Development, validation, and implementation of an ultratrace analysis method for the determination of moenomycin A, in aquatic animal products.

Moenomycin A, an antimicrobial growth promoter widely used as an additive in aquaculture feedstuffs, has been restricted for use in the European Union and China due to its potential risk of promoting resistant strains of pathogenic bacteria and causing residues in aquatic animal products. Although methods for analyzing moenomycin A in feedstuffs have been developed, no established method exists for aquatic matrices. In this study, we present, for the first time, a sensitive and validated high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for the determination of moenomycin A in aquatic animal products. Samples were extracted using methanol and purified with the QuEChERS method employing C18 sorbent. The aliquot was dried under a nitrogen stream, reconstituted with methanol-water solvent, and analyzed by HPLC-MS/MS. The developed method exhibited good linearity (r2 > 0.995) over a wide concentration range (1-100 µg/L) and a low limit of detection (1 µg/kg). Average recoveries ranged between 70 and 110% at spiked concentrations of 1, 50, and 100 µg/kg, with associated intra- and inter-day relative standard deviations of 1.25 to 7.32% (n = 6) and 2.91 to 10.08% (n = 3), for different representative aquatic animal production, respectively. To the best of our knowledge, this is the first reported HPLC-MS/MS method for the quantification of moenomycin A in aquatic animal products. The new approach was effectively employed in the analysis of moenomycin A across various aquatic samples.

Identification and Detection of Intracellular Reactive Sulfur Species Using a Reaction-Mediated Dual-Recognition Strategy.

Reactive sulfur species (RSS) are emerging as a potential key gasotransmitter in diverse physiological processes linking two signaling molecules H2S and SO2. However, the exact roles of H2S and SO2 remain unclear. A major hurdle is the shortage of accurate and robust approaches for sensing of H2S and SO2 in biological systems. Herein, we report a reaction-mediated dual-recognition strategy-based nanosensor, silver nanoparticles (AgNPs)-loaded MIL-101 (Fe) (ALM) hybrids, for the simultaneous detection of H2S and SO2 in a living cell. Upon exposure to H2S, AgNPs can be oxidized to form Ag2S, causing a decrease of surface enhanced Raman spectroscopy (SERS) signals of p,p'-dimercaptoazobenzene. Moreover, SO2 reacts with the amino moiety of MIL-101 to form charge-transfer complexes, resulting in an increment of fluorescent (FL) intensity. The ALM with dual-modal signals can simultaneously analyze H2S and SO2 at a concentration as low as 2.8 × 10-6 and 0.003 µM, respectively. Most importantly, the ALM sensing platform enables targeting mitochondria and detection multiple RSS simultaneously in living cells under external stimulation, as well as displays indiscernible crosstalk between SERS and FL signals, which is very beneficial for the comprehension of physiological issues related with RSS.

Dual-Modal Apoptosis Assay Enabling Dynamic Visualization of ATP and Reactive Oxygen Species in Living Cells.

ATP and reactive oxygen species (ROS) are considered significant indicators of cell apoptosis. However, visualizing the interplay between apoptosis-related ATP and ROS is challenging. Herein, we developed a metal-organic framework (MOF)-based nanoprobe for an apoptosis assay using duplex imaging of cellular ATP and ROS. The nanoprobe was fabricated through controlled encapsulation of gold nanorods with a thin zirconium-based MOF layer, followed by modification of the ROS-responsive molecules 2-mercaptohydroquinone and 6-carboxyfluorescein-labeled ATP aptamer. The nanoprobe enables ATP and ROS visualization via fluorescence and surface-enhanced Raman spectroscopy, respectively, avoiding the mutual interference that often occurs in single-mode methods. Moreover, the dual-modal assay effectively showed dynamic imaging of ATP and ROS in cancer cells treated with various drugs, revealing their apoptosis-related pathways and interactions that differ from those under normal conditions. This study provides a method for studying the relationship between energy metabolism and redox homeostasis in cell apoptosis processes.

Tetrahedral DNA-directed core-satellite assembly as SERS sensor for mercury ions at the single-particle level.

