Plasma sample preparation

Zimmer D. et al., 1999. Comparison of turbulent-flow chromatography with automated solid-phase extraction in 96-well plates and liquid-liquid extraction used as plasma sample preparation techniques for liquid chromatography-tandem mass spectrometry. J Chromatogr A 854 1999. 

Fig. 4 Scheme of enzymatic plasma sample preparation for enantioselective analysis of S- and fChyoscyamine. Atropine-containing plasma samples are mixed either with human serum (procedure A) or with rabbit serum (procedure B). The latter one contains atropinesterase (AtrE) whereas human serum does not. AtrE catalyses the stereoselective hydrolysis of S-Hyo, whereas f -Hyo remains unaffected. 

In addition to that, the stability of D-24851 was also assessed in the dry extract (after drying the organic solvent phase) and the compound proofed to be stable for at least three days (in case of plasma sample preparation at 2-8 °C). The dry extract of urine samples was stable for at least 7 days under the same conditions. 

D. Zimmer, V. Pickard, W. Czembor, C. Muller, Comparison ofTFC with automated SPE in 96-well plates and LEE used as plasma sample preparation techniques for LC-MS-MS, J. Chromatogr. A, 854 (1999) 23. 

Figure 4 LC/MS/MS of a mixture of gabapentin and naproxen using polarity switching. Micromass Quattro II, electrospray ionization rat plasma sample prepared by protein precipitation/extraction using a MetaChem 4.6 X 30-mm Polaris column with 3- Xm packing with a MetaChem 2000-MG2 guard column. Flow rate = 2.0 mL/min split 1 10 to MS. Injection volume 10 uL step gradient 100% 0.1% acetic acid, 0—1.0 min 70/30 0.1% acetic acid/acetoni-trile, 1.0—1.5 min 100% acetonitrile, 1.5—2.0 min. (Courtesy of Laura F. Polchinski, Ion Ebright, and Scott T. Fountain, personal communication.)...

For this reason, when a cassette is composed of structurally diverse compounds, it is usually prudent to use protein precipitation as a plasma sample preparation technique. Although the absolute recovery will vary from compound to compound, this approach can be applied with fairly universal success across a wide structural range of compounds and assay development time will be minimal. Generic solid-phase extraction protocols can also be effective, but will experience a higher (10 to 15% of compounds) failure rate. Chapter 6 contains examples of generic extraction protocols that can be adapted for use with cassettes containing diverse NCEs. 

DK Hieu, et al. Radioimmunoassay of the progestagen dienogest using different methods of plasma sample preparation. Pharmazie 41 711, 1986. 

Figure 5.34 Infusion chromatograms covering the LC/MS assay time (2.5 minutes), obtained using the post-column infusion method shown in Figure 5.33, comparing the ability of different sample preparation methods to remove endogenous sample matrix components that interfere with the ionization of phenacetin. Panels (a) - (f) show the variation with time of the MS signed specific for the infused standard (phenacitin) following on-column injection of 10 p,L of a blank plasma sample prepared by one of the tested sample preparation methods, (a) Protein precipitation, (b) Oasis SPE. (c) Methyl-tertbutyl ether (MTBE) hquid-liquid extraction, (d) Empore C2 disk SPE. (e) Empore C8 disk SPE. (f) Empore Cl 8 disk SPE. Reproduced from Bonfigho, Rapid Commun. Mass Spectrom. 13,1175 (1999), with permission of John Wiley Sons, Ltd.

Another problem is caused by low concentration of K vitamins within complex matrices. In case of plasma sample preparation, for example, often only a limited volume of plasma is available, especially when plasma of newborn babies is investigated. On the other hand, if one uses larger sample volumes, coextracted compounds may interfere with detection of the vitamins, or at least lead to extensive contamination of the chromatographic materials, which results in time-consuming cleaning processes and/or a shorter lifetime of chromatographic columns. 

A major advantage of this hydride approach lies in the separation of the remaining elements of the analyte solution from the element to be determined. Because the volatile hydrides are swept out of the analyte solution, the latter can be simply diverted to waste and not sent through the plasma flame Itself. Consequently potential interference from. sample-preparation constituents and by-products is reduced to very low levels. For example, a major interference for arsenic analysis arises from ions ArCE having m/z 75,77, which have the same integral m/z value as that of As+ ions themselves. Thus, any chlorides in the analyte solution (for example, from sea water) could produce serious interference in the accurate analysis of arsenic. The option of diverting the used analyte solution away from the plasma flame facilitates accurate, sensitive analysis of isotope concentrations. Inlet systems for generation of volatile hydrides can operate continuously or batchwise. 

Gold is a useflil caUbration standard for this method (see Radioactive tracers). Whereas similar sensitivities can be achieved by inductively coupled plasma mass spectrometry (qv), the latter requires more extensive sample preparation to overcome interference by other metals such as copper (64). 

The very low Hg concentration levels in ice core of remote glaciers require an ultra-sensitive analytical technique as well as a contamination-free sample preparation methodology. The potential of two analytical techniques for Hg determination - cold vapour inductively coupled plasma mass spectrometry (CV ICP-SFMS) and atomic fluorescence spectrometry (AFS) with gold amalgamation was studied. 

The complex of the following destmctive and nondestmctive analytical methods was used for studying the composition of sponges inductively coupled plasma mass-spectrometry (ICP-MS), X-ray fluorescence (XRF), electron probe microanalysis (EPMA), and atomic absorption spectrometry (AAS). Techniques of sample preparation were developed for each method and their metrological characteristics were defined. Relative standard deviations for all the elements did not exceed 0.25 within detection limit. The accuracy of techniques elaborated was checked with the method of additions and control methods of analysis. 

