Eluents formic acid

There is increasing interest in copolymer systems, which, due to their chemical heterogeneity, may require very complex eluent systems in order to dissolve the sample and ensure that the separation ensues hy a pure size exclusion mechanism. In these examples, the PLgel is also compatible with eluent systems containing mixed solvents of different polarity (including water as a cosolvent up to 10% hy volume) and in organic solvents modified with acids or bases (e.g., acetic or formic acid, triethanolamine) as it is stable in the pH range of 1-14. 

Purify the tryptic peptides by chromatography on a C18 column to remove salts (follow the manufacturer s directions for peptide purification). Dry the eluent and redissolve the peptides in 20 pi 0.1 percent formic acid. 

Using the Tomtec Quadra 96 workstation, 0.1 mL of the ethyl acetate layer was transferred to a 96-well collection plate containing 0.4 mL of acetonitrile in each sample well. The solution was mixed 10 times by aspiration and dispersion on the Tomtec. The plate was then covered with a sealing mat and stored at 2 to 8°C until LC/MS/MS analysis. The HILIC-MS/MS system consisted of a Shimadzu 10ADVP HPLC system and Perkin Elmer Sciex API 3000 and 4000 tandem mass spectrometers operating in the positive ESI mode. The analytical column was Betasil silica (5 fim, 50 x 3 mm) and a mobile phase of acetonitrile water formic acid with a linear gradient elution from 95 5 0.1 to 73.5 26.5 0.1 was used for 2 min. The flow rate was 1.0 mL/min for the API 3000 and 1.5 mL/min for the API 4000 without any eluent split. The injection volume was 10 jjL and a run time of 2.75 min was employed. 

The diuretic and cholagog effect of Ononis arvensis motivated the development of a new two-dimensional paper chromatographic and TLC method for the measurement of onion in its roots and aerial parts. Samples were dried, ground and extracted with methanol (70 per cent, w/v) for 2 h. The supernatant was diluted and used for TLC separation on a cellulose stationary phase. The first eluent was 3 per cent formic acid, and the second was n-butanol-acetic acid-water (4 1 5, v/v) for both TLC and paper chromatography. Spots were scraped off, extracted with methanol and the absorption was measued at 260 nm. It was found that the average ononin content in roots and aerial parts was 0.153 0.0278 (per cent) and 0.498 0.045 (per cent), respectively. Because of the simplicity, the method was... 

SIS 1 silica stationary phase, eluent ethyl acetate-formic acid-water (6 1 1, v/v) S1S2 silica stationary phase, eluent toluene-ethyl formate-formic acid (4 4 1, v/v) S1S3 C18 stationary phase, eluent methanol-water-acetic acid (25 25 3, v/v) S1S4 silica stationary phase, eluent toluene-pyridine-formic acid (100 20 7, v/v) S1S5 cellulose stationary phase, eluent acetic acid-water (1 1, v/v) S1S6 C18 stationary phase, eluent methanol-water-acetic acid (78 25 3, v/v). Reprinted with permission from A. Pieroni et al. [124]. 

Figure 4.7 Anion exchange separation of carboxylic acids in red wine. Column, Shodex C811, 100 cm x 7.6 mm i.d. eluent, 3 mM perchloric acid flow rate, 0.9 ml min-1 temperature, 60 °C detection, reaction detection using chloro-phenol red at 430 nm. Peaks 1, citric acid 2, tartaric acid 3, malic acid 4, succinic acid 5, lactic acid 6, formic acid and 1, acetic acid.

The PO mode is a specific elution condition in HPLC enantiomer separation, which has received remarkable popularity especially for macrocyclic antibiotics CSPs and cyclodextrin-based CSPs. It is also applicable and often preferred over RP and NP modes for the separation of chiral acids on the cinchonan carbamate-type CSPs. The beneficial characteristics of the PO mode may arise from (i) the offset of nonspecific hydrophobic interactions, (ii) the faster elution speed, (iii) sometimes enhanced enan-tioselectivities, (iv) favorable peak shapes due to improved diffusive mass transfer in the intraparticulate pores, and last but not least, (v) less stress to the column, which may extend the column lifetime. Hence, it is rational to start separation attempts with such elution conditions. Typical eluents are composed of methanol, acetonitrile (ACN), or methanol-acetonitrile mixtures and to account for the ion-exchange retention mechanism the addition of a competitor acid that acts also as counterion (e.g., 0.5-2% glacial acetic acid or 0.1% formic acid) is required. A good choice for initial tests turned out to be a mobile phase being composed of methanol-glacial acetic acid-ammonium acetate (98 2 0.5 v/v/w). 

The use of formic acid, acetic acid and ammonium formate rather than triflu-oroacetic acid can substantially increase sensitivity because their proton affinities are lower than that of the TFA anion - though TFA is often used in the analysis of peptides. It is always advisable to keep the level of acid additives to less than 0.1% v/v, and preferably 0.03-0.05% v/v, in the final eluent. Triethylamine or ammonium hydroxide can be used successfully in negative mode because they promote deprotonation of acidic species. 

