Reduction mode

An ECD measures the current generated by electroactive analytes in the HPLC eluent between electrodes in the flow cell. It offers sensitive detection (pg levels) of catecholamines, neurotransmitters, sugars, glycoproteins, and compounds containing phenolic, hydroxyl, amino, diazo, or nitro functional groups. The detector can be the amperometric, pulsed-amperometric, or coulometric type, with the electrodes made from vitreous or glassy carbon, silver, gold, or platinum, operated in the oxidative or reductive mode. Manufacturers include BSA, ESA, and Shimadzu. 

Barek et al. have reported on the determination of AT-nitroso compounds, azo compounds, heterocychcs, aromatic nitro compounds, heterocychc amines and even benzyl chloride using electrochemical methods such as voltammetry and polarog-raphy. The nitro and AT-nitroso compounds work particularly well in reductive mode [47, 48]. For appropriate analytes, adsorptive stripping voltammetry and anodic stripping voltammetry can give orders of magnitude lower detection hmits than are available from HPLC with electrochemical detection [48]. 

All metals at trace concentration, or in trace quantities, can be analyzed by atomic absorption (AA) spectrophotometry in flame or graphite furnace (electrothermal reduction) mode. A rapid, multi-element analysis may use... 

An FIPLC method using electrochemical detection in the reductive mode for the determination of artemether 28a and its metabolite dihydroartemisinin 29a <1997JCFI(B)145> and for the simultaneous quantification of artesunate 31 and dihydroartemisinin 29a in plasma has been developed <1997JCFI(B)259, 1998JCFI(B)201>. An effective reversed-phase FIPLC method using electrochemical and UV detection has been developed for the simultaneous determination in plant extracts of artemisinin and its bioprecursors such as arteannuin B 32a, and artemisitene 27 <1995JNP798, 2001JIC489>. 

Electrochemical detection, in the reductive mode, has been successively applied by Carignan et al. (380) for the determination of underivatized dimetridazole residues in swine muscle. The major difficulty in this detection mode is the strong interference by oxygen, thus deoxygenation of the entire system is required. 

The first combined HPLC-electrochemical measurements of vitamin K used the reductive mode, but this technique suffered from interference from the reduction of oxygen. A redox method was later developed that eliminated this interference, and provided a 10-fold increase in sensitivity over photometric detection and an improved selectivity. The coulometric detector employed in the redox mode is equipped with a dual-electrode cell in which phylloquinone is first reduced upstream at the generator electrode and the hydroquinone is reoxidized downstream at the detector electrode. 

MeCN/CH2Cl2/aqueous 0.025 M sodium perchlorate, 90 5 5 Amperometric detection (reductive mode), silver electrode, — 1.1 V vs saturated calomel reference electrode... 

Amperometric 1 x10-9g 2 x 104 Responds to -OH functionalities Used for alphatic and aromatic -OH compounds, amines, and indoles pulsed potential units are most sensitive, can be used with gradient elution and organic mobile phases senses compounds in oxidatitive or reductive modes mobile phases must be highly pure and purged of 02... 

Operation of amperometric detectors in the reductive mode is more difficult because dissolved oxygen in the sample and mobile phase will interfere. Nevertheless, the quinone of doxorubicin and the nitro group of chloramphenicol have been reduced [94]. 

K. Bratin, P. T. Kissinger, and C. S. Brunlett, Reductive mode thin-layer amperometric detector for liquid chromatography, J. Liquid Chromatogr., 4-.Y111 (1981). 

Alawi [319] has discussed an indirect method for the determination of nitrite and nitrate in surface, ground and rain water by reaction with excess phenol (nitrite ions first being oxidised to nitrate) and extraction of the o-nitrophenol produced, followed by separation on a reversed phase high performance liquid chromatography column with amperometric detection in the reduction mode. Recoveries were 82% for nitrate and 77% for nitrite in the concentration range 10-lOOOpg L 1. The method is claimed to be free of interferences from other ions. 

