A tri-enzyme system consisting of choline kinase/choline oxidase/horseradish peroxidase was used in the rapid and specific determination of the biomarker for bacterial sepsis infection, secretory phospholipase Group 2-IIA (sPLA2-IIA). These enzymes were individually immobilized onto the acrylic microspheres via succinimide groups for the preparation of an electrochemical biosensor. The reaction of sPLA2-IIA with its substrate initiated a cascading enzymatic reaction in the tri-enzyme system that led to the final production of hydrogen peroxide, which presence was indicated by the redox characteristics of potassium ferricyanide, K3Fe(CN)6. An amperometric biosensor based on enzyme conjugated acrylic microspheres and gold nanoparticles composite coated onto a carbon-paste screen printed electrode (SPE) was fabricated and the current measurement was performed at a low potential of 0.20 V. This enzymatic biosensor gave a linear range 0.01–100 ng/mL (R2 = 0.98304) with a detection limit recorded at 5 × 10−3 ng/mL towards sPLA2-IIA. Moreover, the biosensor showed good reproducibility (relative standard deviation (RSD) of 3.04% (n = 5). The biosensor response was reliable up to 25 days of storage at 4 °C. Analysis of human serum samples for sPLA2-IIA indicated that the biosensor has potential for rapid bacterial sepsis diagnosis in hospital emergency department.

The sensitive and rapid detection of hydrogen peroxide is very important in the areas of clinical and environmental analyses. A sensitive and selective Horseradish peroxidase (HRP)-hydrogen peroxide (H 2 O 2 ) biosensor was developed based on acrylic microspheres. Hydrophobic poly (n-butyl acrylate-N-acryloxysuccinimide) [poly(nBA-NAS)] microspheres were synthesized using photopolymerization in an emulsion to form an enzyme immobilization matrix. The HRP enzyme was covalently immobilized onto the acrylic microspheres via the succinimide functionality. Field emission scanning electron microscope (FESEM) has been utilized to characterize the screen-printed carbon paste electrode (SPE) constructed from enzyme conjugated acrylic microspheres and gold nanoparticles (AuNPs) composite (HRP/nBA-NAS/AuNPs/SPE). Differential pulse voltammetry was used to assess the biosensor performance. The linear response range of the hydrogen peroxide biosensor obtained was from 1.0 × 10 -2 to 1.0 × 10 -10 M (R 2 = 0.99) with the limit of detection (LOD) approximately at 1.0 × 10 -10 M. This is an improvement over many hydrogen peroxide biosensors reported so far. Such improvement may be attributed to the large surface area provided by the acrylic microspheres as a matrix for immobilization of the HRP enzyme. ABSTRAKPengesanan hidrogen peroksida yang sensitif dan cepat adalah sangat penting dalam bidang analisis persekitaran dan klinikal. Biosensor hidrogen peroksida (H 2 O 2 )-peroksidase lobak putih (HRP) yang sensitif dan memilih telah dibangunkan berasaskan mikrosfera akrilik. Mikrosfera hidrofobik poli (n-butil akrilat-N-akriloksuksinimida) [poli(nBA-NAS)] disintesis dengan menggunakan proses fotopempolimeran dalam bentuk emulsi dan ia bertindak sebagai matriks pemegun enzim. Enzim HRP dipegunkan secara kovalen pada mikrosfera akrilik melalui pengfungsian suksinimida. Mikroskop elektron imbasan pancaran medan (FESEM) telah digunakan bagi mencirikan elektrod karbon permukaan bercetak (SPE) yang dibina daripada komposit mikrosfera akrilik berkonjugat enzim dan nanosfera emas (AuNPs) (HRP/nBA-NAS/AuNPs/SPE). Voltametri denyutan pembezaan digunakan untuk penilaian prestasi biosensor. Julat keupayaan linear bagi biosensor hidrogen peroksida diperoleh daripada 1.0 × 10 -2 hingga 1.0 × 10 -10 M (R 2 = 0.99) dengan had pengesanan (LOD) ditemui pada 1 × 10 -10 M. Ini merupakan penambahbaikan berbanding biosensor hidrogen peroksida yang telah terlebih dahulu dilaporkan. Penambahbaikan ini mungkin ditentukan oleh luas permukaan yang besar yang disediakan oleh mikrosfera akrilik sebagai tapak pemegunan enzim HRP. Kata kunci: Biosensor; hidrogen peroksida; mikrosfera akrilik; peroksidase lobak putih

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