Portable Rapid Paper-Based Device for Analysis of Carbamates and Organophosphates in Water Samples
Authors: Marques Fernanda , Mitra Sushanta 
Published: July 2022

Traditionally, the analysis of contaminants in water samples is performed using sophisticated techniques, namely HPLC, GC, and MS, requiring qualified personnel and laboratory infrastructure. Such methods are not feasible for the detection of contaminants in the field. In contrast, paper-based portable devices have been demonstrated as versatile, sustainable, and cheap solutions for such purposes. Paper consists of a pressed sheet of entangled hydrophilic cellulose fibres with excellent porosity. Such features enable the capillarity motion of liquids and the immobilization of active substances on the surface.
Pesticides are critical pollutants in regions where agricultural activity is intense and represent a serious health risk for rural workers and their communities. These chemicals have been applied in enormous quantities to increase crop production in many countries. Carbamates (CBs) and organophosphates (OPs) are employed as active ingredients in industrial formulations of insecticides. They are considered highly toxic due to their inhibition action in the cholinergic transmission found in the nervous system of living organisms.
This work aims to present a low-cost and simple operational paper-based testing device for the fast detection of CBs and OPs in water samples based on Ellman’s method of acetylcholinesterase (AChE) inhibition. We have named the proposed device “dip-and-fold” due to its how-to-use procedure: first, the device’s bottom edge is dipped in the water sample. The liquid moves up by capillarity towards the detection zone, in which the analytes find the AChE previously immobilized on the paper. The top edge of the paper is covered with a thin parafilm layer to avoid leaking.
After a 10-minute incubation, the AChE is inhibited in the presence of CBs and/or OPs. Then, the device is folded to release both enzymatic substrate acetylthiocholine iodide (ACThI) and indicator 5,5′-Dithiobis (2-nitrobenzoic acid) (DTNB) from a secondary paper to the detection zone for the colorimetric reaction. The detection zone area remains white in the presence of CBs and/or OPs (inhibited AChE); otherwise, it becomes yellow in the absence of these pesticides (active AChE).
It is worthy to emphasize that the dip-and-fold has all required ingredients immobilized onto the paper substrates prior to tests, which excludes the necessity of extra reagents and materials. Besides, the sample can be transferred to the device by capillarity motion when dipping it directly into the contaminated water, eliminating any risk of having contact with the pesticides.

Experiment

The dip-and-fold device was fabricated using two analytical papers and a backing card as a foldable scaffold. Blotting paper (Whatman GB003) was used as the main device’s component for the sample loading by capillarity as well as for the immobilization of 3 µL AChE (100 U/mL), using chitosan 1% (w/v, in acetic acid 2%), as a crosslinker. The secondary component was the filter paper (Whatman grade 4) in which the 3µL of the substrate ACThI (25mM) and 3 µL of DTNB (10mM) were impregnated on the surface. The detection tests were carried out using standard solutions of carbofuran, propoxur (both carbamates) and chlorpyriphos (an organophosphate) at concentrations between 0.1 and 0.0001 mM. Control samples of DI water were also tested. A minimum volume of methanol was required to dissolve the pesticides in water. Yellow-coloured results of the tests were evaluated by taking digital photos of the devices using a smartphone. The RGB channels of images were analyzed using ImageJ software.

Results and Conclusion

The result interpretation was performed by comparing the plotted curves of the blue channel (B) values obtained from the images of each tested device for the different analytes and their respective concentrations. The evaluation of the plotted curves confirmed that the yellow colour's intensity is inversely proportional to the concentration of the analytes in the sample, which means, the more intense the yellow, the lower the concentration of pesticides in the sample. The concentrations applied in this study meet the parameters proposed by the Ontario Drinking Water Quality Standards (ODWQS) and may be compared to other paper-based devices described in the literature.
Both propoxur and carbofuran presented consistent results that were observed by a steady decrease of the B values related to their concentration. In contrast, the chlorpyrifos presented a non-linear curve, which can be attributed to the higher amount of methanol used as a co-solvent to dissolve it in water due to its extremely poor solubility.
The dip-and-fold device presented an exclusive, eco-friendly, and economical testing platform for pesticide detection in water, which offers an attractive solution for water assessment in vulnerable areas exposed to these toxic chemicals.

Figure 1