To monitor the deteriorating mercury emissions, it is imperative to propose methods for detecting mercury ions (Hg2+) with sensitivity and selectivity. The SERS spectral-resolved single-particle detection approach can be carried out using dark-field optical microscopy (DFM) combined with Raman spectroscopy. Herein, we have designed a novel yet convenient single-particle detection assay for quantifying Hg2+ using DFM-correlated Raman spectroscopy. In the assay, a tetrahedral DNA-directed core-satellite nanostructure is used as the SERS probe. Especially, one edge of the tetrahedron is made up of a single-stranded DNA containing a Hg2+ aptamer, which reconfigures upon the specific recognition of Hg2+. As a result, the interparticle distance reduces from 4.5 to 1.2 nm, thus generating Raman signal enhancement. As a proof of concept, Hg2+ was detected in a linear range from 1 to 100 nM based on the variation in SERS intensity. Furthermore, the experimental results were supported by the finite difference time domain (FDTD) calculations. Owing to its high sensitivity and selectivity, this method was further employed to detect Hg2+ in practical tap water and lake water samples, revealing that the single-particle detection strategy holds great promise for Hg2+ analysis in real environment analysis.

In Situ Monitoring of Hydrogen Peroxide Released from Living Cells Using a ZIF-8-Based Surface-Enhanced Raman Scattering Sensor.

Hydrogen peroxide (H2O2) widely involves in intracellular and intercellular redox signaling pathways, playing a vital role in regulating various physiological events. Nevertheless, current analytical methods for the H2O2 assay are often hindered by relatively long response time, low sensitivity, or self-interference. Herein, a zeolitic imidazolate framework-8 (ZIF-8)-based surface-enhanced Raman scattering (SERS) sensor has been developed to detect H2O2 released from living cells by depositing ZIF-8 over SERS active gold nanoparticles (AuNPs) grafted with H2O2-responsive probe molecules, 2-mercaptohydroquinone. Combining the superior fingerprint identification of SERS and the highly efficient enrichment and selective response of H2O2 by ZIF, the ZIF-8-based SERS sensor exhibits a high anti-interference ability for H2O2 detection, with a limit of detection as low as 0.357 nM. Satisfyingly, owing to the enhanced catalytic activity derived from the successful integration of AuNPs and ZIF, the response time as short as 1 min can be obtained, demonstrating the effectiveness of the SERS sensor for rapid H2O2 detection. Furthermore, the developed SERS sensor enables real-time detection of H2O2 secreted from living cells under phorbol myristate acetate stimulation, as cells can be cultured on-chip. This study will pave the way toward the development of a metal-organic framework-based SERS platform for application in the fields of biosensing and early disease diagnosis associated with H2O2 secretion, thus exhibiting promising potential for future therapies.

Determination of Methylene Blue and Its Metabolite Residues in Aquatic Products by High-Performance Liquid Chromatography-Tandem Mass Spectrometry.

A sensitive and reliable method was developed to determine methylene blue (MB) and its metabolite residues, including azure A (AZA), azure B (AZB), and azure C (AZC) in aquatic products by HPLC-MS/MS. The samples were extracted by acetonitrile and cleaned up by alumina-neutral (ALN) cartridges. The analytes were separated on a Sunfire C18 column (150 mm × 2.1 mm, 5 µm). The method was validated according to the European criteria of Commission Decision 2002/657/CE. Good linearity between 1-500 µg/L was obtained with correlation coefficients (R2) greater than 0.99. The limit of quantification (LOQ) was 1.0 µg/kg. The average recoveries at three levels of each compound (1, 5, and 10 µg/kg) were demonstrated to be in the range of 71.8-97.5%, with relative standard deviations (RSDs) from 1.05% to 8.63%. This method was suitable for the detection of methylene blue and its metabolite residues in aquatic products.

Label-free chlorine and nitrogen-doped fluorescent carbon dots for target imaging of lysosomes in living cells.

Lysosomes with a single-layered membrane structure are mainly involved in the scavenging of foreign substances and play an important role in maintaining normal physiological functions of living cells. In this work, near-neutrally charged fluorescent carbon dots (CDs) were prepared with lipophilicity through a facile one-pot hydrothermal carbonization of chloranil and triethylenetetramine at 160 °C for 3 h. The as-obtained CDs are proved to have good photostability, low cost, and excellent biocompatibility. Importantly, the as-prepared CDs with high quantum yield of 30.8% show excitation-dependent emission with great stability, and thus, they can be well used for the long-term target imaging of lysosomes in living cells without further modification. Meanwhile, the CDs can quickly enter into the lysosomes within 30 min, and the green fluorescence (FL) of CDs reaches the plateau when incubated for 60 min. By comparing the fluorescent intensity, the information about distribution and amount of lysosomes in different cells can be obtained. The proposed CD-based strategy demonstrates great promise for label-free target imaging of lysosomes in living cells. Graphical abstract The near-neutral carbon dots (CDs) with lipophilicity are used as label-free fluorescent nanoprobes for the long-term imaging of lysosomes in living cells.

