TY - JOUR
T1 - User-friendly one-step disposable signal-on bioassay for glyphosate detection in water samples
AU - Delprat, N.
AU - Martins, L. O.
AU - Blum, L. J.
AU - Aymard, C. M.G.
AU - Leca-Bouvier, B.
AU - Octobre, G.
AU - Doumèche, B.
N1 - Funding Information:
The French Ministry of Education, Research and Technology is gratefully acknowledged for fellowship of one of us (N.D.). Mailis Dissais and Nathan Montmailler are kindly acknowledged for practical help.
Publisher Copyright:
© 2023 Elsevier B.V.
PY - 2023/12/1
Y1 - 2023/12/1
N2 - The onsite detection of glyphosate requires an easy-to-handle, low-cost and disposable assay for untrained users as requested by the ASSURED guidelines. A new strategy based on the expression of fusion proteins is proposed here. A glyphosate oxidase derived from Bacillus subtilis and the 6E10 variant of the dye peroxidase from Pseudomonas putida, both fused with the carbohydrate binding module (CBM) 3a from Clostridium thermocellum, were designed and expressed, leading to GlyphOx-CBM and 6E10-CBM. Cell lysates were used to immobilise both enzymes on cotton buds' heads without any purification. The cotton buds exhibit glyphosate oxidase activity when dipped into a glyphosate-contaminated water sample containing the 6E10-CBM chromogenic substrates. The chromophore could be quantified both in the solution and on the cotton buds’ heads. Photography followed by image analysis allows to detect glyphosate with a linear range of 0.25–2.5 mM and a limit of detection (LoD) of 0.12 mM. When the chromogenic substrates are replaced by luminol, the chemiluminescence reaction allows the detection of glyphosate with a linear range of 2–500 μM and a LoD of 0.45 μM. No interference was observed using glyphosate analogues (glycine, sarcosine, aminomethylphosphonic acid) or other herbicides used in a mixture. Only cysteine was found to inhibit 6E10-CBM. Two river waters spiked with glyphosate lead to recoveries of 64–131%. This work describes a very easy-to-handle and inexpensive signal-on bioassay for glyphosate detection in real surface water samples.
AB - The onsite detection of glyphosate requires an easy-to-handle, low-cost and disposable assay for untrained users as requested by the ASSURED guidelines. A new strategy based on the expression of fusion proteins is proposed here. A glyphosate oxidase derived from Bacillus subtilis and the 6E10 variant of the dye peroxidase from Pseudomonas putida, both fused with the carbohydrate binding module (CBM) 3a from Clostridium thermocellum, were designed and expressed, leading to GlyphOx-CBM and 6E10-CBM. Cell lysates were used to immobilise both enzymes on cotton buds' heads without any purification. The cotton buds exhibit glyphosate oxidase activity when dipped into a glyphosate-contaminated water sample containing the 6E10-CBM chromogenic substrates. The chromophore could be quantified both in the solution and on the cotton buds’ heads. Photography followed by image analysis allows to detect glyphosate with a linear range of 0.25–2.5 mM and a limit of detection (LoD) of 0.12 mM. When the chromogenic substrates are replaced by luminol, the chemiluminescence reaction allows the detection of glyphosate with a linear range of 2–500 μM and a LoD of 0.45 μM. No interference was observed using glyphosate analogues (glycine, sarcosine, aminomethylphosphonic acid) or other herbicides used in a mixture. Only cysteine was found to inhibit 6E10-CBM. Two river waters spiked with glyphosate lead to recoveries of 64–131%. This work describes a very easy-to-handle and inexpensive signal-on bioassay for glyphosate detection in real surface water samples.
UR - http://www.scopus.com/inward/record.url?scp=85171379728&partnerID=8YFLogxK
U2 - 10.1016/j.bios.2023.115689
DO - 10.1016/j.bios.2023.115689
M3 - Article
C2 - 37716158
AN - SCOPUS:85171379728
SN - 0956-5663
VL - 241
JO - Biosensors and Bioelectronics
JF - Biosensors and Bioelectronics
M1 - 115689
ER -