A Simple and Efficient Colorimetric Detection of Creatinine Based on Citrate-Stabilized Gold Nanoparticles

A Simple and Efficient Colorimetric Detection of Creatinine Based on Citrate-Stabilized Gold Nanoparticles | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article A Simple and Efficient Colorimetric Detection of Creatinine Based on Citrate-Stabilized Gold Nanoparticles Xianfa Lv, Tongrui Shi, Xia Bai, Zheng Guan, Rujian Jiang, Lu Zhou, and 1 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4785879/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 27 Aug, 2024 Read the published version in Plasmonics → Version 1 posted 7 You are reading this latest preprint version Abstract Creatinine level is a crucial indicator in the clinical assessment and diagnosis of renal diseases, achieving simple and accurate detection of urinary creatinine levels in resource-limited point-of-care settings is of great significant in the timely prevention and diagnosis of kidney diseases. As a popular zero-dimensional material, gold nanoparticles (AuNPs) exhibit intriguing optical properties and thus have become a promising material for many sensing detection applications. Here, we proposed a simple, efficient and sensitive quantitative detection of creatinine by studying the relative absorbance (ΔA) of AuNPs in absence and presence of creatinine. The method relies on the aggregation of AuNPs via ligand-exchanged of citrate ions and creatinine on the surface of AuNPs to achieve colorimetric detection. With this assay, the limit of detection for creatinine was as low as 0.16 mM, and the dynamic detection range was 0.5 to 20 mM under optimized conditions. In our experiments, the specificity of proposed method was investigated and successfully applied to detect creatinine in urine sample. It reveals that the proposed colorimetric protocol has demonstrated a high sensitivity and selectivity for creatinine, and has a potential practicability in clinical diagnostics. citrate-stabilized gold nanoparticles (AuNPs) creatinine colorimetric method relative absorbance (ΔA) Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 1. Introduction Kidney disease has been one of the leading causes of death worldwide [ 1 ], but less than 10% of patients with early renal dysfunction are diagnosed and treated in a timely manner [ 2 ]. Therefore, it is important to timely identify and diagnose kidney diseases, especially in resource-limited settings. Creatinine is a nitrogenous breakdown product produced during the metabolism of creatine and creatine phosphate in the muscle tissue [ 3 ], and the normal kidney function filters and excretes creatinine with the urine [ 4 ]. The normal urinary creatinine levels are in the range of 0.3 to 3.1 g/L (2.4 to 27.0 mM) [ 5 ], but vary depending on age and gender [ 6 ]. Changes in kidney function and creatinine metabolism can significantly affect urine-creatinine levels. Creatinine levels that deviate from the normal range can indicate various kidney diseases such as chronic kidney diseases (CKDs) and acute kidney injury (AKI) [ 7 , 8 ]. Thus, creatinine levels are considered to be one of the most effective indicators for evaluating renal function and preventing kidney diseases. To date, various methods are available for the determination of creatinine levels, including high performance liquid chromatography [ 9 ], nuclear magnetic resonance [ 10 ], liquid chromatography-isotope dilution mass spectrometry [ 11 ], etc. Although these methods can offer sensitive and accurate measurement, most of them are time-consuming and expensive as well as require sophisticated instruments and/or trained staff [ 12 ]. Therefore, it is essential to develop a simple and cost-effective method for the detection of creatinine. Generally, colorimetric assays have been studied as an ideal candidate for detection of creatinine, such as Jaffe and enzymatic methods [ 13 ]. The Jaffe method is based on the reaction of creatinine with alkaline picrate to produce an orange-red complex that can be quantitatively monitored by spectrophotometry [ 14 – 16 ]. The enzymatic method depends on the hydrolysis of creatinine by creatinase [ 13 , 17 ]. The Jaffe method is more susceptible interferences from some organic compounds (such as ascorbic acid, glucose, bilirubin and certain proteins), which can lead to false positive results [ 18 ]. Whereas, the enzymatic method exhibits better anti-interference ability but is more expensive than Jaffe method, and sources of key enzyme is difficult to obtain [ 19 ]. To overcome the limitations of traditional methods, different types of sensors have been developed, such as surface-enhanced Raman spectroscopy [ 20 ], nanotechnology-based sensors [ 21 ], electrochemical biosensors [ 22 ], etc. Among them, noble metal (Au, Ag and Cu) nanomaterial-based colorimetric sensing displays high sensitivity due to its unique localized surface plasmon resonance (LSPR) effect [ 23 – 25 ]. Typically, the LSPR effect of the metallic nanoparticles is dependent on the size, shape, and distance between nanoparticles, particularly the gold nanoparticles (AuNPs) with high extinction coefficients (10 8 ~ 10 10 M − 1 · cm − 1 ) [ 26 ]. Specifically, according to the Mie theory, when AuNPs are in the strong coupling plasmonic regime (low interparticle distance), a red-shift and an amplification of the LSPR as the interparticle distance decreases are observed [ 27 ]. That is, when the analyte induces the aggregation of AuNPs, the LSPR peak and colour of AuNPs colloid can undergo remarkable changes, thereby achieving colorimetric detection of the analyte. For example, Yi He et al. demonstrated a colorimetric method through studying the relationship between the red-shift values of AuNPs LSPR peak before and after introducing the creatinine [ 28 ]. Sittiwong et al. further refined the pretreatment of creatinine in urine samples by extracting creatinine from urine samples using sulfonic acid-functionalized silica gel, and then determining creatinine based on the changes in LSPR signals before and after creatinine-induced aggregation of AuNPs [ 29 ]. Joseph et al. developed a smartphone-based sensor kit for monitoring creatinine through polyol functionalized AuNPs [ 30 ]. The currently reported plasmonic-based colorimetric sensing assays provide an effective research tool for accurate and sensitive monitoring creatinine levels. However, the difficulty in accurately