Direct sampling of solids may be carried out using laser ablation. In this technique a high-power laser, usually a pulsed Nd-YAG laser, is used to vaporize the solid, which is then swept into the plasma for ionization. Besides not requiring dissolution or other chemistry to be performed on the sample, laser ablation ICPMS (LA-ICPMS) allows spatial resolution of 20-50 pm. Depth resolution is 1-10 pm per pulse. This aspect gives LA-ICPMS unique dit nostic capabilities for geologic samples, surface features, and other inhomogeneous samples. In addition minimal, or no, sample preparation is required. 

K. Ki onkvist, M. Gustavsson, A.-K. Wendel and H. Jaegfeldt, Automated sample preparation foi the determination of budesonide in plasma samples by liquid chi omatography and tandem mass specti ometi y , 7. Chromatogr. A 823 401-409 (1998). 

R. Herraez-Heruandez, A. J. H. Eouter, N. C. van de Merbel and U. A. Th Brinkman, Automated on-line dialysis for sample preparation for gas cliromatogruphy determination of benzodiazepines in human plasma , 7. Pharm. Biomed. Anal. 14 1077-1087 (1996). 

The quantification of kinins in human tissues or body fluids has been limited due to the inherent difficulties in accurately measuring the concentration of ephemeral peptides. Today HPLC-based and RIA/capture-ELA measurements are established to determine kinins in human plasma, liquor or mine. Serine protease inhibitors need to be added to prevent rapid degradation of the kinins in vitro during sample preparation. Kinins and their degradation products have been studied in various biological milieus such as plasma/ serum, urine, joint fluids, kidney, lung and skeletal muscle [2]. Under normal conditions, the concentration of kinins in these compartments is extremely low for... 

According to the characterizations by TEM and XRD, the sample prepared from a CH4/H2 plasma was composed of nanocrystalline diamond and disordered microcrystalline graphite. Then nondiamond carbon was effectively removed with an increase in [CO]. It is therefore concluded that the VDOS of the nanocrystalline diamond and DEC films extracted from the HREELS data is in qualitative agreement with the characterizations of TEM and XRD. Although the HREELS probes only the region near the surface, the agreement suggests that the surface dynamics do not differ dramatically from those of the bulk. 

Sample preparation for the common desorption/ionisation (DI) methods varies greatly. Films of solid inorganic or organic samples may be analysed with DI mass spectrometry, but sample preparation as a solution for LSIMS and FAB is far more common. The sample molecules are dissolved in a low-vapour-pressure liquid solvent - usually glycerol or nitrobenzyl alcohol. Other solvents have also been used for more specialised applications. Key requirements for the solvent matrix are sample solubility, low solvent volatility and muted acid - base or redox reactivity. In FAB and LSIMS, the special art of sample preparation in the selection of a solvent matrix, and then manipulation of the mass spectral data afterwards to minimise its contribution, still predominates. Incident particles in FAB and LSIMS are generated in filament ionisation sources or plasma discharge sources. 

TXRF has also been used for the characterisation of single, colourless textile fibres (polyesters, modified cellulose and wool), yielding a fingerprint trace-element pattern, suitable for forensic purposes [276,277], Sample preparation involved dissolution/predigestion in HN03 and matrix removal (O2 cold plasma). 

Deng, Y., Wu, J., Lloyd, T.L., Chi, C.L., Olah, T.V., Unger, S.E. (2002). High-speed gradient parallel liquid chromatography/tandem mass spectrometry with fully automated sample preparation for hioanalysis 30 seconds per sample from plasma. Rapid Commun. Mass Spectrom. 16, 1116-1123. 

An alternative system proved to be both simpler and more user friendly (Unger et al., 2004 Machtejevas et al., 2006). Thus far we have used this configuration to analyze human plasma, sputum, urine, cerebrospinal fluid, and rat plasma. For each particular analysis we set up an analytical system based on a simple but specific strategy (Figure 9.5). The analysis concept is based on an online sample preparation and a two-dimensional LC system preseparating the majority of the matrix components from the analytes that are retained on a RAM-SCX column followed by a solvent switch and transfer of the trapped peptides. The SCX elution used five salt steps created by mixing 20 mM phosphate buffer (pH 2.5) (eluent Al) and 20 mM phosphate buffer with 1.5 M sodium chloride (eluent Bl) in the following proportions 85/15 70/30 65/45 45/55 0/100 with at the constant 0.1 mL/min flow rate. Desorption of the... 

FIGURE 9.6 The peptide and small protein map from a 100 pL human plasma injection. Columns sample preparation SCX RAM analytical column chromolith performance RP-18, 100 x 0.1 mm I.D. Minute fractions were analyzed using MALDI-TOF MS. Fraction numbers correspond to the time scale. Dot size is related to signal intensity. 

Additionally, the inj ected matrix must also be miscible with the solvents used in the separations. For normal phase mode separations, all water must be removed from the injected matrix. Since many of the complex matrixes, such as plasma, urine, and other biological fluids contain a large amount of water, this requires more time consuming sample preparation. However, water can be injected into a polar organic or reverse phase mode separation. Even within the same mode, mobile phases that are very different can cause large disturbances in the baseline. Oda et al., (1991) solved this problem by inserting a dilution tube followed by a trap column in order to dilute the mobile phase used on the achiral column. Following the dilution tube, a trap column was used to reconcentrate the analyte of interest before the enantiomeric separation. 

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