Loop-mediated isothermal amplification for visual detection of Vibrio parahaemolyticus using gold nanoparticles.

Loop-mediated isothermal amplification (LAMP) eradicates the need of thermocycler in DNA amplification. Signals are usually obtained via fluorometry or turbidimetry, but such methods need improvement in order to become more effortless and reliable. The authors describe a set of six specific primers targeting the species-specific tlh gene of Vibrio parahaemolyticus which were used in accelerated LAMP reaction. Gold nanoparticles (AuNPs) were functionalized with streptavidin (Avidin-AuNPs), and engineered to signal the LAMP reaction. Two of the loop primers for LAMP were biotinylated and then can produce a DNA that can cause clusterization of Avidin-AuNPs based on the formation of avidin-biotin complex. This leads to a color change of the solution from red to blue. Amplification is completed within 30 min and can be visually detected within 5 min. The detection limit of the method is found to be 8.6 cfu per reaction. This visual detection scheme does not require any fluorescent reagents and detection instruments. Conceivably, the method has a wide scope because such Avidin-AuNPs can be used as nanoprobes for a variety of other LAMP products. This rapid and universal strategy holds promise in point of care testing and food testing, particularly in resource-limited regions. Graphical abstract Six specific primers (two of them are biotinylated) were used to realize the accelerated Loop-Mediated Isothermal Amplification. Streptavidin modified gold nanoparticles (Avidin-AuNPs) cluster on the DNA products, leading to the apparent change of color from red to blue, which is readily identified even by unaided eye.

Correction to: Loop-mediated isothermal amplification for visual detection of Vibrio parahaemolyticus using gold nanoparticles.

The published version of this article, unfortunately, contained error. Modifications have been made to the Abstract, Introduction, Results and discussion, and Acknowledgements section. The original article has been corrected.

Improved LC/MS/MS Quantification Using Dual Deuterated Isomers as the Surrogates: A Case Analysis of Enrofloxacin Residue in Aquatic Products.

Extensive and high residue variations in enrofloxacin (ENR) exist in different aquatic products. A novel quantitative method for measuring ENR using high-performance liquid chromatography-tandem mass spectrometry was developed employing enrofloxacin-d5 (ENR-d5) and enrofloxacin-d3 (ENR-d3) as isotope surrogates. This reduced the deviation of detected values, which results from the overpass of the linear range and/or the large difference in the residue between the isotope standard and ENR, from the actual content. Furthermore, high residue levels of ENR can be directly diluted and re-calibrated by the corresponding curve with the addition of high levels of another internal surrogate without repeated sample preparation, avoiding the overflow of the instrument response. The validation results demonstrated that the method can simultaneously determine ENR residues from MQL (2 µg/kg) to 5000 × MQL (method quantification limit) with recoveries between 97.1 and 106%, and intra-precision of no more than 2.14%. This method realized a wide linear calibration range with dual deuterated isomers, which has not been previously reported in the literature. The developed method was successfully applied to the analysis of ENR in different aquatic products, with ENR residue levels varying from 108 to 4340 µg/kg and an interval of precision in the range of 0.175~6.72%. These results demonstrate that batch samples with a high variation in ENR residues (over the linear range with a single isotope standard) can be detected by the dual isotope surrogates method in a single sample preparation process.

Multi-Residue Screening of Pesticides in Aquatic Products Using High-Performance Liquid Chromatography-Tandem High-Resolution Mass Spectrometry.

