"Nanofiltration" Enabled by Super-Absorbent Polymer Beads for Concentrating Microorganisms in Water Samples

Abstract

Detection and quantification of pathogens in water is critical for the protection of human health and for drinking water safety and security. When the pathogen concentrations are low, large sample volumes (several liters) are needed to achieve reliable quantitative results. However, most microbial identification methods utilize relatively small sample volumes. As a consequence, a concentration step is often required to detect pathogens in natural waters. Herein, we introduce a novel water sample concentration method based on superabsorbent polymer (SAP) beads. When SAP beads swell with water, small molecules can be sorbed within the beads, but larger particles are excluded and, thus, concentrated in the residual non-sorbed water. To illustrate this approach, millimeter-sized poly(acrylamide-co-itaconic acid) (P(AM-co-IA)) beads are synthesized and successfully applied to concentrate water samples containing two model microorganisms: Escherichia coli and bacteriophage MS2. Experimental results indicate that the size of the water channel within water swollen P(AM-co-IA) hydrogel beads is on the order of several nanometers. The millimeter size coupled with a negative surface charge of the beads are shown to be critical in order to achieve high levels of concentration. This new concentration procedure is very fast, effective, scalable, and low-cost with no need for complex instrumentation.

Figure 1. Schematic diagram illustrating the water sample concentration procedures using super-absorbent polymer (SAP) beads.

The standard operation includes: (1) adding synthesized SAP beads into a water sample that requires a concentration step in order for a specific analytical procedure; (2) exposure of the beads for given amount of time for water uptake; (3) separating the swollen beads to collect the concentrate. The SAP beads can be recycled after drying.

Figure 2. Fabrication and characterization of millimeter-sized poly(acrylamide-co-itaconic acid) beads.

(a) Schematic of the fabrication process of the P(AM-co-IA) beads using a millifluidic system. (b) Picture of the P(AM-co-IA) beads as prepared. (c) Optical microscope image of one P(AM-co-IA) bead as prepared. (d) Size change of the P(AM-co-IA) beads when soaking in deionized water. The numbers at the top-right corner of the inset images indicate the soaking time. (e) Picture of the fully swollen P(AM-co-IA) hydrogel beads.

Figure 3. Performance of applying millimeter-sized P(AM-co-IA) beads to concentrate water samples containing microorganisms (here: E. coli or bacteriophage MS2).

(a–c) Change in water volumes and E. coli concentrations during sample concentration. The initial E. coli concentrations are different: (a) ~2 × 10²CFU/mL; (b) ~9 × 10² CFU/mL; and (c) ~9 × 10³CFU/mL. (d) Summary of the average and cumulative recovery efficiencies of the 5 concentrating steps for all concentration tests with different initial E. coli concentrations. (e) Change in water volumes and MS2 concentrations during water concentration. The initial MS2 concentration is ~6×10³PFU/mL. Dashed lines in (a–c,e) indicate theoretical concentrations calculated from the volume changes assuming 100% recovery during the concentration procedure.

Figure 4. Estimation of the water channel size within the P(AM-co-IA) hydrogel beads.

(a) Performance of applying millimeter-sized P(AM-co-IA) beads to concentrate water samples containing 5 nm gold nanoparticles (Au NPs). (b,c) Scanning electron microscope (SEM) images with energy dispersive spectrometry (EDS) elementary analysis of the outer surface (b) and cross-section (c) of the beads after drying with water containing Au NPs. Left: SEM images; Right: corresponding Au signal mapping by EDS. (d) Performance of applying millimeter-sized P(AM-co-IA) beads to concentrate water samples containing methyl orange (MO). Inset: P(AM-co-IA) hydrogel beads after use and intercalated by MO. Dashed lines in (a,d) indicate theoretical concentrations calculated from the volume changes assuming 100% recovery during the concentration procedure.