controlling the extent of creatinine-induced aggregation of AuNPs, and the complex procedures for functionalizing AuNPs and extracting creatinine limit the utility of the method developed assays in point-of-care conditions. Therefore, there is a pressing need for a simple and practical method to quantify creatinine in biological fluids. In this work, we developed a simple method for creatinine detection by studying the relative absorbance (ΔA) of citrate-stabilized AuNPs in absence and presence of creatinine. The optimized experimental parameters such as the pH values, the AuNPs concentrations and the reaction time were investigated. Under the optimized conditions, the AuNPs-based colorimetric determination of creatinine showed a better specificity in the concentration range from 0.5 to 20 mM with the limit of detection of 0.16 mM. Furthermore, the proposed protocol has been successfully used to detect the creatinine level in urine. It demonstrates that the colorimetric detection based on citrate-stabilized AuNPs provides a facile route for the high sensitivity and specificity of creatinine in clinical diagnosis. 2. Experimental Section 2.1 Chemicals Hydrogen tetrachloroaurate (III) trihydrate (HAuCl 4 ·3H 2 O) and artificial urine solutions were purchased from Sinopharm Chemical Reagent Co., Ltd. Trisodium citrate (Na 3 C 6 H 5 O 7 ·2H 2 O), hydrochloric acid (HCl), nitric acid (HNO 3 ), sodium hydroxide (NaOH), boric acid (H 3 BO 3 ), acetic acid (CH 3 COOH), phosphoric acid (H 3 PO 4 ), creatinine (C 4 H 7 N 3 O), urea (CH 4 N 2 O) and uric acid (N 5 H 4 N 4 O 3 ) were obtained from Aladdin (Shanghai, China). Britton-Robinson (B-R) buffer solutions were prepared by H 3 BO 3 , CH 3 COOH and H 3 PO 4 solutions all at a concentration of 4 mM, and NaOH solution (0.2 mol/L) was used to adjust the different pH values. Different concentrations of creatinine solution were prepared in the B-R buffer solution (44 mM, pH 7.0). All reagents were analytical grade and used as received, and Millipore ultrapure double deionized water (18 MΩ) was used for all solution preparations. All glassware was cleaned by configured aqua regia (3:1; HCl/HNO 3 ) for 15 min, then rinsed thoroughly with distilled water several times and dried in a hot air oven at 85˚C. 2.2 Instruments Ultraviolet-Visible (UV-Vis) absorption spectra were monitored with a spectrophotometer (Shimadzu UV-2450, Japan). Mean hydrodynamic diameter of AuNPs were determined by dynamic light scattering (DLS) with Malvern laser particle size analyser (C-W10X10XB22, China). The morphology of AuNPs was characterized by using a high-resolution field emission scanning electron microscope (SEM, JEOL JSM-7900F, Japan). 2.3 Preparation of the Citrate-Stabilized AuNPs The synthesis of AuNPs colloid was according to the classical sodium citrate reduction method [ 31 ]. Briefly, 4 mL of sodium citrate (1%) was quickly added to 96 mL HAuCl 4 (0.007%) boiling solution under vigorous stirring until the colour of mixture solution change to red. After that, the reaction mixture was stirred and refluxed for an additional 15 min, and then cooled to room temperature. The AuNPs colloid was successfully prepared and stored in a clean tube at 4°C for further use. According to the Beer-Lambert law, the concentration of AuNPs colloid was estimated to be 1.97 nM using an extinction coefficient of 2.7×10 8 M − 1 · cm − 1 at 521 nm [ 32 ]. 2.4 Detection of Creatinine Firstly, 2.5 mL AuNPs colloid dispersed in 2.5 mL B-R buffer solution was respectively added to each of the six test tubes and incubated for 15 min at room temperature. Secondly, 60 µL of the newly prepared creatinine solution with different concentrations (0.5 mM, 1 mM, 5 mM, 10 mM, 15 mM, and 20 mM) were respectively added to the above AuNPs solutions. After incubating at room temperature for 233 min, the mixture solution was subjected to measure UV-Vis absorption spectrum, and the visual observation photographs were taken at the same time. 2.5 Creatinine Detection in Urine Sample To prevent the influence of proteins that may be present in the urine, the artificial urine samples were filtered through a 0.22 µm membrane and then diluted 100 times with B-R buffer solution for immediate testing. Subsequently, 60 µL of creatinine solution (0.5 mM, 10 mM, 20 mM) prepared with the diluted artificial urine was added to the AuNPs solution, respectively. Then, the detection of creatinine was performed as described above. 3. Results and Discussion 3.1 Working Principle of Creatinine Detection Figure 1 a shows the UV-Vis absorption spectra of citrate-stabilized AuNPs colloid in the wavelength range of 400–800 nm before and after addition of creatinine at room temperature. The AuNPs colloid prepared by the citrate reduction method displays characteristic LSPR band located at 521 nm. After the addition of creatinine, the LSPR peak of AuNPs at 521 nm was weakened and slightly red-shifted to 525 nm. And it was noted that a new absorption band emerged at 663 nm due to the aggregation of AuNPs induced by linkage with creatinine, and meanwhile a red-to-blue colour changes could be observed (Fig. 1 a, inset). It is demonstrated that the introduction of creatinine can induce the aggregation of AuNPs, thereby triggering the colour change of AuNPs colloid. Moreover, to further confirm that the introduction of creatinine could cause the AuNPs aggregation, the particle size distribution and morphology of AuNPs before and after introduction of creatinine were performed by using DLS and SEM (Fig. 1 b- 1 d), respectively. The hydrodynamic diameter distribution of the citrate-stabilized AuNPs in the absence and presence of creatinine were illustrated in Fig. 1 b. The AuNPs without creatinine show the mono-dispersity with an average hydrodynamic diameter of 29 nm. Upon the addition of creatinine, the aggregated AuNPs exhibit two major peaks at 37 nm and 251 nm. The former peak corresponds to the average size of citrate-stabilized AuNPs with a slight increase in size, which is consistent with the slightly red-shifted UV-Vis spectrum. The latter peak was attributed to the aggregated AuNPs, whose particle size increase significantly. In addition, the SEM im-ages of AuNPs before and after incubation with creatinine are shown in Fig. 1 c and 1 d. It can be seen that the citrate-stabilized AuNPs were uniformly distributed with an average particle size of 23 ± 2 nm by using Nano Measurer, according to the