Low LOD Values Research Articles

Analytical quality-by-design guided development of a novel and robust HPTLC method for quantifying plumbagin from Plumbago species

The study utilizes an analytical quality-by-design (AQbD) approach to establish a robust and efficient HPTLC method for plumbagin quantification in plumbago species. The highest-risk critical method parameters (CMP; concentration of plumbago and composition of mobile phase) and most appropriate critical analytical attributes (CAA; Peak area and retention factor [Rf]) were employed in the AQbD methodology. These CMP were tuned for higher plumbagin recovery using the central composite design (CCD) experiment. The created approach was validated by the requirements outlined in the International Conference on Harmonization (ICH) Q2 (R2). Every plate involved in the proposed method validation has been subjected to the passing requirements of the system suitability test (SST). The low LOD (16.93 ng/mL) and LOQ (49.68 ng/mL) values and the minimal divergence in intra- and inter-day studies validate the developed method’s high precision and sensitivity. Further experiments employing the proposed method to determine the amount of plumbagin from the two distinct plant sources harvested under two different climate conditions determined the highest plumbagin content in P. indica, which is in line with the results. To summarize, the AQbD method may be used in other plant systems to optimize various factors and get a greater concentration of the desired plant metabolite.

Development and validation of a UV spectrophotometric method for simultaneous estimation of teneligliptin hydrobromide hydrate and pioglitazone hydrochloride in pharmaceutical dosage form

BackgroundThis study presents the development and validation of a UV spectrophotometric method for the simultaneous estimation of teneligliptin hydrobromide hydrate (TEN) and pioglitazone hydrochloride (PIO) in pharmaceutical dosage forms. The method measures absorbance at 243 nm for TEN and 265 nm for PIO. Validation was conducted according to ICH guidelines, covering specificity, linearity, range, precision, accuracy, limit of detection, limit of quantification, and robustness.ResultsThe method exhibited linearity in the 2–25 µg/ml concentration range for both TEN and PIO, with correlation coefficients close to 1. Precision studies showed low % RSD values, indicating excellent repeatability and minimal intra- and inter-day variability. Accuracy was confirmed through recovery studies, with results within the 98–102% range. The method demonstrated sensitivity with low LOD and LOQ values. Robustness testing showed stability and consistency under varying conditions. Analysis of tablet formulations confirmed the accurate quantification of both drugs.ConclusionThe developed UV spectrophotometric method offers a simple, sensitive, and cost-effective solution for the simultaneous estimation of TEN and PIO in pharmaceutical formulations. It is suitable for routine analysis and quality assessment of tablet dosage forms.

Blue light emitting graphene quantum dots/ Rhodamine B doped gold nanostars for ratiometric detection of methotrexate

In this work, an innovative ratiometric sensing platform was developed for the determination of methotrexate (MTX), an antifolate drug, a chemotherapy agent, and an immune system suppressant based on blue emission graphene quantum dots/Rhodamine B doped gold nanostars (B-GQDs/Au NSt-RB). The developed sensor was a dual-emission fluorescent probe with two major emission peaks at 440 nm (B-GQDs) and 580 nm (Au NSt-RB) by exciting at 330 nm. Based on the inhibiting effect of MTX on the system's fluorescence density, the stable ratiometric fluorescent probe was used for the rapid determination of MTX in aquatic solutions and spiked human serum samples. The results indicated good linear correlations over the logarithmic concentration range of 0.3 nM–50.0 μM. In addition, B-GQDs/Au NSt-RB can further realize highly sensitive detection of MTX with a low LOD value of 2.28 × 10−10 M. The RSD% values obtained for the intra-day and inter-day precision were 0.63–3.86 %. With recoveries of 98.2–100.1 % and 98.7–100.5 %, respectively. The short-term temperature and freeze-thaw tests confirmed the higher stability of the developed sensor. In addition, the calculated recoveries for MTX recognition in real samples were in the range of 98–102 %. These findings suggested the excellent potential of the ratiometric fluorescence B-GQDs/Au NSt-RB sensor for detecting MTX in real plasma samples.