Pesticide residues in aquatic products are of great concern due to the risk of environmental transmission and their extensive use in aquaculture. In our work, a quick screening approach was developed for the qualitative and semi-quantitative screening of 87 pesticide residues in aquatic products. The sample preparation was investigated, including extract solvent, extract methods, buffer salts, lipid removal, cleanup materials and filter membranes for aquatic products. Samples were extracted using a modified QuEChERS procedure, and two clean-up procedures were developed for UHPLC-Q/Orbitrap MS analysis based on the fat content of the aquatic products. The screening detection limits for all studied pesticides were distributed between 1 and 500 µg/kg in the three representative matrices. Seventy-one pesticides could be analyzed with a screening limit between 1 and 25 µg/kg in grass carp and crayfish, sixty-one pesticides could be screened for limits between 1 and 50 µg/kg in crab. The accuracy results showed that recoveries ranged from 50 to 120% for 60, 56 and 52 pesticides at medium-level for grass carp, crayfish and crab, respectively. At high spiking levels, 74, 65 and 59 pesticides were recovered within the range of 50-120% for the three matrices, respectively. The relative standard deviations of most compounds in different matrices were less than 20%. With this method, the local farmed aquatic products were tested for pesticide residues. In these samples, ethoxyquinoline, prometryn and phoxim were frequently detected. The majority of these confirmed compounds did not exceed 2.00 µg/kg. A grass carp with trichlorfon at 4.87 µg/kg and two carps with ethoxyquinoline at 200 µg/kg were detected, indicating the potential dietary risk.

Rapid Sample Enrichment, Novel Derivatization, and High Sensitivity for Determination of 3-Chloropropane-1,2-diol in Soy Sauce via High-Performance Liquid Chromatography-Tandem Mass Spectrometry.

A novel, simplified derivatization method and a rapid sample preparation process using carbon yarn as a sorbent for the determination of 3-chloropropane-1,2-diol (3-MCPD) in soy sauce via high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was developed. 3-MCPD was first enriched and purified with carbon yarn and then eluted with a methanol-water solution. Subsequently, the analyte underwent derivatization with p-(dimethylamino)-phenol for sensitive detection via HPLC-MS/MS. The limit of detection and the limit of quantitation for 3-MCPD were validated to be 0.5 and 1.0 µg/kg, respectively. Spiking experiments showed recoveries between 83 and 94%, with a relative standard deviation of ≤10%. The method was further validated with a certified reference material. Furthermore, 11 real soy sauce samples from local markets were tested by using this method. These results reveal the widespread 3-MCPD contamination. Consequently, this study offers a preferable alternative for the sensitive, accurate, and precise determination of 3-MCPD in soy sauce.

A dual-responsive nanozyme sensor with ultra-high sensitivity and ultra-low cross-interference towards metabolic biomarker monitoring.

Accurate, sensitive and selective detection of metabolic biomarkers in biofluids are of vital significance for health self-monitoring and chronic disease prevention. Here, for the first time, a smart dual-responsive nanozyme sensor (DNS) was developed for simultaneous analysis of glucose and caffeine utilizing stimuli-responsive yolk-shell gold nanoparticles (GNPs)-embedded MIL-53 (Al) (GNPs@MIL-53) structures. After the introduction of glucose, GNPs@MIL-53 displays excellent glucose oxidase (GOx)-like activity to induce the conversion of glucose to gluconic acid and H2O2. H2O2 can oxidize 3,3',5,5'-tetramethylbenzidine (TMB) with the generation a bright-blue color, enabling in-field visualization and surface enhanced Raman scattering (SERS) detection of glucose. Upon the addition of caffeine, 2-aminoterephthalic acid modified MIL-53 can react with the caffeine to form intermolecular hydrogen-bonded complexes, leading to strong cyan fluorescence and significant Raman enhancements. The DNS with multi-channel signal outputs can simultaneously determine glucose and caffeine at concentrations of as low as 3 × 10-8 M and 1.2 × 10-11 M, respectively. Importantly, the DNS-based analytical system not only enables visual discrimination and accurate assay of glucose and caffeine in biofluids, but also exhibits negligible cross-interference between glucose and caffeine determination. The combined characteristics of high selectivity, enhanced accuracy and superior quantitative performance make our platform suitable for the point-of-care monitoring of chronic-disease-related metabolic biomarkers.

1,4-Benzenedithiol-Bridged Nanogap-Based Individual Particle Surface-Enhanced Raman Spectroscopy Mechanical Probe for Revealing the Endocytic Force.