SEM images (Fig. 1 c). After incubation with creatinine, significant aggregation of AuNPs is observed in Fig. 1 d. These results suggested that creatinine can induce the aggregation of citrate-stabilized AuNPs. To explore the origins of the AuNPs aggregation induce by creatinine, the surface charge and structural properties of AuNPs before and after introduction of creatinine was analysed. The principle of creatinine detection using citrate-stabilized AuNPs is illustrated in Fig. 2 . Before introducing of creatinine, the structure of citrate-stabilized AuNP consists a central carboxylate group interacting with AuNP surface and the remaining two terminal carboxylate group (see Fig. 2 a) [ 33 ]. Under neutral conditions (B-R buffer solution, pH 7.0), the electrostatic repulsion between the negatively charged carboxylate group of citrate anions adsorbed on AuNPs maintains the homogeneously dispersion of the AuNPs. It has been reported that the creatinine has two tautomeric (amino tautomer and imino tautomer) and can be interconverted between two tautomeric under neutral condition (see Fig. 2 b) [ 5 , 12 , 34 ]. After addition of creatinine, the citrate adsorbed on the AuNPs can be replaced by the creatinine ligands, due to the coordinated interaction between the nitrogen containing functional group (creatinine) and AuNPs is much stronger than that of citrate ions [ 28 , 35 , 36 ]. After ligand-exchanged, creatinine and its two tautomeric are neutral charged at pH 7.0, which greatly reduces the number of surface negative charge of AuNPs, resulting in the aggregation of AuNPs (see Fig. 2 a and 2 c). Besides, creatinine and its two neutral tautomers can disrupt the electrostatic balance of citrate-stabilized AuNPs by forming hydrogen bonds with the exposed carboxylate groups of the citrate molecule, thereby causing the aggregation of AuNPs (see Fig. 2 c) [ 12 ]. Thus, creatinine could induce the aggregation of AuNPs and accompanying a colour change through the synergistic effect of ligand-exchanged and hydrogen bonds with the citrate ions adsorbed on the AuNPs, thereby achieving colorimetric detection. 3.2 Optimization of Detection Conditions 3.2.1 Effect of pH The pH value of AuNPs colloid is a crucial factor affecting the sensitivity and accuracy of creatinine detection. Here, the media samples were prepared using B-R buffer solutions with pH values of 4.0 to 10.0, and AuNPs colloid dispersed in 5 mL B-R buffer solution at different pH values. To evaluate the influence of pH on the stability of AuNPs colloid, we studied the effect of pH value of AuNPs over the absorbance (Fig. 3 a). With the increasing of pH value from 4.0 to 7.0, the absorbance at 521 nm progressively increases (Fig. 3 b), accompanied by a change in colour from purple to red (Fig. 3 c). This is because at pH < 7, citrate is fully protonated and the number of surface negative charge is greatly reduced, resulting in the aggregation of AuNPs [ 12 , 37 ]. While, with the increasing of pH value from 7.0 to 10.0, the absorbance at 521 nm progressively decreases (Fig. 3 b), accompanied by a change in colour from red to purple (Fig. 3 c). It can be interpreted that the two free terminal carboxylic acid groups of citrates adsorbed on the surface of AuNPs can be induced to bind to the surface of AuNPs through an increase of pH [ 38 ], leading to a decrease in surface negative charge and thus causing the aggregation of AuNPs. At neutral pH, the citrate is deprotonated, the negative surface charge result in electrostatic repulsion between AuNPs and maintained the red wine colour indicating no aggregation took place. It was noticed that a new absorption band in the longwave direction appeared at higher or lower pH due to the aggregation of AuNPs, accompanied by a visible colour change from red to purple. Therefore, pH value has a significant impact on the stability of AuNPs, especially at higher or lower pH value. We choose pH 7.0 for the following experiments. 3.2.2 Effect of AuNPs concentration The AuNPs concentrations are another significant factor affecting the limit of detection and sensitivity of creatinine. Here, different concentrations of AuNPs (0.59 to 1.37 nM) were prepared using B-R buffer solution (pH 7.0) at room temperature. The absorbance of various concentrations of AuNPs before and after the addition of creatinine was determined and shown in Fig. 4 a. As anticipated, with the increasing of AuNPs concentrations from 0.59 to 1.37 nM, the absorbance at 521 nm in the absence and presence of creatinine gradually increases (Fig. 4 a). While, with the increasing of AuNPs concentrations from 0.59 to 0.98 nM, the ΔA at peak at 521 nm progressively increases and then decreases from AuNPs concentrations of 0.98 to 1.37 nM (Fig. 4 b). It can be interpreted that higher AuNPs concentrations require more creatinine to compete with citrate ions for binding sites, and a small amount of creatinine cannot effectively induce the aggregation of higher concentrations of AuNPs, resulting in an insignificant change in ΔA at 521 nm after the addition of creatinine [ 39 ]. Therefore, AuNPs concentration of 0.98 nM was chosen for the following experiments. 3.2.3 Effect of reaction time Figure 5 a displays the UV-Vis spectra of AuNPs in the presence of 0.5 mM creatinine at different incubation time. With the increasing of incubation time, the absorption band at 521 nm decreased, and remained unchanged until about 103 min, but meanwhile, the new peak at 650 nm red-shifted and increased. To determine the effect of creatinine concentration on reaction time, we also studied the UV-Vis absorption spectra of AuNPs in the presence of 10 mM and 20 mM creatinine with different reaction times (Fig. 5 b and 5 c). Similarly, upon reaction time, the absorption band at 521 nm decreased, and remain unchanged at about 125 min and 135 min, respectively. And meanwhile the absorption band at 650 nm red-shifted and increased. Figure 5 d shows the relative absorbance as a function of the incubation time in the presence of different concentrations of creatinine. The time constant τ for AuNPs reaction with creatinine of different concentration was further derived by using the exponential decay formula \(\:y={y}_{0}+A\text{*}{e}^{-x/t}\) . From our study, it can be found that τ is 14 ± 1 min, 24 ± 4 min and 24 ± 2 min for the concentration of 0.5 mM, 10 mM and 20 mM creatinine, respectively. The reaction time when the reaction