Development of an RP-HPLC Method for Quantifying Diclofenac Diethylamine, Methyl Salicylate, and Capsaicin in Pharmaceutical Formulation and Skin Samples.

An RP-HPLC method with a UV detector was developed for the simultaneous quantification of diclofenac diethylamine, methyl salicylate, and capsaicin in a pharmaceutical formulation and rabbit skin samples. The separation was achieved using a Thermo Scientific ACCLAIMTM 120 C18 column (Waltham, MA, USA, 4.6 mm × 150 mm, 5 µm). The optimized elution phase consisted of deionized water adjusted to pH = 3 using phosphoric acid mixed with acetonitrile in a 35:65% (v/v) ratio with isocratic elution. The flow rate was set at 0.7 mL/min, and the detection was performed at 205 nm and 25 °C. The method exhibits good linearity for capsaicin (0.05-70.0 µg/mL), methyl salicylate (0.05-100.0 µg/mL), and diclofenac diethylamine (0.05-100.0 µg/mL), with low LOD values (0.0249, 0.0271, and 0.0038 for capsaicin, methyl salicylate, and diclofenac diethylamine, respectively). The RSD% values were below 3.0%, indicating good precision. The overall greenness score of the method was 0.61, reflecting its environmentally friendly nature. The developed RP-HPLC method was successfully applied to analyze Omni Hot Gel® pharmaceutical formulation and rabbit skin permeation samples.

A ratiometric fluorescent probe with 1,2,4-triazole recognition unit for fluoride ion detection based on synchronous fluorescence spectroscopy

As an important inorganic chemical with wide applications and high toxicity, the detection of fluoride-containing products and environmental samples has gained the most interest. In this work, a ratiometric fluorescent probe (NIN2) is constructed with a triazole group as the new recognition site for the detection of F−. The synchronous fluorescence technology was introduced to obtain the well discrimination of two emission bands. The value of wavelength intervals (Δλ = λem. - λex.) is optimized to be set as 70 nm with the minimized overlap of two emission bands centered around 380 nm and 470 nm, respectively. In this way, NIN2 can detect F− in the region of 0–0.2 μM with a good linear relationship (R2 = 0.9989) and low LOD value (36 nM). Only F− can induce obvious ratiometric response (I470nm/I380nm) from 0.2 to 3.4 while the ratios of the samples containing other cations and anions are as low as the blank sample. For investigating the recognition mechanism of triazole, 1H, 13C and 19F NMR spectral technologies were used to verify the interaction between triazole and F−. Furthermore, the DFT calculation was also performed to discuss the electronic transition of NIN2 before and after interaction with F−. We believe this work provides a new recognition moiety for detecting F−.

Uporedno spektrofotometrijsko određivanje 3-hidroksiflavona bazirano na kompleksima cinka i aluminijuma i njihovi antioksidatni profili

Flavonoids, as plant-derived compounds, were essential active components in traditional medicine for centuries. Their potential or confirmed effects include antiviral, antimutagenic, antiinflammatory, antibacterial, vasodilatory, and anticancer properties. The promotion of a plantbased diet, along with the benefits of consuming flavonoids, has recently become increasingly attractive. 3-Hydroxyflavone (3HF) is the structural spine of flavonols, an important subgroup of flavonoids. Although 3HF itself does not exist in plants per se, it exerts many of its effects because of its characteristics that allow it to prevent free radical generation. This work is focused on the characterization of 3HF complexes with zinc(II) and aluminium(III) ions (Zn-3HF and Al-3HF, respectively). Besides this, a simple, fast, and low-priced spectrophotometric method for 3HF determination, with very low LOD and LOQ, based on Zn-3HF and Al-3HF formation, was established. A slight advantage is given to the modification with Al3+ ion on pH 4.91, due to very low LOD and LOQ values of 1.83×10-7 molL-1 , and 5.50×10-7 molL-1 , respectively, and a high correlation coefficient, R =0.99986. Furthermore, the antioxidant ability of Zn-3HF, Al-3HF, and parent 3HF was examined by the ABTS and DPPH tests. They brought the Zn-3HF complex to the fore as a potential antioxidative agent.