FIGURE 11.8 Isocratic elution separations of 20pg (panel A) and 10mg (panel B) of a melanotropin mixture. Column 4.6 x 250 mm ODS Eluent 0.25% formic acid, 0.5% triethylamine, 24% acetonitrile in water. Components 1, unknown 2, MSH sulfoxide 3, acetyl-MSH sulfoxide 4, MSH 5, acetyl-MSH. (Reprinted with permission from Elsevier from Barnthouse, K.A. et al., J. BiotechnoL, 66, 125, 1998. Copyright.)... 

The HPLC-FTIR technique has recently been used to identify six catechins and two methyl-xanthines present in green tea extracts." " A reversed-phase separation of the compounds was performed on a C-18 column equilibrated at 30°C using an isocratic mobile phase of acetonitrile-0.1% formic acid (15 85), prior to introduction to the deposition interface linked to the FTIR detector. The solvent was evaporated at 130°C and spectra were collected every 6 sec during the run. Two distinct designs for HPLC-FTIR interfaces have been developed flow cells and solvent elimination systems. Flow cell systems acquired spectra of the eluent in the solvent matrix through IR transparent, nonhydroscopic windows. The... 

Two ion chromatographic techniques were utilized to quantify formic acid in both diesel engine exhaust and mine air subjected to diesel emissions. A commonly used anion separation system utilizing a weak borate eluent adequately separated the acids of interest in diesel exhaust. It was, however, affected by the presence of strong acids during subsequent consecutive analyses. 

Volatile buffers were reconsidered for the modified method. Triethylamine was ruled out primarily because it could not be obtained in high purity and because the secondary and primary amines contaminating it could potentially react with solutes present in the water sample. Preliminary evidence of reaction between ethidium bromide and triethylammonium bicarbonate was obtained, but the reaction product was not characterized. The components of volatile buffers that appeared acceptable on the basis of chemical purity were ammonia, acetic acid, and formic acid. A few exploratory experiments were conducted involving the elution by ammonium formate and ammonium acetate of EB or quinaldic acid exchanged onto AG MP-50 or IRA 900. These experiments showed that 1 M ammonium formate in water was a very poor eluent, but that EB could be eluted from AG MP-50 with 1 M ammonium formate in methanol. Elution was essentially complete with 6 bed volumes of the methanolic eluent, whereas neither methanol alone nor aqueous 1 M ammonium formate was able to elute this solute. This situation pointed out the necessity for a counterion to displace exchanged solutes and, additionally, indicated that the displaced solute be highly soluble in the eluting solvent. 

High-performance LC was also used for determination of TBZ after its extraction from marmalades and curds with ethyl acetate (13). The use of a buffered mobile phase improved the response of the UV detector, and column performance remained constant throughout 2 months of daily use with a detection limit of 100 ppb. Three detectors (UV, fluorimetric, and electrochemical) were used for the determination of OPP, BP, and TBZ in plant materials (45). The compounds were extracted with dichloromethane and separated on an RP-18 column with a methanolic formic acid buffer as eluent. It was not possible to determine TBZ using an electrochemical detector, although the extraction recovery varied between 80 and 95%. 

Figure 6.11 Pseudo-on-flow 1 H NMR chromatograms of a spiked soil sample (a) with and (b) without31P decoupling. Conditions column, Purospher RP18, 250 x 4 mm, 5 pm eluents, A - acetonitrile, B - 1 % formic acid in D20 gradient, t = Omin A/B (1/99), t = 5 min A/B (10/90), t = 10min A/B (90/10) flow, l.Oml/min spectrometer, Bruker Avance 500MHz probe head, TX1 H/ C/31 4 mm LC probe 64 scans were acquired per row...

Karlson and Frankenberger [60] have developed a simple column ion-chromatographic column method for the determination of selenite in soil extracts with the simultaneous determination of chloride, nitrite, nitrate and phosphate. Separation of the anions was conducted on a low-capacity ion-exchange column, and anions were quantified by conductiometric detection. The eluent stream consisted of 1.5 mmol/1 phthalic acid and adjusted to pH 2.7 with formic acid. 

Eluent A 3% aqueous acetonitrile containing 0.1% formic acid. 

Fig. 8.22. Reconstructed ion chromatogram from the CEC—ESI-MS analysis of cefuroxime axetil diastereomers. Column, 900 x 0.05 mm i.d. packed with 3 pm Hypersil ODSA eluent, 5 mmol/1 sodium tetraborate, pH 9.0, 80% acetonitrile applied voltage, 30 kV detection, ESI-MS, 400-700 amu sheath liquid, 0.3% aqueous formic acid, 50% methanol, 10 pl/min injection, electrokinetic, 30 kV, 5 s. (Reproduced from ref. [98] with permission of John Wiley Sons).

The N7-guanine adduct (22) is a urinary excretion product derived from the reaction of sulfur mustard with DNA. It can be isolated from urine by SPE on C18. GC/MS analysis of the derivatized adduct was problematic. A sensitive method was developed for the underivatized compound using LC/ESI/MS/MS, monitoring the fragmentation MH+, m/z 256 —> [CH2CH2SCH2CH2OH]+, m/z 105 on a triple-sector quadrupole instrument (60). LC separation was on a C18 column eluted with water-acetonitrile-formic acid. The detection limit was 8pg injected (S/N 5 1), 0.2 ng/ml in urine. Rao et al. (61) also reported characterization by LC/ESI/MS but using 25 mM NH4HC03 in 20 % MeOH as eluent. 

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