reached equilibrium were calculated from the first order reaction kinetic equation to be 133 ± 13 min, 233 ± 13 min and 226 ± 40 min, separately. As consequence, the optimal incubation time was selected to be 233 min. 3.3 Detection of Creatinine Under the optimized conditions, the colorimetric determination of creatinine was performed. As the creatinine concentration increased, the colour of AuNPs solution gradually changed from red to purple and then to blue (Fig. 6 a), which could be visible to naked eyes above 0.5 mM. Thus, it can be visually detected by the naked eye when the creatinine concentration is higher than 0.5 mM. Furthermore, the colour change of AuNPs in presence of creatinine was quantitatively determined by using UV-Vis spectrophotometry. Figure 6 b shows the UV-Vis spectra of AuNPs with different concentrations of creatinine. As the creatinine concentration increased, the absorption bands at 521 nm progressively decreased and red-shifted to 525 nm and 529 nm, while the absorption bands of 600 to 700 nm emerged and gradually increased. The differences in absorbance at 521 nm of AuNPs with and without creatinine was used to compare the aggregation of AuNPs caused by different concentrations of creatinine. Figure 6 c shows the relationship between ΔA and creatinine concentration. The corresponding standard dose-response curve was a linear relationship described by the fitting curve \(\:\text{y}=0.00746\text{x}+0.0519\) (see Fig. 6 c), and the limit of detection (LOD) of creatinine was determined to be 0.16 mM (S/N = 3), which was calculated via the linear regression method (LOD = 3σ/slope) [ 40 , 41 ]. The reproducibility of this method was further evaluated by a series of six parallel experiments of three different concentrations of creatinine (0.5 mM, 10 mM, 20 mM), corresponding to a relative standard deviation (RSD) of 3.4%, 1.8% and 4.8%, respectively. Therefore, our colorimetric methods based on citrate-stabilized AuNPs has high sensitivity and better reproducibility to reliably detect the trace creatinine level. Table 1 presents the summary of certain current methods for creatinine determination and their detection capabilities. It can be found that the LOD of our method is lower than that of the commonly used Jaffe-based method in the clinic, indicating that the established method can meet the requirements for clinical detection. Compared with some of other AuNPs-based creatinine assays, our method has a wider linear range or higher linear correlation, and allows for creatinine detection without requiring functionalized modification of AuNPs or pretreatment of creatinine. Thus, our suggested AuNPs-based colorimetric detection of creatinine strategy has a LOD equivalent to or lower than that of common methods, and our strategy is remarkable for its simplicity and effectivity. Table 1 Comparison of the present method with some previously reported methods for urinary creatinine detection Methods Limit of detection Analytical range (mM) R 2 Reference Smartphone-based method 0.084 µM 0.001–0.05 0.975 [ 42 ] Electrochemical method 0.04 mM 0–1 0.98 [ 43 ] AuNPs aggregation-based colorimetric method 0.08 mM 0.1–20 0.99 [ 28 ] Jaffe-based method 0.72 mM 0–6 0.991 [ 44 ] Polyol functionalized AuNPs methods 0.7 nM 0-0.002 0.994 [ 30 ] Amperometric method 0.21 mM 0.5-2 - [ 45 ] Solid phase extraction method 0.12 mM 0.13–0.35 0.998 [ 29 ] Monodisperse AuNPs-based method 0.16 mM 0.5–20 0.999 This work 3.4 Assay Specificity The presence of inorganic ions and other organic compounds in the urine sample may lead to a false-positive of creatinine estimation. In order to verify the specific recognition of creatinine in urine samples by the proposed sensor, some potential interfering substances, including glucose (Glu), glycine (Gly), ascorbic acid (AA), urea, uric acid (UA), sodium chloride (NaCl), ammonium chloride (NH 4 Cl), potassium phosphate (K 3 PO 4 ), sodium sulfate (Na 2 SO 4 ) and magnesium chloride (MgCl 2 ) were detected under the same optimum conditions. Figure 7 shows the ΔA for interfering substances (10 mM) and creatinine (1 mM). Obviously, the ΔA in presence of the above interfering substances is much smaller than that of creatinine, indicating that these substances did not interfere with the recognition and detection of creatinine. The observed high selectivity in this case was attributed to the stronger affinity between the three nitrogen atoms of creatinine and AuNPs within the concentration range of this method under the specified experimental conditions. Moreover, the pKa of uric acid and glycine were 5.5 and 2.3, respectively, which were lower than the pH of medium in this assay (7.0). Therefore, uric acid and glycine were negatively charged. When they were adsorbed onto the surface of AuNPs through nitrogen atoms, they did not disrupt the electrostatic equilibrium of AuNPs particles. Consequently, the proposed sensor can be used to specifically detect creatinine. 3.5 Detection of Creatinine in artificial urine Samples To evaluate the practical application, different amounts of creatinine were added into the diluted artificial urine samples (1:100) and then detected by the proposed sensor. As shown in Fig. 8 , it can be found that the ΔA of AuNPs is increased with increasing the concentrations of creatinine, which is similar to the situation in buffer solution. It is indicated that our proposed method can successfully detect the creatinine level in urine samples. Table 2 specifically presents the comparison of the detection results of creatinine in B-R buffer solution and diluted artificial urine. As can be seen in Table 2 , the amount of creatinine added and the assay values were comparable in both buffer solution and diluted urine, respectively, indicating the high accuracy of our method. Compared with detection results of creatinine in buffer solution, we performed the same six sets of parallel measurements of 0.5 mM, 10 mM and 20 mM creatinine in diluted urine, and the corresponding relative standard deviations were 8.4%, 1.6% and 4.3%, respectively. It demonstrated that our method has good reproducibility even in diluted urine. Moreover, the spiked recoveries of 0.5 mM, 10 mM and 20 mM creatinine in diluted urine were calculated as 110%, 103.4% and 105.3%, respectively, indicating the high reliability of our method. Therefore, our method is suitable for the detection of creatinine in artificial urine. Table 2 Comparison