Development and validation of a green RP-HPLC method for detection and quantitation of meropenem trihydrate in the bulk: A comparison with HPTLC method

This research introduces an eco-friendly green Reverse-Phase High Performance Liquid Chromatography (RP-HPLC) method for detecting and quantifying meropenem trihydrate. Traditional methods use buffered solutions, gradient mobile phases, and longer retention times, but this method offers a rapid RP-HPLC technique with a C8 column and isocratic mobile phase (60% methanol, 40% ultra-pure water), eliminating the need for buffers and acetonitrile. It features a short 2.1-minute retention time and a high r² value of 0.9995. It delivers accuracy (101.1-102.3%) and precision (RSD ≤ 2%), coupled with low LOD and LOQ values of 1.72 and 5.20 µg/ml. Aqueous dilution simplifies sample preparation, reducing degradation and interference. The method is compared with an HPTLC method, showing an extended linear range (6.25-200 µg/ml for HPLC, 7.81-62.5 µg/ml for TLC) and high sensitivity, making it significant for meropenem trihydrate quality control in bulk and dosage forms.

Dual-channel fluorescent probe for discriminative detection of H2S and N2H4: Exploring sensing mechanism and real-time applications

A highly efficient system incorporates the real-time visualization of the two toxic molecules (H2S and N2H4) and the recognition of corresponding transforms using a fluorescent sensor. In this paper, a dual-responsive probe (QS-DNP) based on methylquinolinium-salicyaldehyde-2,4-dinitrophenyl was developed that can simultaneously detect H2S and N2H4 at two independent fluorescent channels without signal crosstalk. QS-DNP showed excellent anti-interference, high selectivity, outstanding water solubility, low LOD values (H2S: 51 nM; N2H4: 40 nM), low cytotoxicity, and mitochondrial localization properties. The 2,4-dinitrophenyl site was sensitive to H2S, and the CC bridge was reactive to N2H4, with strong fluorescence at 680 and 488 nm, respectively. The wavelength gap between these two channels is 192 nm; verify that there is no signal crosstalk throughout detection. By this means, the probe was used to simultaneously detect H2S and N2H4 in real soil samples, food samples, and living cells. The endogenous H2S and N2H4 were monitored in HeLa cells and investigated the mitochondria organelle of living cells with a positive charge on QS-DNP. Overall, all results emphasize that the QS-DNP probe is a powerful tool for the simultaneous detection of H2S and N2H4 and presents a potential new sensing approach.

Investigations on the electrochemical behavior of sunitinib and metabolites N-desethyl-sunitinib and sunitinib-N-oxide and its selective determination using molecularly imprinted polymer-based sensor

Sunitinib (SUN) is an orally administered tyrosine kinase inhibitor and is mainly metabolized via N-deethylation to form the primary active metabolite N-desethyl-sunitinib (M1) and via oxidation to sunitinib-N-oxide (M2). In this study, we first investigated the electrochemical behavior of sunitinib and its N-desethyl and N-oxide metabolites on a bare glassy carbon electrode (GCE) by cyclic voltammetry (CV) technique. Based on the results, a plausible electrooxidation mechanism is proposed, yielding the oxo-desfluoro-sunitinib (quinoneimine metabolite) via oxidative defluorination of sunitinib. After this phase of the study, due to the low sensitivity and selectivity results obtained from the bare GCE, we aimed to develop the first molecularly imprinted polymer (MIP)-based electrochemical sensor for the sensitive and selective determination of SUN. The MA@SDS/MIP-GCE sensor was prepared by thermal polymerization of methacrylic acid (MA) as a functional monomer in the presence of SUN on a GCE surface under the optimized parameters such as template/monomer ratio, drop amount, polymerization temperature and time, removal solution and time, and rebinding time, which affect the structure and response of the sensor. The surface characterization of the developed sensor was performed by scanning electron microscopy (SEM), and the electrochemical characterization was performed by CV and electrochemical impedance spectroscopy (EIS). The electroanalytical performance of the MA@SDS/MIP-GCE sensor was tested by differential pulse voltammetry (DPV) under the optimized conditions in comparison with the NIP-based sensor. The linearity of the developed sensor for the determination of SUN was investigated in the concentration range of 0.5 – 5.0 nM for standard and serum solutions with low LOD and LOQ values. The application of the MIP sensor for the determination of SUN in pharmaceutical dosage form (capsule) and serum samples was demonstrated with excellent recoveries. The results showed that the MA@SDS/MIP-GCE sensor is a sensitive, specific, selective, rapid, and economical option for the quantitative determination of SUN.