1,4-Benzenedithiol (BDT)-bridged core-satellite assemblies, as surface-enhanced Raman spectroscopy (SERS) mechanical probes, can be employed for real-time monitoring of the dynamics of endocytic forces and the accompanying trajectory of nanoparticles during the endocytosis process. These mechanical probes exhibit good responses in terms of SERS intensity ratios while undergoing mechanical pressure. Force tracing and the accompanying trajectory of nanoparticles are resolved accurately to render the endocytosis process in live cells. Density functional theory simulation results further proved the sensing scheme due to the electrons transforming between BDT and gold nanoparticles. Furthermore, this SERS mechanical probe is a valid method to visualize endocytic forces at multiple locations and establish a direct criterion to discriminate between cancer cells and normal cells.

Ratiometric Fluorescence Immunoassay Based on Carbon Quantum Dots for Sensitive Detection of Malachite Green in Fish.

Malachite green (MG) is a synthetic poisonous organic compound that has been banned in many countries as a veterinary drug for aquaculture. An efficient, fast and sensitive method is urgently needed for monitoring the illegal use of malachite green (MG) in aquaculture. In this study, a novel ratiometric fluorescence immunoassay was established. Nitrogen-doped carbon quantum dots were used as ratiometric fluorescent probes with a fluorescence peak at 450 nm. Horseradish peroxidase was employed to convert o-phenylenediamine to 2,3-diaminophenazine, with a new fluorescence peak at 580 nm and a strong absorption at 420 nm. The inner filter effect between N-CQD fluorescence and DAP absorption was identified. It allows for the ratiometric detection of MG using a fluorescent immunoassay. The results demonstrated a linear ratiometric fluorescence response for MG between 0.1 and 12.8 ng·mL-1. The limit of detection of this method was verified to be 0.097 µg·kg-1 with recoveries ranging from 81.88 to 108%, and the relative standard deviations were below 3%. Furthermore, this method exhibited acceptable consistency with the LC-MS/MS results when applied for MG screening in real samples. These results demonstrated a promising application of this novel ratiometric fluorescence immunoassay for MG screening with the merits of rapid detection, simple sample preparation, and stable signal readout. It can be an alternative to other traditional methods if there are difficulties in the availability of expensive instruments, and achieve comparable results or even more sensitivity than other reported methods.

Simple and rapid detection of free 3-monochloropropane-1,2-diol based on cysteine modified silver nanoparticles.

A rapid colorimetric method using cysteine-modified silver nanoparticles (Cys-AgNPs) is applied for the detection of 3-monochloropropane-1,2-diol (3-MCPD). Indeed, in the presence of 3-MCPD, the color of Cys-AgNPs solution changes from yellow to pink within five minutes at 100 °C and pH 9.3. This change is mainly attributed to the ability of amino group of cysteine to react with 3-MCPD to form N-(2,3-dihydroxypropyl)-amino acid grafted on AgNPs (3-MCPD-Cys-AgNPs) in alkaline medium. This color change makes 3-MCPD to be clearly detectable by unassisted visual means even at 0.1 µg⋅mL-1. Besides, using UV-Vis spectroscopic technique, a linear range from 0.1 µg⋅mL-1 to 1.25 µg⋅mL-1 for 3-MCPD detection is obtained, with a calculated detection limit of 0.084 µg⋅mL-1. These results suggest that this sensing technique is sensitive to 3-MCPD and may have a substantial application in the rapid detection of food contaminants particularly, where quality and safety of food products are paramount concern.

Green preparation of carbon quantum dots with wolfberry as on-off-on nanosensors for the detection of Fe3+ and l-ascorbic acid.

A green and facile hydrothermal synthesis approach is proposed for the preparation of nitrogen-doped carbon quantum dots (N-CQDs) with wolfberry. These N-CQDs were developed as a highly sensitive fluorescent 'on-off-on' switch sensor for the sensing of Fe3+ and l-ascorbic acid (AA). The N-CQDs displayed superior fluorescence characteristics of CQDs with a quantum yield up to 22%. The N-CQDs were demonstrated to selectively react with Fe3+, leading to fluorescence quenching effect, which was successfully used for the detection of Fe3+ with a limit of detection at 3 µmoL•L-1. The addition of AA is supposed to repair the surface defects, and result in the fluorescence recovery. Based on this effect, the strategy of 'on-off-on' detection of AA was established with a limit of detection at 1.8 µmoL•L-1. Furthermore, the practical application of the detection of Fe3+ lake water and AA in medical tablet was demonstrated, promising an effective and efficient 'on-off-on' nanosensor with low-cost, green synthesis for Fe3+ and AA detection.

