Fluorometric Determination of Alkaline Phosphatase Activity in Food Using Magnetoliposomes as On-flow Microcontainer Devices

Abstract

Liposomes containing magnetic gold nanoparticles (AuNPs) and an enzymatic substrate (4-methylumbelliferyl-phosphate) have been used as on-flow microcontainers for reagent preconcentration in a flow injection method for the determination of alkaline phosphatase (ALP) activity. The dynamic range of the calibration graph was 6.4 × 10(-3)-0.25 U L(-1) ALP, and the detection limit was 1.9 × 10(-3) U L(-1). The precision, expressed as relative standard deviation (RSD%), was in the range of 0.7-2.4%. The overall method showed a sampling frequency of 10 h(-1). The method was applied to the determination of ALP in milk samples with recovery values ranging between 87.5 and 104.6%. The residual ALP activity in milk samples subjected to temperature treatments was also determined. The results obtained in the analysis of all milk samples were compared with those obtained by applying a previously described flow injection method.

Separation and purification of hydrophobic magnetite-gold hybrid nanoparticles by multiphase density gradient centrifugation

“…Although the synthesis of Fe 3 O 4 @AuNPs-C12SH has been previously described [11], it was again optimized in order to increase the final amount of the NPs obtained. This study also required to optimize again the synthesis of AuNPs, AuNPs-C12SH, Fe 3 O 4 NPs and Fe 3 O 4 @AuNPs.…”

“…Hydrophobic magnetic-gold NPs coated with C12SH (Fe 3 O 4 @AuNPs-C12SH) were obtained using a procedure described elsewhere [11]. The first step was the synthesis of Fe 3 O 4 NPs by the co-precipitation method [14], in which 25 mL of an aqueous solution containing 0.8 mol · L −1 iron (III) chloride, 0.4 mol · L −1 iron (II) chloride and 40 mmol · L −1 hydrochloric acid was dropwise added to 250 mL of 1.5 mol · L −1 sodium hydroxide solution under vigorous stirring using a nonmagnetic stirrer.…”

“…The method has been applied to the purification of hydrophobic magnetic-gold NPs coated with 1-dodecylthiol (Fe 3 O 4 @AuNPs-C12SH). These NPs have been previously synthesized for the preparation of magnetoliposomes [11], which were used to develop a method for alkaline phosphatase determination, but they can be contaminated with secondary NPs such as AuNPs, AuNPs-C12SH, Fe 3 O 4 NPs and Fe 3 O 4 @AuNPs. In addition to the use of different sucrose concentrations to form the layers, ficoll and percoll at constant concentrations have been used for the stabilization of the layers.…”

A Graphene Quantum Dots-Enzyme Hybrid System for the Fluorescence Assay of Alkaline Phosphatase Activity and Inhibitor Screening

“…Until recently, a number of methods for detecting the activity of ALP have been reported, including spectrofluorometry, 14,15 colorimetry, 16 chromatographic, 17 electrochemical, 18,19 chemiluminescence, 20 and surface enhanced resonance Raman scattering. 21 Among those, spectrofluorometry has become the most critical method because of the advantages of higher sensitivity, more convenience, less sophisticated instrument requirements, and a rapider response.…”

Ultrasensitive surface-enhanced Raman scattering detection of alkaline phosphatase

“…5,6 Moreover, the detection methods of ALP, which are based on nanomaterials, also have been reported. [7][8][9][10] Surface-enhanced Raman scattering (SERS) techniques are powerful tools for ultrasensitive detection. 11 In applications, the strong SERS signals have always been collected from molecules with both high affinity to SERS materials and strong intrinsic Raman activity.…”