A pyridoxal based bio-compatible fluorometric chemosensor for recognition of Zn2+ ions: Theoretical approaches and application in live cell imaging

A simple pyridoxal-based chemosensor (HL = 4-(((furan-2-ylmethyl)imino)methyl)-5-(hydroxymethyl)-2-methylpyridin-3-ol) was synthesized which selectively detects Zn2+ ions in HEPES buffer at pH = 7.4 (MeOH:H2O, 9:1, (v/v)) among other competitive metal ions. Around 10-fold increase in fluorescence intensity was seen at 472 nm after excition at 390 nm. HL displays blue fluorescence with a large stoke′s shift (∼80 nm) in presence of Zn2+ ions. A 1:1 binding stoichiometry between HL and metal ions (Zn2+) and the PET-OFF-CHEF-ON mechanism had been established by different spectroscopic techniques and theoretical studies. Low LOD value of the chemosensor towards Zn2+ (lower than the WHO's guideline of 76.5 µM) and strong binding constant (K = 1.54 × 105 M−1) suggest the applicability of HL towards biological cell imaging studies under physiological conditions. MTT assay was carried out to evaluate the toxicity of probe HL up to maximum concentrations with PC3 cell line.

Innovative UV Protocols Based on Straightforward Mathematical Filtration for Concurrent Estimation of Two Antidiabetic Drugs in Their Brand-New Combination: A Comparative Study.

In 2019, the U.S. Food and Drug Administration approved a brand-new combination of linagliptin and empagliflozin in a formulation called Glyxambi® tablets for managing type 2 diabetes mellitus. Nowadays, spectrophotometric techniques occupy the first place among their peers in terms of ease of application, friendliness to the environment, and low costs. This research discusses the development of two very simple spectrophotometric protocols based on zero-order spectra for the determination of linagliptin and empagliflozin. The developed protocols were the induced dual-wavelength and absorption correction protocols. Linagliptin could be determined directly at 305 nm, at which the empagliflozin spectrum was zero-crossing. Empagliflozin was determined using the two developed protocols. The induced dual-wavelength technique was developed by calculating the equality factor of linagliptin to cancel its interference. The absorption correction technique was developed by measuring the correction absorption factor. The concentration ranges of linagliptin and empagliflozin were 1-10 µg/mL and 3-30 µg/mL, respectively. Excellent recovery results were found in bulk, dosage form, and synthetic mixtures. Low LOD and LOQ values were obtained, indicating the high sensitivity of the protocols. The statistical Student's t-test was performed to compare the results of the applied and reported protocols, indicating no difference between them. The proposed protocols have the advantages of being straightforward, affordable, and requiring no sophisticated manipulations, just simple mathematical calculations. The proposed protocols are acceptable for routine usage in QC laboratories and in future research applications. Two novel univariate methods were developed for quantitative analysis of linagliptin and empagliflozin in their pharmaceutical and laboratory mixtures, and produced satisfactory results.

