Elution program —

Use of 10 pm LiChrosorb RP18 column and binary eluent of methanol and aqueous 0.1 M phosphate buffer (pH 4.0) according to suitable gradient elution program in less than 20-min run time with satisfactory precision sensitivity of spectrophotometric detection optimized, achieving for all additives considered detection limits ranging from 0.1 to 3.0 mg/1, below maximum permitted levels Simultaneous separation (20 min) of 14 synthetic colors using uncoated fused silica capillary column operated at 25 kV and elution with 18% acetonitrile and 82% 0.05 M sodium deoxycholate in borate-phosphate buffer (pH 7.8), recovery of all colors better than 82%... 

Chemical Purity. Selected eluates from a 4 liter elution program were subjected to analyses for chemical and biological purity. 

Table 2.1.4 High-performance liquid chromatography elution program...

U(VI). The long separation column—preconditioned, loaded, and washed with 3 M HN03—was credited with the apparent retention of Np(V) and Am(III) under this elution program. These authors also examined UTEVA-Resin for this actinide analysis application but concluded that TRU-Resin was to be preferred for retention of actinides from urine. 

Figure 6.2 Different shapes of elution programs in chromatography. Description of programs (a) step (b) linear (c) convex (d) concave (e) multisegment.

With simple, continuous elution programs the elution conditions for the individual peaks (in terms of peak width and detector sensitivity) will either be constant throughout the chromatogram, or will vary in a continuous way. 

A MicroPak MCH-10 reverse phase column was chosen for separation of the hexane and ether extracts. The monomolecular bonded phase provides efficient separation of both polar and non-polar substances and rapid equilibration to initial activity after gradient elution programs. The reverse phase column provides symmetrical, narrow peaks for the cannabinoic acids, which tend to tail on polar, normal phase columns (e.g. silica). 

To reduce interference from endogenous fats, the extract was partially purified by chromatographing five successive 20pl extract injections (equivalent to 0.5 ml milk each) on a MicroPak CN-10 column. A gradient elution program from hexane to 5% methanol in dichloro-methane was used for the purification step. The elution region corresponding to the neutral nonpolar cannabinoids was collected from each run. The fractions were then combined and analyzed on another MicroPak CN-10 column. 

The injection volume for these analyses is 1-50 yl. Under the conditions noted in the preceding table, benzidine elutes from the column in approximately 8.5 minutes. Following the elution of benzidine, a gradient elution program to 100% methanol in 5 minutes is effected to clean the column of later eluting compounds. This cleanup step is of critical importance in the analysis of bulk dye samples because of the presence of extensive contaminating material in the chloroform extract. For quantitation, the area of the sample peak is compared with that of appropriate standards. The limit of detection is 10 ng benzidine/injection. However, due to the relatively poor ex-... 

Figure 10 CEC-UV chromatogram (240 nm) of a mixture of 10 corticosteroids (100 qg/mL) using a linear gradient elution program. Voltage = 30 kV, HPLC injection volume = 10 pL, flow-rate = 10 pL/min for 3 min, then decreased to 100 pL/ min. Gradient program = initial ammonium acetate, 5 mM, in acetonitrile/water (17/83), held for 3 min, then ramped to 38% acetonitrile at 15 min and maintained to end of run. Column = Hypersil ODS, 3 pm, 42 cm total length, 30 cm packed length, 30.1 cm to window, 1 = triamcinolone, 2 = hydrocortisone and prednisolone co-eluting, 3 = cortisone, 4 = methylprednisolone, 5 = betamethasone, 6 = dexamethasone, 7 = adrenosterone, 8 = fluocortolone, 9 = triamcinolone aceto-nide. (Reprinted from Ref. 57, with permission.)...

Elution program (98 2) for 3 min to (2.88) over 23 min, hold for 10 min back to initial conditions over 2 min with equilibration of 8 min before next injection. 

The adaptation of the ABTS" method as an on-hne test required that the chromogen radical should be stable for sufficient time in different solvents to permit the utilization of isocratic or gradient elution programs. The on-line reaction time between ABTS" and potential antioxidants was an additional potential limiting factor. 

Another significant aspect was the stability of the radical chromogen ABTS in different solvents, in isocratic or gradient elution programs. We found that in the mobile phases used in our determinations (saline solutions and mixtures of organic solvents in different proportions), the observed fall was less than 0.01 expressed as — AAbs730nm/min. This stability is high... 

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