Trichloroacetic acid elute

Chromatography by ion exchange on a sulfonated poly(styrene-co-divinyl benzene) phase has been proposed as a replacement for titrimetry.57 Eluted by a dilute solution of a neutral salt such as sodium ethanesulfonate, the conductance of the protons can be measured in the absence of a suppressor from sub-millimolar to molar concentration. The response factors of mono-, di-, and trichloroacetic acid and of o-phthalic acid were large and essentially equivalent to ethanesulfonic acid, while the response factor of acetic acid was far smaller. A syringe pump has generated pressures as high as 72,000 psi (5000 bar) in a capillary column packed with 1 p particles, generating a fraction capacity of 300 peaks in 30 minutes.58... 

In ion-exclusion chromatography, ions are separated from nonelectrolytes by an ion-exchange column. Nonelectrolytes penetrate the stationary phase, whereas half of the ions are repelled by the fixed charges. Because electrolytes have access to less of the column volume, they are eluted before nonelectrolytes. The chromatogram here shows the separation of trichloroacetic acid (TCA, pKa = 0.5), dichloroacetic acid (DCA, pK.a = 1.1), and... 

The elution sequence was as follows 0.006M trichloroacetic acid (pH2.5), yielding first As(III) and then monomethylarsonate 0.2M trichloroacetic acid yielding As(V) 1.5M NH4OH followed by 0.2M trichloroacetic acid yielding dimethylarsinite. 

It was found that most of the 2-deoxyribosylic compounds in a trichloroacetic acid extract of the bacteria are present chemically in such a combination that snake-venom treatment is required in order to make them microbiologically active. Furthermore, when the extract was tested without venom pretreatment but in the presence of thymidine, the growth-promoting effect of the 2-deoxyribosylic compounds was 10 times that displayed without thymidine and reached 60% of the activity obtained after venom treatment. The nature of the substances responsible for this effect has not yet been described. The acidic nature of the 2-deoxyri-bosylic compounds was shown by the fact that 98% of the mixture was adsorbed to a Dowex anion exchanger. In the eluted fractions, thymidine 5-phosphate, thymidine 5-pyrophosphate, and thsmiidine 5-triphosphoric acid were identified. From the major 2-deoxyribosylic fraction, thymidine rhamnosyl pyrophosphate (XIX) has been isolated. This substance was also identified in the thymidine-requiring mutant of Escherichia coli 15 T-. 

The reaction mixture contained l-5-HTP as substrate, pyridoxyl phosphate as a cofactor, pargylcine HQ, and the enzyme. The reaction was terminated by the addition of trichloroacetic acid (TCA), and after addition of the internal standard the reaction mixture was clarified by centrifugation. The sample was prepurifled on Amberlite, and the 5-HT eluted and injected onto the HPLC column for quantitation. The results obtained the following the incubation of 5-HTP with the homogenate are shown in Figure 9.2B. The formation of... 

Fig. 2. Gel filtration profile of I-labeled human prolactin, [ I]hPRL (VLS No. 3) io-dinated by the chloramine-T method and purified on a 1.5 x 30 cm Sephadex G-100 column, precoated with bovine serum albumin. Elution solvent was 10 mM phosphate buffer, pH 7.5, containing 0.15 M NaCl and 1 10,000 Merthiolate. The labeled hormone was purified just prior to use. The talc-resin-trichloroacetic acid (TCA) test results showed that peak III material was suitable for use in radioimmunoassay (Tower et al. ).

The serum stability was studied by incubation of I-DOTATATE in human serum overnight at ambient temperature. Determination of serum protein binding was carried out by gel permeation chromatography using a Sephadex G-25 column (2.5 cm x 30 cm) and elution using 0.05M phosphate (elution volume 1 mL fractions). The l-DOTATATE was incubated in serum at ambient temperature for 18 h and then treated with 10% trichloroacetic acid for precipitation of the protein bound fraction. The supernatant was tested by HPLC to estimate its purity. 

Iodine-131 was introduced in one tyrosyl residue of the protein chain by adding 28 MBq of to 20 pg of anti-CD20 at pH7.4 and 10 pL of 0.13 pg/pL chloramine T. After incubating the reaction mixture for 1 min at room temperature, the radiolabelling yield was determined by protein precipitation with 10% trichloroacetic acid solution and purification was carried out by gel permeation with Sephadex G-25 (PD-10 columns, Pharmacia). Elution was carried out using 50mM phosphate saline buffer with 0.2% BSA [16.4]. The specific activity and extent of iodine incorporation were also determined. 

SpA-containing Staphylococci (Section 3.3), fixed with trichloroacetic acid (TCA) or formalin (Section 3.3.1), may also serve as immunosorbent for many mammalian antibodies (Table 7.1). Both fixation procedures are satisfactory but yield products with different properties. Fixation of Staphylococci with hot TCA (Lindmark, 1982) removes the negatively charged cell-wall polymer teichoic acid, producing an IgG-sorbent which can bind 1.4 mg human IgG per ml of a 10% (v/v) suspension of bacteria and is stable for about 5 months. Formalin-fixed bacteria (Kessler, 1976) bind 35% more IgG and are stable for at least 1 year. However, IgG can be eluted quantitatively from TCA-fixed bacteria but not from formalin-fixed bacteria, probably due to the interaction between IgG and teichoic acid, unless 80 mM MgCh is included in the acid buffer. 