Repurposing of waste ammonium sulfate as S,N-doped carbon quantum dots: A sensitive and selective fluorescent probe for the determination of tetracycline

BackgroundCurrently, many types of pharmaceuticals are used to treat disease and bacterial infections. Tetracycline (TC) is a widely applied human and veterinary medicine. However, the widespread usage and lack of awareness in the management of TC lead to its unavoidable discharge into the environment. The consequences include the proliferation of resistant strains of bacteria and adverse impacts on nontarget organisms and microbial ecosystems, which means that TC is an emerging substance of concern and that it is crucial to develop determination platforms with cost-effective and fast-response features. MethodsCarbon quantum dots (CQDs) have received tremendous attention owing to their great performance in fluorescent probing. In the present work, ammonium sulfate-doped carbon quantum dots (S,NCQDs) are fabricated through a facile hydrothermal synthesis pathway. Significant FindingsThe results illustrate that the S,NCQDs exhibit sensitive determination ability toward tetracycline via a static quenching mechanism and possess several superior features, including a rapid response, good sensitivity and stability, broad linear detection range (1–100 μM), and low LOD value (0.1203 μM), when applied in the determination of TC. Additionally, the application of S,NCQDs valorizes ammonium sulfate waste from semiconductor manufactory waste and has great potential in fluorescence sensing.

A novel thiazole-based covalent organic framework as turn-off fluorescent sensor for detection of aniline and triethylamine in liquid and vapor phase

Due to the diverse chemical structures and varying basicity of amines, the development of rapid, portable, and reversible methods for the selective recognition of trace organic amines in the environment remains a challenge. Herein, a novel recyclable and selective turn-off fluorescent sensor for organic amines detection based on covalent organic framework (COF) was developed. As-synthesized COF, TAPB-BTDD exhibits good crystallinity, high chemical stability and strong fluorescence, which can selectively recognize triethylamine (TEA) and aniline in both liquid and vapor phase. The COF material demonstrates impressively low LOD values for aniline and triethylamine in solution, estimated to be 27 nM and 170 nM, respectively. Moreover, when detecting aniline and triethylamine vapors, the calculated LOD values are 20.1 ppb and 106.4 ppb, respectively. In addition, TAPB-BTDD showed basically unchanged emission intensities and rapid response to the amines after ten recycling tests, suggesting that the COF has high stable recycle repeatability. The combination of experimental analysis and density functional theory (DFT) calculations demonstrates that the unique fluorescence-quenching response of TAPB-BTDA can be attributed to the photoinduced electron transfer (PET) between the amines and the COF. This COF-based sensor offers a promising solution for the accurate and reliable detection of organic amines in various environmental settings.

Green and fast ultrasound-assisted extraction procedures for Fe, Mn, Mg and Ca analysis in cane syrups by FAAS

New methodologies for the analysis of Fe, Mn, Mg, and Ca content in cane syrups using FAAS combined with ultrasound-assisted extraction (UAE) were developed and validated. For multivariate optimization of UAE, a simplex-centroid mixture design and individual desirability functions were applied to achieve the best composition of extraction solvents (HNO3, HCl, and water). Under optimal conditions of ultrasound-assisted extraction (100 W, 37 kHz, 10 min, 5.0 mL of extraction solvent (ultrapure water for Fe and Mn and HNO3:H2O (75:25%, v/v), corresponding to a 1.19 mol L−1 HNO3 solution for Mg and Ca)), method validation showed good linearity, confirmed by regression analysis, and low LOD and LOQ values of 0.266–0.284 μg g−1 and 0.114–0.861 μg g−1. Estimates of repeatability and mean precision (RSD = 0.44–2.11%), Horrat values (0.07–0.26), and assessment of trueness by comparing sample preparation methods using a paired t-test demonstrated the excellent precision and trueness of the developed methods. The methods were applied to twenty-five cane syrup samples, and the results of the multivariate analysis showed that the metal ions can be used as indicators of geographical origin or contamination during manufacturing or processing, thus helping to ensure the food quality and safety of this sweetener.