The development of the reductive mode LCEC technique has been slow because of difficulties in preparing convenient and reliable working electrodes for use with a high efficiency chromatographic separation. In addition, problems are encountered with dissolved oxygen and heavy metals. Solid electrodes have been used with limited success for reductive LCEC. Mercury pool electrodes (44-47), the DME (48-53), and platinum wire electrodes coated with mercury (49) are generally not satis-... 

Dissolved oxygen seriously limits the range of the reductive mode detector if not removed from the mobile phase and sample. 

Nitrogen a Dohrmann microcoulometric reactor being used in reductive mode was replaced with an Antek Model 771 pyroreactor which uses a chemiluminescent nitrogen detector. 

The major contributions which result in the reduced chromatographic efficiency have been ascribed to slow mass transfer principally due to poor wetting of the surfactant modified stationary phase (109), poor mass transfer between the micelle and stationary phase (113), and poor mass transfer in the stationary phase (100,106). In some cases, the use of small amounts of alcohol additives (MeOH, n-PrOH) and operation at elevated temperature (MO0 C) result in chromatographic efficiencies comparable to that seen in traditional LC using hydro-organic mobile phases (109,113,154,206). In our own work, we have found n-pentanol to be superior to n-propanol in this regard (refer to Table IX) (112). Further work is clearly needed in this efficiency area in order to clarify the exact reason(s) for the reduction in efficiency. It appears that a combination of factors can contribute to this effect with the dominant efficiency reduction mode dependent upon the nature of the solute, micellar mobile phase, and stationary phase packing material employed (100,112,135). 

Alternatively, the reaction could be performed in a reduction mode, using the ketone as substrate and up to 50% v/v isopropanol as the reductant, affording the corresponding (S)-alcohol in 99% ee at conversions ranging from 65 to 92%. 

Electrochemical detectors can be used in the oxidative mode for a wide range of drugs, including cannabinoids, haloperidol, morphine, paracetamol, phenothiazines, salicylic acid, and tricyclic antidepressants. Operation in the reductive mode is more difficult as dissolved oxygen must be removed from the eluent (K. Bratin et al, J. liq. Chromat., 1981,, 1777-1795). Reductive applications include chloramphenicol and benzodiazepines. 

The results were interpreted in terms of steric requirements of the actual reducing species. It was suggested that attack of BH4" proceeds exclusively along the 1,4-reduction mode, whereas alkoxyborohy-drides (formed as reaction products) prefer the 1,2-reduction mode. The pyridine borine itself does not reduce enones under the reaction conditions, but it inhibits formation of alkoxyborohydrides. - The same trend was observed with aluminum hydride reductions. When LiAlH4 was first reacted with pyridine to form lithium tetrakis(dihydro-N-pyridyl)aluminate, 1,4-reduction predominated. ... 

Stereoselective reduction of an enone lactone was a key step in the construction of the 20-hydroxyec-dysone side chain. Totally different mixtures of products were obtained when the reduction was carried out with sodium borohydride or by catalytic hydrogenation (Scheme 30). In all cases, the 1,4-reduction mode is preferred. With borohydride, however, this process is followed by a subsequent reduction of the saturated ketone and base-catalyzed rearrangement of the 5-lactone into a y-lactone. 

High catalytic activities, with turnovers of up to 9(X) cycles min , is displayed in the transfer hydrogenation of a,p-unsaturated ketones, such as benzylideneacetone and chalcone, using 2-propanol and catalytic amounts of [Ir(3,4,7,8-Me4-phen)COD]Cl (phen = 1,10-phenanthroline COD = 1,5-cyclo-octadiene) in a weakly alkaline medium. Other Ir-chelated complexes are also active catalysts in this reaction, with over 95% selectivity for the 1,4-reduction mode. Divalent lanthanide derivatives, such as Sml2 or Ybh in stoichiometric quantities, in THF and t-butyl alcohol or methanol reduce ethyl cinnamate and cinnamic acid to give the saturated derivatives. " Similarly, 3-methylcyclohexenone is reduced to 3-methylcyclohexen-l-ol in 67% yield, but a,p-unsaturated aldehydes are nonselectively reduced with these systems. 

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