Analytica Chimica Acta 797 (2013) 57–63 Contents lists available at ScienceDirect Analytica Chimica Acta j ourna l ho mepage: www.elsevier.com/locate/aca Biparametric multicommutated flow analysis system for determination of human serum phosphoesterase activity Lukasz Tymecki ∗ , Kamil Strzelak, Robert Koncki University of Warsaw, Department of Chemistry, Pasteura 1, 02-093 Warsaw, Poland h i g h l i g h t s • PEDD-based MCFA system for bipara- metric enzyme assays is developed. • The system is useful for simultaneous ACP and ALP assays. • The system is validated with real human serum samples. g r a p h i c a l a b s t r a c t a r t i c l e i n f o Article history: Received 4 March 2013 Received in revised form 21 July 2013 Accepted 28 August 2013 Available online 31 August 2013 Keywords: Flow analysis Paired emitter detector diode Enzyme assays Acid phosphatase Alkaline phosphatase Clinical diagnostics a b s t r a c t A multicommutated flow analysis (MCFA) system constructed of microsolenoid valves and pumps offer- ing simultaneous determination of activity of acid phosphatase (ACP) and alkaline phosphatase (ALP) in human serum samples has been developed. The MCFA system is based on optoelectronic flow-through detector made of two light emitting diodes and operating according to paired emitter detector diode (PEDD) principle. This photometric PEDD device has been dedicated for detection of p-nitrophenol (NP) generated in the course of enzymatic hydrolysis of p-nitrophenyl phosphate and optimized for the deter- mination of NP in human serum samples. The developed PEDD-based MCFA system allows independent optimization of conditions for reaction and detection steps of photometric ACP and ALP bioassays. More- over, it allows elimination of photometric interferences from serum matrix components according to two-points kinetic mode of measurement. The single measurement cycle takes 12 min, consists of four measurements (two for each phosphoesterase) and enables determination of serum ACP and ALP activi- ties at physiological and pathological levels. The real analytical utility of the developed MCFA system has been confirmed by analysis of control sera as well as real human serum samples from healthy persons and oncological patients. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Since several enzymes have been recognized as markers of health status, the evaluation of selected enzyme activities in human serum is fundamental for clinical diagnostics. Phosphomo- noesterases (phosphoric monoester hydrolases categorized with the EC number 3.1.3) belong to the group of indicative enzymes mostly assayed in the clinical practice. These nonspecific enzymes are responsible for dephosphorylation (removal of phosphoric ∗ Corresponding author. Tel.: +48 22 8220211. E-mail address: luktym@chem.uw.edu.pl (L. Tymecki). groups from phosphoesters of several alcohols, sugars, nucleotides, alkaloids and proteins). From the diagnostic point of view the most important are determinations of alkaline phosphatase (ALP, EC. 3.1.3.1) and acid phosphatase (ACP, EC. 3.1.3.2) levels. As it is indi- cated by the enzyme names, ALP exhibits maximal activity at higher pHs (8–10), whereas ACP at relatively low pHs (4–6). Increased serum ALP levels occur with all forms of cholesta- sis, particularly with obstructive jaundice caused by the blocking of bile ducts. A rise in serum ALP activities is also observed in case of several bone disorders [1]. The highest elevations of ALP occur in Paget’s disease (bone inflammation). Other skeletal disor- ders include hyperparathyroidism, osteomalacia (bone softening), fractures and rickets, osteogenic sarcoma and malignant tumors. A 0003-2670/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.aca.2013.08.047