Other physiological fluids

The analysis of blood for organic acids presents a more complex problem with respect to sampling and sample preparation, although subsequent analytical stages may be identical. Although several different procedures for the preparation of blood for organic acid analysis have been recorded, relatively little attention has been paid to sampling and the type of specimen to be used. 

The technique of venepuncture may be important for certain organic acids, for example lactate, 3-hydroxybutyrate, etc., and results obtained on blood obtained by use of a tourniquet tend to be higher than those on samples obtained without stasis. Similarly, exercise may adversely affect the levels of acids observed. In general, blood samples should be taken without stasis, preferably after overnight fast, from the resting patient. 

Levels of acids in whole blood differ from those in plasma or serum and for most work, blood plasma is the most convenient for use, although whole blood has also been employed (Hagenfeldt, 1968). Serum obtained from clotted blood may give erroneous results since during the time required for the blood to clot, changes in the levels and composition of the organic acids can occur. Heparin is the most useful anticoagulant and sodium heparin tubes rather than heparin solutions are recommended (Perry and Jellum, 1974). The plasma 

The method of treatment of the plasma may differ depending on the acids of interest. If long-chain fatty acids and a lipid extract are required, deproteinization and extraction with chloroform/methanol is suitable (Johnston, 1971 Nelson, 1972 Kuksis, 1977). A suitable procedure is described below. Both free fatty acids and total fatty acid (free + esterified) may be determined in this type of extract. 

A suggested procedure is described below. Direct solvent extraction of acidified sodium chloride-saturated deproteinized plasma may also be employed if suitable for the metabolites of interest, but incomplete extraction of some components can result. 

---

It should be noted that with the use of a column guard to protect the analytical column from irreversible adsorption of proteins, it is sometimes possible to inject serum, urine, or other physiological fluids directly into the chromatograph without prior sample processing, especially if very small samples are used. 

Blood, urine, cerebrospinal, and other physiological fluids contain a great number of posttranslationally modified amino acids (approximately 170 have been studied to date) and in a wider range of concentrations than protein hydrolysates [6], Additionally, plant sources produce about 500 nonprotein amino acids and, in geological samples, highly unusual amino acids may indicate extraterrestrial origin [7, 8],... 

If the study being outsourced is a pharmacokinetic or clinical pharmacology study, does the CRO have bioanalytical chemistry laboratory facilities and expertise appropriate for the analysis of plasma, serum, or other physiological fluid samples for the desired analyte(s) Can your bioanalytical chemistry methods be transferred to their laboratories and validated or will samples be shipped to the sponsor or to another CRO that has the necessary method (s) up and running for analysis ... 

Concentrations of major carbohydrates in other physiological fluids are usually sufficiently high to permit a reliable profile analysis with the flame-ionization detector. Thus, the GC carbohydrate analyses have been described for plasma [412,413] as well as the seminal fluid from both normal and sterile men [164]. Several attempts have been made to relate the polyol concentrations in the human cerebrospinal fluid to certain pathological conditions [414-416]. If higher sensitivities are needed in the carbohydrate determinations, it is of advantage to consider perfluoroacyl derivatives and the electron-capture detector [402]. 

Quantification of free amino acids in serum, urine, or other physiological fluids The separation here is complicated by the presence of many interfering substances, such as small peptides and uncommon amino acids. This analysis is requested often for screening purposes. 

Simmonds HA, Duley JA, Davies PM. Analysis of purines and pyrimidines in blood, urine and other physiological fluids. Chapter 25. In Hommes F Ed, NY Wiley-Liss. Techniques in Diagnostic Human Biochemical Genetics A laboratory Manual 1991 p 397-424... 

Chalmers, R. A. (1974), A study of the acidic metabolites present in the urine of normal and mentally-subnormal human subjects and of patients with some metabolic diseases, together with some observations on other physiological fluids. Ph.D Thesis, Council for National Academic Awards. 

---