of the sensing results of creatinine in B-R solution and in diluted artificial urine Sample Added (mM) Detected (mM) Recovery (%) RSD (n = 6, %) Creatinine in buffer 0.5 0.48 96 3.4 10 9.73 97.3 1.8 20 19.58 97.9 4.8 Creatinine in urine 0.5 0.55 110 8.4 10 10.34 103.4 1.6 20 21.06 105.3 4.3 4. Conclusions In summary, we proposed a strategy by studying the relative absorbance of citrate-stabilized AuNPs in absence and presence of creatinine to quantitively detect urine creatinine level. The experimental conditions that may affect the sensitivity and accuracy of creatinine detection, including the pH, the concentration of AuNPs and reaction time, were optimized. Subsequently, under optimal conditions, the relative absorbance of citrate-stabilized AuNPs with creatinine concentration range of 0.5 to 20 mM were obtained, and the LOD was calculated to be 0.16 mM. Moreover, the control experiments for detection of inorganic ions and other organic compounds in the urine sample exhibited that the proposed methods had superior selectivity for the detection of creatinine. Furthermore, the method was successfully applied to the detection of creatinine level in spiked artificial urine. The proposed method based on the relative absorbance of citrate-stabilized AuNPs has a great potential for detecting creatinine in resource-limited point-of-care settings sites. Declarations Competing Interests The authors declare no competing interests. Funding This work is supported by the National Science Foundation of China (No. 52305317), Natural Science Foundation of Shandong Province (Nos. ZR2022QB040, ZR2022QH006 and ZR20230B113). Author Contribution “H.C. and L.Z. conceived the idea of the work. X.L., T.S., X.B. and Z.G. performed design, material preparation, data collection, and analysis. X.L. wrote the original manuscript text. 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Adv Funct Mater 30: 2005400. https://doi.org/10.1002/adfm.202005400 He Y, Zhang XH, Yu HL (2015) Gold nanoparticles-based colorimetric and visual creatinine assay Microchim Acta 182: 2037-2043. https://doi.org/10.1007/s00604-015-1546-0 Sittiwong J, Unob F (2015) Detection of urinary creatinine using gold nanoparticles after solid phase extraction. Spectroc Acta Pt A-Molec Biomolec Spectr 138: 381-386. https://doi.org/10.1016/j.saa.2014.11.080 Mercy JSI, Vasimalai N (2023) Point-of-care sensor kit for on-site monitoring of creatinine in clinical samples using polyol functionalized gold nanoparticles by UV–visible spectrophotometry. Microchem J 193: 109173. https://doi.org/10.1016/j.microc.2023.109173 Sangwan S, Seth R (2021) Synthesis, characterization and stability of gold nanoparticles (AuNPs) in different buffer systems. 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Sens Actuator B-Chem 255: 2779-2784. https://doi.org/10.1016/j.snb.2017.09.092 Liu GZ, Luais E, Gooding JJ (2011) The fabrication of stable gold nanoparticle-modified interfaces for electrochemistry. Langmuir 27: 4176-4183. https://doi.org/10.1021/la104373v El Badawy AM, Luxton TP, Silva RG, Scheckel KG, Suidan MT, Tolaymat TM (2010) Impact of environmental conditions (pH, ionic strength, and electrolyte type) on the surface charge and aggregation of silver nanoparticles suspensions. Environ Sci Technol 44: 1260-1266. https://doi.org/10.1021/es902240k Park JW, Shumaker-Parry JS (2014) Structural study of citrate layers on gold nanoparticles: Role of intermolecular interactions in stabilizing nanoparticles. J Am Chem Soc 136: 1907-1921. https://doi.org/10.1021/ja4097384 Liu QJ, Han P, Gong WW, Wang H, Feng XY (2018) Colorimetric determination of the pesticide chlorothalonil based on the aggregation of gold nanoparticles. Microchim Acta 185: 354. https://doi.org/10.1007/s00604-018-2890-7 Gegenschatz SA, Chiappini FA, Teglia CM, Pena AMD, Goicoechea HC (2022) Binding the gap between experiments, statistics, and method comparison: A tutorial for computing limits of detection and quantification in univariate calibration for complex samples. Anal. Chim. Acta 1209: 339342. https://doi.org/10.1016/j.aca.2021.339342 Li DX, Wang S, Wang L, Zhang H, Hu JD (2019) A simple colorimetric probe based on anti-aggregation of AuNPs for rapid and sensitive detection of malathion in environmental samples. Anal Bioanal Chem 411: 2645-2652. https://doi.org/10.1007/s00216-019-01703-7 Zhang Q, Yang R, Liu G, Jiang SY, Wang JR, Lin JQ, Wang TY, Wang J, Huang ZF (2024) Smartphone-based low-cost and rapid quantitative detection of urinary creatinine with the Tyndall effect. Methods 221: 12-17. https://doi.org/10.1016/j.ymeth.2023.11.011 Wei F, Cheng S, Korin Y, Reed EF, Gjertson D, Ho CM, Gritsch HA, Veale J (84) Serum creatinine detection by a conducting-polymer-based electrochemical sensor to identify allograft dysfunction. Anal Chem 84: 7933-7937. https://doi.org/10.1021/ac3016888 Randviir EP, Kampouris DK, Banks CE (2013) An improved electrochemical creatinine detection method via a Jaffe-based procedure. Analyst 138: 6565-6572. https://doi.org/10.1039/c3an01431b Matsui R, Sakaki T, Osakai T (2012) Amperometric determination of creatinine with a dialysis membrane‐covered nitrobenzene/water interface for urine analysis. Electroanalysis 24: 2325-2331. https://doi.org/10.1002/elan.201200497 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Published Journal Publication published 27 Aug, 2024 Read the published version in Plasmonics → Version 1 posted Editorial decision: Revision requested 07 Aug, 2024 Reviews received at journal 30 Jul, 2024 Reviewers agreed at journal 26 Jul, 2024 Reviewers invited by journal 24 Jul, 2024 Editor assigned by journal 24 Jul, 2024 Submission checks completed at journal 24 Jul, 2024 First submitted to journal 23 Jul, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4785879","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":337181080,"identity":"85eb3dee-2609-42bf-ad30-d6b4c7f66e6f","order_by":0,"name":"Xianfa Lv","email":"","orcid":"","institution":"School of Chemistry and Pharmaceutical Engineering, Shandong First Medical University \u0026 Shandong Academy of Medical Sciences, Taian 271000, Shandong, China","correspondingAuthor":false,"prefix":"","firstName":"Xianfa","middleName":"","lastName":"Lv","suffix":""},{"id":337181082,"identity":"3551c861-c49b-49ba-8d1d-6dede84498f1","order_by":1,"name":"Tongrui Shi","email":"","orcid":"","institution":"School of Chemistry and Pharmaceutical Engineering, Shandong