Validated ion exchange HPLC method for the quantification of levothyroxine – a narrow therapeutic index drug – used for the treatment of hypothyroidism

Drugs with narrow therapeutic index (NTI-drugs) have been defined by the FDA as drugs with small differences between therapeutic and toxic doses that might lead to serious therapeutic failures or life-threatening adverse drug reactions. Levothyroxine sodium pentahydrate (LT4), a synthetic T4 hormone used for the treatment of hypothyroidism (a condition where there is a hormonal imbalance in the thyroid gland that is responsible for the regulation of several physiological, metabolic, cardiovascular, and neurological processes). LT4 is designated by the FDA as a narrow therapeutic index drug and is available in the market in the form of very low dose pharmaceutical formulations ranging from 25 mcg to 150 mcg. This requires that the pharmaceutical dosage form should contain the exact labeled amount of the active ingredient, LT4, such that safety and efficacy are maintained. Therefore, it is necessary to develop an a precise, accurate and sensitive analytical method for LT4 quantification considering the treatment doses being in micrograms. In the present work, an ion exchange HPLC method has been developed and validated for the determination of LT4 as per ICH guidelines. The developed method was found to be simple, sepcific, precise and accurate. The low LOD and LOQ values allowed the quantitfication of the active ingredient in different pharmaceutical products qualifying the method to be applied in quality control assays.

Tryptanthrin-malononitrile adduct for highly efficient turn-off fluorescent sensing of aniline

The fluorescence quenching of 2-(8-chloro-12-oxo-12H-indolo[2,1-b]quinazolin-6-ylidene)-malononitrile (5a) by aniline was studied in detail. Time correlated single photon counting measurements (TCSPC) indicated that the quenching involved both static and dynamic mechanisms without ground state complex formation. The selectivity towards aniline sensing in presence of other amines/aromatics and the anti-interference studies together with the low LOD value indicates the potential of 5a as a molecular probe for aniline sensing which is unaffected by pH alteration as well.

A reliable ratiometric fluorescence sensing of heparin and its antidote based on cationic carbon quantum dots and acid red 87

A reliable, selective, and sensitive ratiometric fluorescence sensor was fabricated for the detection of protamine (PROT) and heparin (HEPR). It is based on a rational combination of Acid Red 87 (AR 87) as an anionic dye and polyethyleneimine modified carbon quantum dots (PEI@CQDs) as cationic fluorescent nanoparticles via electrostatic interaction. Addition of positively charged PROT quenched the negatively charged AR 87 via static quenching; while, the addition of HEPR restored the fluorescence of AR 87 via the formation of PROT-HEPR complex. Simultaneously, the fluorescence emission of PEI@CQDs remained constant. Thus, a ratiometric response (FAR 87/FPEI@CQDs) was linear with the concentration of PROT and HEPR in the range of 0.006–20 µg mL−1 and 0.018–22 µg mL−1 with detection limits (S/N) of 1.7 ng mL−1 and 6.0 ng mL−1, respectively. The analytical sensing platform offers several advantages including good selectivity, low LOD values, low-cost, and fast-response. Importantly, the sensing platform was used to detect PROT and HEPR in human serum and urine samples with satisfactorily recoveries % and RSDs %.

Determination of 27 Glucocorticoids in Urine by Ultra-High-Performance Liquid Chromatography-Tandem Mass Spectrometry Using UniSprayTM Source

Glucocorticoids (GCs) are a group of the most important and commonly used anti-inflammatory, antiallergenic, and immunosuppressive drugs. Like narcotics, they can be addictive if taken at increasing doses to achieve greater analgesic effects. The purpose of the study was to develop initial and confirmation testing analytical methods that would allow for the identification of glucocorticoid class substances in human urine to be used for routine analyses for the purposes of prosecution in the case of abuse, for clinical toxicology, medical jurisprudence, as well as for routine testing of athletes for the presence of prohibited substances in sports by means of ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) using a new generation ionization source UniSprayTM (U.S). This new method allows for the simultaneous detection of 27 glucocorticoids in human urine using LC-MS/MS. The tests conducted yielded relatively low LOD and LOQ values, ranging from 0.06 ng/mL to 0.14 ng/mL and 0.75 ng/mL for LOD and LOQ, respectively.