This statement too may cause raised eyebrows but an objective assessment of errors and costs can support it. Thus, De Courcy s trichloroacetic acid reagent (D2) is capable of convenient application to the detection of pregnanetriol in the nonketonic fraction from one-sixteenth of a 24-hour collection from men (100-150 ml) on simple LLC paper chromatograms (a TLC method could no doubt also be devised for this). As she pointed out, the same quantity can be obtained from one-thousandth of a 24-hour collection (1.5-2 ml) of urine from some patients with adrenal hyperplasia. Quantitative estimation by the same reaction after elution from the paper or TLC chromatogram could also be used either by one of the many variations of the micro-17-ketogenic steroid methods or by the same fluorescence reaction (e.g., W7). Further confirmation of identity could be obtained by using a small fraction of the latter after the periodate oxidation for a paper LLC, or TLC, chromatogram and application of the Zimmerman reaction to demonstrate the formation of etiocholanolone (B32). 

Sample preparation Equilibrate a Sep-Pak silica SPE cartridge with 5 mL ethyl acetate. 1 mL Serum -i- 3 mL 5% trichloroacetic acid -I- 4 mL ethyl acetate, vortex vigorously, centrifuge at 1500 g for 5 min. Remove organic layer and repeat extraction twice with 3 mL portions of ethyl acetate. Combine extracts, evaporate to about 1.5 mL, add to the SPE cartridge. Wash with 8 mL ethyl acetate, elute with 4 mL MeOH ammonium hydroxide (90 10). Evaporate the eluent to dryness under nitrogen, reconstitute in 100 jiL MeOH, inject. 

In order to gain enough amount proteins for trypsin digestion and mass spectrometry detection, precipitate eluted proteins by trichloroacetic acid (TCA) and sodium deoxycholate (DOC) before loading onto a sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). 

Next, total rat hver proteins were examined for the presence of linkages characteristic of ADP-ribosyl-cysteine. Trichloroacetic acid insoluble extracts of hver tissue were treated to remove non-covalently bound ADP-ribose and subsequently treated with mercuric ion and analyzed for ADP-ribose. Fig. 2 shows such an analysis along with control experiments. Panel A shows that analysis of an extract treated with mercuric ion exhibited a fluorescent peak that migrated at the expected elution position of etheno-ADP-ribose, the fluorescent derivative used for quantitative determination of ADP-ribose. Panel B shows the analysis in which mercuric ion was omitted from a parahel sample of hver extract. Panel C shows the result obtained when chloroacetaldehyde, which is required for the formation of the fluorescent derivative of ADP-ribose, was omitted. This control rules out the possibility of endogenously fluorescent compounds present in the ceh extract that were released by mercuric ion. Panel D shows that a small amoimt of authentic etheno-ADP-ribose added to extracts prepared as in... 

Fio. 1-5. Nucleotides of the human erythrocyte. Trichloroacetic acid extracts of fresh erythrocytes were chromatographed on a column of Dowex-1 resin (formate form) eluted with a gradient of ammonium formate. [From G. Bartlett, Patterns of phosphate compounds in red blood cells of man and animals, in Adwinces in. Experimental Medicine and Biology (G. J. Brewer, ed.), Vol. 6, p. 245. Plenum Press, New York, 1970. Reproduced with permission.)... 

GSH (1 mM), and NADPH (1 mM) in potassium phosphate buffer (100 mM, pH 7.4) for 30 min [24], The total incubation volume was 2 mL. The incubation reactions were initiated by the addition of an NADPH solution after a 3-min preincubation, and were stopped by the addition of 300 pL of trichloroacetic acid (10%). After centrifugation (13,000 rpm for 10 min), supernatants were loaded onto SPE cartridges (Oasis extraction cartridges, Waters Corp., Milford, MA). The cartridges were washed with 1 mL of water and then eluted with 2 mL of methanol. The methanol fractions were dried and reconstituted with 200 pL of a water/acetonitrile mixture (v/v, 95 5). Aliquots (20 pL) of the reconstituted solutions were injected into LC-MS/MS. 

Many laboratories have succeeded in fractionating histones by selective extraction and differential precipitation. The selective extraction procedure is based on the difference in the amino acid composition of the various types of histones. For example, a histone-rich lysine can be obtained by extraction from whole thymus with 5% perchloric acid followed by precipitation with 10% trichloroacetic acid. If the histones found in the perchloric acid extract are passed through a carboxymethyl-cellulose column and eluted from the column with borate buffer at pH 9, then different components called 1, 2, and 3 can be recovered. 

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