First Medical University \u0026 Shandong Academy of Medical Sciences, Taian 271000, Shandong, China","correspondingAuthor":false,"prefix":"","firstName":"Tongrui","middleName":"","lastName":"Shi","suffix":""},{"id":337181083,"identity":"cd69cbd5-0f53-4424-ad4f-5732bd5ac1de","order_by":2,"name":"Xia Bai","email":"","orcid":"","institution":"School of Chemistry and Pharmaceutical Engineering, Shandong First Medical University \u0026 Shandong Academy of Medical Sciences, Taian 271000, Shandong, China","correspondingAuthor":false,"prefix":"","firstName":"Xia","middleName":"","lastName":"Bai","suffix":""},{"id":337181085,"identity":"c7527a1a-34b7-4172-bc89-cc0958cacb35","order_by":3,"name":"Zheng Guan","email":"","orcid":"","institution":"School of Chemistry and Pharmaceutical Engineering, Shandong First Medical University \u0026 Shandong Academy of Medical Sciences, Taian 271000, Shandong, China","correspondingAuthor":false,"prefix":"","firstName":"Zheng","middleName":"","lastName":"Guan","suffix":""},{"id":337181086,"identity":"7e63b42c-2adc-4deb-b18d-da9b33748a0f","order_by":4,"name":"Rujian Jiang","email":"","orcid":"","institution":"School of Chemistry and Pharmaceutical Engineering, Shandong First Medical University \u0026 Shandong Academy of Medical Sciences, Taian 271000, Shandong, China","correspondingAuthor":false,"prefix":"","firstName":"Rujian","middleName":"","lastName":"Jiang","suffix":""},{"id":337181087,"identity":"0202d6e6-d9ec-45ac-b7be-77be1b563ef3","order_by":5,"name":"Lu Zhou","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAyUlEQVRIiWNgGAWjYBACPgYGNiBlw8Bw+ACIz0xYCxtESxoDw7EE0rQcJkWLRPqzBx/bztvzHeNOk2CosE5sYD97gICWhHTDmW23E2ce490mwXAmPbGBJy+BkJZj0rxttxMM7vduk2BsO5zYIMFjQEBLYhtQyzl7A5AtjP+I0pLMBtRygHEDWEsDMVp4nrFJzjiXDPLLZouEY+nGbTw5+LXws6c/k/hQZgcMMd6NNz7UWMv2s5/BrwUVJDBAomkUjIJRMApGAYUAACNfP8DQ58SsAAAAAElFTkSuQmCC","orcid":"","institution":"School of Chemistry and Pharmaceutical Engineering, Shandong First Medical University \u0026 Shandong Academy of Medical Sciences, Taian 271000, Shandong, China","correspondingAuthor":true,"prefix":"","firstName":"Lu","middleName":"","lastName":"Zhou","suffix":""},{"id":337181089,"identity":"618dbd3f-ccf2-4527-80dd-ea256fa4be9f","order_by":6,"name":"Hongyu Chen","email":"","orcid":"","institution":"School of Chemistry and Pharmaceutical Engineering, Shandong First Medical University \u0026 Shandong Academy of Medical Sciences, Taian 271000, Shandong, China","correspondingAuthor":false,"prefix":"","firstName":"Hongyu","middleName":"","lastName":"Chen","suffix":""}],"badges":[],"createdAt":"2024-07-23 05:45:36","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4785879/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4785879/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s11468-024-02510-2","type":"published","date":"2024-08-27T15:57:35+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":62864783,"identity":"814cf072-e97f-40fe-820c-cd5e1ed0bb24","added_by":"auto","created_at":"2024-08-20 11:18:44","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":3772074,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e UV-Vis absorption spectra of citrate-stabilized AuNPs colloid in the absence and presence of creatinine (10 mM). The inset shows the corresponding photographic images of AuNPs colloid. \u003cstrong\u003eb\u003c/strong\u003e Hydrodynamic particle size distribution of AuNPs in the absence and presence creatinine (10 mM, pH 7.0). SEM images of citrate-stabilized AuNPs in the absence \u003cstrong\u003ec\u003c/strong\u003e and presence \u003cstrong\u003ed\u003c/strong\u003ecreatinine (10 mM, pH 7.0)\u003c/p\u003e","description":"","filename":"Figure1.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/298aa55894c0805c68f0d54b.png"},{"id":62864785,"identity":"d40ce262-1118-4fdc-b33d-96e9651085be","added_by":"auto","created_at":"2024-08-20 11:18:44","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":6381286,"visible":true,"origin":"","legend":"\u003cp\u003eSchematic of the mechanism of creatinine detection based on citrate-stabilized AuNPs\u003c/p\u003e","description":"","filename":"Figure2.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/76b1e98b02fba2a18a237242.png"},{"id":62864789,"identity":"101d5c75-a8f6-462b-85e5-bb524887e3b2","added_by":"auto","created_at":"2024-08-20 11:18:44","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":5373482,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e UV-Vis spectra of the AuNPs colloid in different pH medium (4.0, 5.0, 6.0, 7.0, 8.0, 9.0 and 10.0). \u003cstrong\u003eb\u003c/strong\u003e The corresponding effect of the pH on the absorbance of AuNPs. \u003cstrong\u003ec\u003c/strong\u003eThe photographic images of AuNPs colloid formed in different pH medium\u003c/p\u003e","description":"","filename":"Figure3.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/af3454a9186d8007673a657c.png"},{"id":62864784,"identity":"1caa1afa-e48a-4335-bbae-a46b2acd7796","added_by":"auto","created_at":"2024-08-20 11:18:44","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":1543764,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e Absorbance of AuNPs with different concentrations before and after addition of creatinine. \u003cstrong\u003eb\u003c/strong\u003e Effect of AuNPs concentrations on the relative absorbance (ΔA=A\u003csub\u003e0\u003c/sub\u003e-A, where A\u003csub\u003e0\u003c/sub\u003e and A are the absorbance at 521 nm in the absence and presence of creatinine, respectively)\u003c/p\u003e","description":"","filename":"Figure4.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/59cb282b1d68b8e9c50d5031.png"},{"id":62864782,"identity":"161c5a78-a344-4b28-8037-367d32aea19f","added_by":"auto","created_at":"2024-08-20 11:18:44","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":7049425,"visible":true,"origin":"","legend":"\u003cp\u003eTime-dependent UV-Vis spectra of AuNPs in the presence of \u003cstrong\u003ea\u003c/strong\u003e 0.5 mM, \u003cstrong\u003eb\u003c/strong\u003e 10 mM and \u003cstrong\u003ec\u003c/strong\u003e 20 mM creatinine. \u003cstrong\u003ed\u003c/strong\u003e The ΔA (ΔA=A\u003csub\u003e0\u003c/sub\u003e-A, where A\u003csub\u003e0\u003c/sub\u003e and A are the absorbance at 521 nm in the absence and presence of creatinine, respectively) of AuNPs in the presence of 0.5 mM, 10 mM and 20 mM creatinine as a function of reaction time\u003c/p\u003e","description":"","filename":"Figure5.