High-pressure liquid chromatography Detection systems

Since ISEs can be used in continuous flow systems or in flow systems with sample injection (flow injection analysis, FIA)21 their application is wide, not limited to discrete samples. Analysis time becomes shorter, with faster recycling. Additionally, in flow systems the experimental assembly and data analysis can be controlled automatically by microcomputer, including periodic calibration. Another development is the use of sensors for the detection of eluents of chromatographic columns in high-pressure liquid chromatography (HPLC). Miniaturization has permitted an increase in the use of sensors in foods, biological tissues, and clinical analyses in general. 

Internal Standard Solution. A solution containing 125 ag/ml ofN-propyl-4-aminophenol in methanol. Method. To 50 lal of plasma, serum, or whole blood in a 1-ml polypropylene cenhifuge tube add 100 p.1 of the internal standard solution, mix, and cenfiifuge for 2 minutes in a microcentrifuge. Examine 5 pi of the supernatant liquid by high pressure liquid chromatography, using die system described below, with ultraviolet detection at 240 nm. 

Dissolve the residue in 150 Ltl of tetrahydrofuran and examine by high pressure liquid chromatography using the system described below with ultraviolet detection at 240 nm, and applying 10 Lll to the column. 

High pressure liquid chromatography (HPLC) was used to identify phenols and separate the products of the reaction. The system included a Dupont 870 pump module, 850 absorbance detector, and a 4.6 mm x 25 cm Zorbax ODS liquid chromatography column (all from DuPont and Co., Wilmington, DE). Analysis of 2-CP was achieved under the following conditions mobile phase 33% MeOH in water at 1.5 mL/min PCP analysis mobile pliase 50% MeOH in water at 1.5 mL/min UV detection for 2-CP and PCP at 254 nm. 

For analytical purposes cholesterol oxidase has been immobilized on various carriers (Table 6). Electrochemical, optical, and calorimetric indication have been used as detection methods. Combination of a thermistor-coupled flow-through system with immobilized COD permitted the measurement of 0.03-0.15 mmolA cholesterol (Mattiasson et al., 1976). Ogren et al. (1980) described an immobilized COD reactor for the analysis of steroid fractions obtained by high pressure liquid chromatography. The UV absorption at 240 nm of enzymatically formed cholestenone was used as the measuring signal. Linearity was found between 10 and 80 pmol/1. 

The fluorimetric method of Bates and Rapoport [8], based on the oxidation of PSP toxins in alkaline conditions to form fluorescent derivatives, was incorporated into a detection method with the PSP toxins separated in a chromatographic column by Buckley et al. [17]. This method set the basis for the development of a high pressure liquid chromatography with postcolumn reaction system that was subsequently improved to achieve a better toxin separation and adequate sensitivity [18]. Sullivan et al. [ 19] evaluated its applicability to shellfish toxicity monitoring, by comparing the results obtained by the HPLC method and the standard Association of Official Analytical Chemists (AOAC) mouse bioassay. They found, in general, a good correlation between the two methods. However, Cl and C2 toxins could not be separated and individually quantified. Further improvements and modifications... 

Biochemical analysis by high-pressure liquid chromatography (HPLC) and mass spectrometry (MS) enables analysis of the perfundate for metabolites of a liver cell culture or for tracer compounds used to assess the permeability of barrier organ systems. Also ELISA assays may be used to detect secretion of compounds such as albumin and urea from liver cultures. 

Other analytical methods such as reversed-phase high-pressure liquid chromatography (RPHPLC) have also been used to determine the quantity of protein released from the drug delivery system. Figure 9 shows a release profrle of insulin from an ATRIGEL formulation. Insulin was detected in its native form by RPHPLC through day 15. 

---