Highly Sensitive Multiplex Detection of Molecular Biomarkers Using Hybridization Chain Reaction in an Encoded Particle Microfluidic Platform

In the continuous combat against diseases, there is the need for tools that enable an improved diagnostic efficiency towards higher information density combined with reduced time-to-result and cost. Here, a novel fully integrated microfluidic platform, the Evalution™, is evaluated as a potential solution to this need. Encoded microparticles combined with channel-based microfluidics allow a fast, sensitive and simultaneous detection of several disease-related biomarkers. Since the binary code is represented by physically present holes, 210 different codes can be created that will not be altered by light or chemically induced degradation. Exploiting the unique features of this multiplex platform, hybridization chain reaction (HCR) is explored as a generic approach to reach the desired sensitivity. Compared to a non-amplified reference system, the sensitivity was drastically improved by a factor of 104, down to low fM LOD values. Depending on the HCR duration, the assay can be tuned for sensitivity or total assay time, as desired. The huge potential of this strategy was further demonstrated by the successful detection of a multiplex panel of six different nucleic acid targets including viruses and bacteria. The ability to not only discriminate these two categories but, with the same effort, also virus strains (human adenovirus and human bocavirus), virus subtypes (human adenovirus type B and D) and antibiotic-resistant bacteria (Streptococcus pneumonia), exemplifies the specificity of the developed approach. The effective, yet highly simplified, isothermal and protein-enzyme-free signal amplification tool reaches an LOD ranging from as low as 33 ± 4 to 151 ± 12 fM for the different targets. Moreover, direct detection in a clinically relevant sample matrix was verified, resulting in a detection limit of 309 ± 80 fM, approximating the low fM levels detectable with the gold standard analysis method, PCR, without the drawbacks related to protein enzymes, thermal cycling and elaborate sample preparation steps. The reported strategy can be directly transferred as a generic approach for the sensitive and specific detection of various target molecules in multiplex. In combination with the high-throughput capacity and reduced reagent consumption, the Evalution™ demonstrates immense potential in the next generation of diagnostic tools towards more personalized medicine.

An ultrasensitive "mix-and-detect" kind of fluorescent biosensor for malaoxon detection using the AChE-ATCh-Ag-GO system.

Malaoxon, a highly toxic metabolite of malathion, can lead to severe harm or death if ingested. This study introduces a rapid and innovative fluorescent biosensor that relies on acetylcholinesterase (AChE) inhibition for detecting malaoxon using Ag-GO nanohybrid. The synthesized nanomaterials (GO, Ag-GO) were evaluated with multiple characterization methods to confirm their elemental composition, morphology, and crystalline structure. The fabricated biosensor works by utilizing AChE to catalyze the substrate acetylthiocholine (ATCh), which generates positively charged thiocholine (TCh) and triggers citrate-coated AgNP aggregation on the GO sheet, leading to an increase in fluorescence emission at 423 nm. However, the presence of malaoxon inhibits the AChE action and reduces the production of TCh, resulting in a decrease in fluorescence emission intensity. This mechanism allows the biosensor to detect a wide range of malaoxon concentrations with excellent linearity and low LOD and LOQ values of 0.001 pM to 1000 pM, 0.9 fM, and 3 fM, respectively. The biosensor also demonstrated superior inhibitory efficacy towards malaoxon compared to other OP pesticides, indicating its resistance to external influences. In practical sample testing, the biosensor displayed recoveries of over 98% with extremely low RSD% values. Based on the results obtained from the study, it can be concluded that the developed biosensor has the potential to be used in various real-world applications for detecting malaoxon in food, and water samples, with high sensitivity, accuracy, and reliability.