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/0527a6ff2b7255555e50ddb5.png"},{"id":62865201,"identity":"128ed509-a01d-4049-b1bd-dd4b95df2f1a","added_by":"auto","created_at":"2024-08-20 11:26:44","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":5267989,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003ea\u003c/strong\u003e The photographs of the AuNPs solution with different concentrations of creatinine. \u003cstrong\u003eb\u003c/strong\u003e The UV-Vis absorption spectra of AuNPs with different concentrations of creatinine and C the corresponding standard dose-response curve of creatinine\u003c/p\u003e","description":"","filename":"Figure6.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/b036a13bf3a3fa75a442e509.png"},{"id":62864788,"identity":"38b09f70-06b1-43d2-8096-df46c17cd2b4","added_by":"auto","created_at":"2024-08-20 11:18:44","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":1152865,"visible":true,"origin":"","legend":"\u003cp\u003eThe relative absorbance (ΔA) of the AuNPs in the presence of creatinine or other interferences (the concentration of creatinine was 1 mM and the other interferences were 10 mM)\u003c/p\u003e","description":"","filename":"Figure7.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/e62a36e61cacdf203043f52b.png"},{"id":62864787,"identity":"65808d12-0d0c-47e8-9714-0d6ff730e103","added_by":"auto","created_at":"2024-08-20 11:18:44","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":1856615,"visible":true,"origin":"","legend":"\u003cp\u003eThe relative absorbance (ΔA) of AuNPs with different concentrations of creatinine in buffer solution and diluted artificial urine samples\u003c/p\u003e","description":"","filename":"Figure8.png","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/9cee7340fd5da2e3f539b71a.png"},{"id":63820994,"identity":"1d9c9e8e-5628-4602-9e25-3f6373f8d77a","added_by":"auto","created_at":"2024-09-02 16:10:55","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":63599945,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4785879/v1/4ff208c6-aa28-4b77-b1e0-749c55a925c5.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"A Simple and Efficient Colorimetric Detection of Creatinine Based on Citrate-Stabilized Gold Nanoparticles","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eKidney disease has been one of the leading causes of death worldwide [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e], but less than 10% of patients with early renal dysfunction are diagnosed and treated in a timely manner [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Therefore, it is important to timely identify and diagnose kidney diseases, especially in resource-limited settings. Creatinine is a nitrogenous breakdown product produced during the metabolism of creatine and creatine phosphate in the muscle tissue [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e], and the normal kidney function filters and excretes creatinine with the urine [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. The normal urinary creatinine levels are in the range of 0.3 to 3.1 g/L (2.4 to 27.0 mM) [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e], but vary depending on age and gender [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Changes in kidney function and creatinine metabolism can significantly affect urine-creatinine levels. Creatinine levels that deviate from the normal range can indicate various kidney diseases such as chronic kidney diseases (CKDs) and acute kidney injury (AKI) [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Thus, creatinine levels are considered to be one of the most effective indicators for evaluating renal function and preventing kidney diseases.\u003c/p\u003e \u003cp\u003eTo date, various methods are available for the determination of creatinine levels, including high performance liquid chromatography [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e], nuclear magnetic resonance [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e], liquid chromatography-isotope dilution mass spectrometry [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e], etc. Although these methods can offer sensitive and accurate measurement, most of them are time-consuming and expensive as well as require sophisticated instruments and/or trained staff [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Therefore, it is essential to develop a simple and cost-effective method for the detection of creatinine. Generally, colorimetric assays have been studied as an ideal candidate for detection of creatinine, such as Jaffe and enzymatic methods [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. The Jaffe method is based on the reaction of creatinine with alkaline picrate to produce an orange-red complex that can be quantitatively monitored by spectrophotometry [\u003cspan additionalcitationids=\"CR15\" citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. The enzymatic method depends on the hydrolysis of creatinine by creatinase [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. The Jaffe method is more susceptible interferences from some organic compounds (such as ascorbic acid, glucose, bilirubin and certain proteins), which can lead to false positive results [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. Whereas, the enzymatic method exhibits better anti-interference ability but is more expensive than Jaffe method, and sources of key enzyme is difficult to obtain [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eTo overcome the limitations of traditional methods, different types of sensors have been developed, such as surface-enhanced Raman spectroscopy [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e], nanotechnology-based sensors [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e], electrochemical biosensors [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e], etc. Among them, noble metal (Au, Ag and Cu) nanomaterial-based colorimetric sensing displays high sensitivity due to its unique localized surface plasmon resonance (LSPR) effect [\u003cspan additionalcitationids=\"CR24\" citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. Typically, the LSPR effect of the metallic nanoparticles is dependent on the size, shape, and distance between nanoparticles, particularly the gold nanoparticles (AuNPs) with high extinction coefficients (10\u003csup\u003e8\u003c/sup\u003e ~ 10\u003csup\u003e10\u003c/sup\u003e