Ion reversed phase

Takayama and coworkers (60) introduced the h.p.l.c. separation technique for such amphiphilic molecules as lipid A, and in earlier experiments they applied paired-ion reverse-phase h.p.l.c. for the preparation of homogeneous fractions deriving from 4,-monophosphated lipid A of S. typhimur-ium. The purified preparations obtained were suitable for f.a.b. - m.s. analysis. However, monophosphated lipid A isolated in this way expressed a considerable heterogeneity with respect to the number and location of 0-acyl residues (60). In order to further improve the purification procedure, as well as to obtain lipid A derivatives suitable for n.m.r. spectroscopy, Qureshi et al. (174) prepared the dimethyl phosphate derivative of S. minnesota (R595) lipid A, which, after purification by reverse-phase h.p.l.c. (C18), could be analyzed by1 H-n.m.r. The n.m.r. spectrum of, for example, the heptaacyl lipid A dimethyl monophosphate fraction, unequivocally revealed 0-acyl substitution [14 0(3-OH)J at position 3 and a free hydroxyl group at position 4 of GlcN(I). 

Separation can now be achieved by using any one and/or a combination of the various modes available. These modes include size exclusion, normal phase, reversed-phase, paired-ion reversed-phase and ion-exchange. The principles, theories and the... 

A paired-ion, reversed-phase high-performance liquid chromatographic method was developed for the simultaneous determination of sweeteners (dulcin, saccharin-Na, and acesulfame-K), preservatives (sodium dehydroacetate, SA, salicyclic acid, BA, succinic acid, methyl-para-hydroxybenzoic acid, ethyl-para-hydroxybenzoic acid, n-propyl-para-hydroxybenzoic acid, n-butyl-para-hydroxybenzoic acid, and isobutyl-para-hydroxybenzoic acid), and antioxidants (3-tertiary-butyl-4-hydroxyanisole and tertiary-butyl-hydroquinone). A mobile phase of acetonitrile-50 ml aqueous tr-hydroxyisobutyric acid solution (pH 4.5) (2.2 3.4 or 2.4 3.6, v/v) containing 2.5 mM hexadecyltrimethylammonium bromide and a Clg column with a flow rate of 1.0 ml/min and detection at 233 nm were used. This method was found to be very reproducible detection limits ranged from 0.15 to 3.00 p,g. The retention factor (k) of each additive could be affected by the concentrations of hexadecyltrimethylammonium bromide and a-hydroxyisobu-tyric acid and the pH and ratio of mobile phase. The presence of additives in dried roast beef and sugared fruit was determined. The method is suitable for routine analysis of additives in food samples (81). 

Figure 9.110 Separation of adenine nucleotides and adenosine by paired-ion reversed-phase HPLC Standards of AMP, adenosine, ADP, ATP, and cAMP (approximately 2 nmol each) in Tris-HG (pH 7.4) were injected onto a C18 /xBondapak reversed-phase column (300 mm X 7.8 mm) and eluted with 65 mM KH2P04 (pH 3.6), 1 mM tetrabutylammonium phosphate, and 3% acetonitrile. The flow rate was 2 mL/min, and detection was at 254 nm. (From Rossomando and Jahngen, 1983.)...

D. Bushec. I.S. Krull. R.N. Savage and S.B. Smith. Jr., Metal calion/anion speciation via paired-ion, reversed phase HPLC with refractive index and/or inductively coupled plasma emission spee-troseopic detection methods, / Eiq. Chromatogr.. S. 463. 1982. 

Urinary iodide was measured using paired-ion reversed-phase high-performance liquid chromatography (HPLC) with electrochemical detection employing a silver working electrode (Rendl etal., 1994). A detection limit of 5 pg was achieved. 

Size exclusion, reversed-phase, paired ion reverse-phase, and ion exchange (cation and anion) HPLC have been used for selenium speciation. Size exclusion, also used in combination with affinity chromatography, has been used for the determination of selenoproteins and studies of interactions of selenium with proteins in the body. Ion-exchange and reversed-phase HPLC are often used for separation... 

Kwok, R.P., Rose, W.P., Tabor, R., and Pattison, T.S., 1981. Simultaneous determination of vitamins Bl, B2, B6, and niacinamide in multivitamin pharmaceutical preparations by paired-ion reversed-phase high-pressure liquid chromatography. Journal of Pharmaceutical Sciences. 70 1014—1017. 

The combination of GC with mass spectrometry has been used to study pyridoxine metabolism in tumor cells. Using radiolabeled PN and paired-ion, reversed-phase HPLC, Tryfiates and coworkers (158) demonstrated that about 30% of the radiolabel was associated with a product showing a retention time different from any of the known B vitamers. Using mass spectrometry, with and without prior GLC separation, the product was eventually identified as adenosine-N -diethylthioether-N -pyridoximine-5 -phosphate (158,159). Recent studies from Tryfiates and Bishop indicate that the above-mentioned novel Be vitamer... 

Mondello L, Tranchida PQ, Stanek V, Jandera P, Dugo G, Dugo P. Silver-ion reversed-phase comprehensive two-dimensional liquid chromatography combined with mass spectrometric detection in lipidic food analysis. J Chromatogr A. 2005 1086 91-8. 

Finley, J.M. and Duang, D., Resolution of ascorbic, dehydroascorbic and diketogulonic adds by paired-ion reversed-phase chromatography, J. Chromatogr., 207, 449-453, 1981. 

We report here on the direct interfacing of paired-ion, reversed phase HPLC methods for the initial separation of inorganic Se species, sele-nate and selenite, followed by an on-line, real-time, continuous interfacing with DCP There was, as before, a direct interfacing via a short, flexible Tefzel connector, which permited continuous HPLC effluent introduction into the DCP spray chamber at conventional flow rates for inorganic anion separations to be effective. The overall approach, HPLC-DCI was very similar to what has been reported for Cr and Sn... 

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