An Effective and Efficient Sample Preparation Method for 2-Methyl-Isoborneol and Geosmin in Fish and Their Analysis by Gas Chromatography-Mass Spectrometry.

The intensive aquaculture strategy and recirculating aquaculture system often lead to the production of off-flavor compounds such as 2-methyl-isoborneol (2-MIB) and Geosmin (GSM). The regular purge and trap extraction followed by analysis with gas chromatography-mass spectrometry (GC-MS) usually involve a complicated assembly of facilities, more working space, long sample preparation time, and headspace solid-phase microextraction (SPME). In this work, a method with easier sample preparation, fewer and simplified facilities, and without SPME on GC-MS analysis is developed for the determination of 2-MIB and GSM in fish samples. Unlike previous methods, solvent extract from samples, QuEChERS-based cleanup, and solid-phase extraction for concentration are applied. The LOD (S/N > 3) and LOQ (S/N > 10) of this method were validated at 0.6 µg/kg and 1.0 µg/kg for both 2-MIB and GSM, which are under the sensory limit (1 µg/kg). Application of this method for incurred fish samples demonstrated acceptable analytical performance. This method is suitable for large-scale determination of 2-MIB and GSM in fish samples, owing to the use of simple facility and easy-to-operate procedure, rapid sample preparation, and shorter time for GC-MS analysis without SPME.

Electrochemically renewable SERS sensor: A new platform for the detection of metabolites involved in peroxide production.

The accurate detection of hydrogen peroxide (H2O2)-involved metabolites plays a significant role in the early diagnosis of metabolism-associated diseases, whereas most of current metabolite-sensing systems are often hindered by low sensitivity, interference of coexisting species, or tedious preparation. Herein, an electrochemistry-regenerated surface-enhanced Raman scattering (SERS) sensor was developed to serve as a universal platform for detecting H2O2-involved metabolites. The SERS sensor was constructed by modifying newly synthesized 2-mercaptohydroquinone (2-MHQ) molecules on the surface of gold nanoparticles (AuNPs) that were electrochemically predeposited on an ITO electrode. Metabolites were detected through the changes in the SERS spectrum as a result of the reaction of 2-MHQ with H2O2 induced by the metabolites. Combining the superiority of SERS fingerprint identification and the specificity of the related enzymatic reactions producing H2O2, the designed SERS sensor was highly selective in detecting glucose and uric acid as models of H2O2-involved metabolite with limits of detection (LODs) of 0.159 µM and 0.0857 µM, respectively. Moreover, the sensor maintained a high SERS activity even after more than 10 electrochemical regenerations within 2 min, demonstrating its effectiveness for the rapid detection of various metabolites with electrochemistry-driven regulation. Importantly, the presented SERS sensor showed considerable practicability for the detection of metabolites in real serum samples. Accordingly, the SERS sensor is a new detection platform for H2O2-involved metabolites detection in biological fluids, which may aid the early diagnosis of metabolism-related diseases.

Sensitive and selective SERS probe for detecting the activity of γ-glutamyl transpeptidase in serum.

γ-Glutamyl transpeptidase (GGT) has attracted considerable attention for its regulatory effect on glutathione metabolism in living organisms; further, its close relationship with physiological dysfunctions such as hepatitis and liver cancers has enhanced its applicability. Therefore, the accurate detection of GGT levels is particularly important for the early diagnosis of diseases. Thus, we herein report the development of a surface-enhanced Raman spectroscopic (SERS) probe, namely bis-s,s'-((s)-4,4'-thiolphenylamide-Glu) (b-(s)-TPA-Glu), that comprises of a γ-glutamyl moiety for detection of the GGT activity. In this system, detection was achieved by observing differences in the SERS spectral profiles of the b-(s)-TPA-Glu probe and its corresponding hydrolysis product that resulted from the catalytic action of GGT. This SERS probe system exhibited a high selectivity toward GGT due to a combination of its specific catalytic action and the distinctive spectroscopic fingerprint of the SERS technique. The developed SERS approach was also found to be approximately linear in the range of 0.2-200 U/L, and a limit of detection of 0.09 U/L was determined. Furthermore, the proposed SERS method was suitable for detection of the GGT activity of clinical serum samples and also for evaluation of the inhibitors of GGT. Consequently, this approach is considered to be a promising diagnostic and drug screening tool for GGT-associated diseases.