NAN)]+ (NAN = Me4-phen, Ph2-phen) (38), each containing two biotin units, have been synthesised and characterised by Lo and co-workers [79], Photoexcitation of these iridium(III) diimine bis(biotin) complexes in fluid solutions at 298 K and in alcohol glass at 77 K results in intense and long-lived 3MLCT (djr(Ir) — ti (NaN))/3IL (ir —> ji ) (Me4-phen) emission. HABA assays and emission titrations indicate that the two biotin moieties of each complex are functional. RET-based emission-quenching experiments, microscopy studies using avidin-conjugated microspheres, and HPLC analysis all reveal that the complexes with a... 

Enzyme assays were conducted in a 10 mL screw-neck glass test tube containing 100 fiL of lysate, 90 fiL of a 250 /ng/mL solution of 6-mercaptopurine in 0.01 M HC1, and 15 /uL of 250 mM sodium phosphate buffer (pH 9.2). Reactions were initiated by the addition of 32 fiL of a 3 1 mixture of 250 fiM S-adenosyl-L-methionine and 30 mM dithiothreitol. The final pH was 7.5. After a 1-hour incubation at 37°C, the reaction was stopped by the addition of 850 fjL of ice-cold 3.5 mM dithiothreitol and 50 fih of 1.5 M H2S04. The tubes were then heated at 100°C for 2 hours. To each tube, 500 fiL of 3.4 M NaOH was added, immediately followed by 8 mL of toluene-amyl alcohol-phenyl mercuric acetate. The tubes were shaken for 10 minutes and centrifuged. Then 6 mL of the toluene layer was transferred to a glass-stoppered conical test tube and 0.2 mL of 0.1 M HC1 added. After vortex-mixing and centrifuging, the toluene layer was discarded. Samples (50 fiL in 0.1 M HC1) were used for HPLC analysis. Product formation was linear for up to 120 minutes and 150 /u,L of lysate. 

The product from Step 4 (0.130 mol), 205 ml of isopropyl alcohol and potassium t-butylate (0.156 mol) were mixed, transferred into a 380 ml stirred stainless steel autoclave, and 4.0 ml of the prepared ruthenium asymmetric catalyst solution added. The mixture was hydrogenated at ambient temperature at 4 x 10 Pa 3 hours. Thereafter, the mixture was filtered, concentrated, and the residue dissolved in a water/ethyl acetate mixture. It was treated with a 5% NH4CI solution and the organic phase dried. HPLC analysis of an aliquot indicated that the cis/trans product ratio was 99 1, respectively, with an enantiomeric excess of (S,S)-cis-l,4-dibenzyl-piperidin-3-ol of 91%. 

Analysis of Reagent Purity the enantiomeric purity of the reagent can be evaluated by capillary GC analysis of its methyl ester on a chiral stationary phase, HPLC analysis of the corresponding l-(a-naphthyl)ethylamide, or by LiAlfli reduction to the corresponding alcohol, which is analyzed by chiral GC." ... 

The rate of imine disappearance increased from MeOH (relative rate 1.0) to EtOH (2.0), PrOH (2.0) and fPrOH (3.3), in agreement with the easier oxidation of these alcohols. BuOH does not follow this correlation since it induces a relative rate of only 1.8, giving rise to the formation of more side products. In the case of iPrOH, involvement of the intermediate hydroxyalkyl radical was corroborated through detection of its disproportion product acetone. These results show that the solvent can be directly involved in the oxidative step. Formation of hydrodimers in the absence of olefins thus can be explained by the oxidation of the alcohols. It is noted that in the presence of olefins no alcohol addition products could be detected by HPLC analysis, although methanol is present in a 500-fold molar excess. 

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