M\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e \u0026middot; cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. Specifically, according to the Mie theory, when AuNPs are in the strong coupling plasmonic regime (low interparticle distance), a red-shift and an amplification of the LSPR as the interparticle distance decreases are observed [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. That is, when the analyte induces the aggregation of AuNPs, the LSPR peak and colour of AuNPs colloid can undergo remarkable changes, thereby achieving colorimetric detection of the analyte. For example, Yi He et al. demonstrated a colorimetric method through studying the relationship between the red-shift values of AuNPs LSPR peak before and after introducing the creatinine [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. Sittiwong et al. further refined the pretreatment of creatinine in urine samples by extracting creatinine from urine samples using sulfonic acid-functionalized silica gel, and then determining creatinine based on the changes in LSPR signals before and after creatinine-induced aggregation of AuNPs [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]. Joseph et al. developed a smartphone-based sensor kit for monitoring creatinine through polyol functionalized AuNPs [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. The currently reported plasmonic-based colorimetric sensing assays provide an effective research tool for accurate and sensitive monitoring creatinine levels. However, the difficulty in accurately controlling the extent of creatinine-induced aggregation of AuNPs, and the complex procedures for functionalizing AuNPs and extracting creatinine limit the utility of the method developed assays in point-of-care conditions. Therefore, there is a pressing need for a simple and practical method to quantify creatinine in biological fluids.\u003c/p\u003e \u003cp\u003eIn this work, we developed a simple method for creatinine detection by studying the relative absorbance (ΔA) of citrate-stabilized AuNPs in absence and presence of creatinine. The optimized experimental parameters such as the pH values, the AuNPs concentrations and the reaction time were investigated. Under the optimized conditions, the AuNPs-based colorimetric determination of creatinine showed a better specificity in the concentration range from 0.5 to 20 mM with the limit of detection of 0.16 mM. Furthermore, the proposed protocol has been successfully used to detect the creatinine level in urine. It demonstrates that the colorimetric detection based on citrate-stabilized AuNPs provides a facile route for the high sensitivity and specificity of creatinine in clinical diagnosis.\u003c/p\u003e"},{"header":"2. Experimental Section","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Chemicals\u003c/h2\u003e \u003cp\u003eHydrogen tetrachloroaurate (III) trihydrate (HAuCl\u003csub\u003e4\u003c/sub\u003e\u0026middot;3H\u003csub\u003e2\u003c/sub\u003eO) and artificial urine solutions were purchased from Sinopharm Chemical Reagent Co., Ltd. Trisodium citrate (Na\u003csub\u003e3\u003c/sub\u003eC\u003csub\u003e6\u003c/sub\u003eH\u003csub\u003e5\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u0026middot;2H\u003csub\u003e2\u003c/sub\u003eO), hydrochloric acid (HCl), nitric acid (HNO\u003csub\u003e3\u003c/sub\u003e), sodium hydroxide (NaOH), boric acid (H\u003csub\u003e3\u003c/sub\u003eBO\u003csub\u003e3\u003c/sub\u003e), acetic acid (CH\u003csub\u003e3\u003c/sub\u003eCOOH), phosphoric acid (H\u003csub\u003e3\u003c/sub\u003ePO\u003csub\u003e4\u003c/sub\u003e), creatinine (C\u003csub\u003e4\u003c/sub\u003eH\u003csub\u003e7\u003c/sub\u003eN\u003csub\u003e3\u003c/sub\u003eO), urea (CH\u003csub\u003e4\u003c/sub\u003eN\u003csub\u003e2\u003c/sub\u003eO) and uric acid (N\u003csub\u003e5\u003c/sub\u003eH\u003csub\u003e4\u003c/sub\u003eN\u003csub\u003e4\u003c/sub\u003eO\u003csub\u003e3\u003c/sub\u003e) were obtained from Aladdin (Shanghai, China). Britton-Robinson (B-R) buffer solutions were prepared by H\u003csub\u003e3\u003c/sub\u003eBO\u003csub\u003e3\u003c/sub\u003e, CH\u003csub\u003e3\u003c/sub\u003eCOOH and H\u003csub\u003e3\u003c/sub\u003ePO\u003csub\u003e4\u003c/sub\u003e solutions all at a concentration of 4 mM, and NaOH solution (0.2 mol/L) was used to adjust the different pH values. Different concentrations of creatinine solution were prepared in the B-R buffer solution (44 mM, pH 7.0). All reagents were analytical grade and used as received, and Millipore ultrapure double deionized water (18 MΩ) was used for all solution preparations. All glassware was cleaned by configured aqua regia (3:1; HCl/HNO\u003csub\u003e3\u003c/sub\u003e) for 15 min, then rinsed thoroughly with distilled water several times and dried in a hot air oven at 85˚C.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Instruments\u003c/h2\u003e \u003cp\u003eUltraviolet-Visible (UV-Vis) absorption spectra were monitored with a spectrophotometer (Shimadzu UV-2450, Japan). Mean hydrodynamic diameter of AuNPs were determined by dynamic light scattering (DLS) with Malvern laser particle size analyser (C-W10X10XB22, China). The morphology of AuNPs was characterized by using a high-resolution field emission scanning electron microscope (SEM, JEOL JSM-7900F, Japan).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 Preparation of the Citrate-Stabilized AuNPs\u003c/h2\u003e \u003cp\u003eThe synthesis of AuNPs colloid was according to the classical sodium citrate reduction method [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. Briefly, 4 mL of sodium citrate (1%) was quickly added to 96 mL HAuCl\u003csub\u003e4\u003c/sub\u003e (0.007%) boiling solution under vigorous stirring until the colour of mixture solution change to red. After that, the reaction mixture was stirred and refluxed for an additional 15 min, and then cooled to room temperature. The AuNPs colloid was successfully prepared and stored in a clean tube at 4\u0026deg;C for further use. According to the Beer-Lambert law, the concentration of AuNPs colloid was estimated to be 1.97 nM using an extinction coefficient of 2.7\u0026times;10\u003csup\u003e8\u003c/sup\u003e M\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e \u0026middot; cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e at 521 nm [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Detection of Creatinine\u003c/h2\u003e \u003cp\u003eFirstly, 2.5 mL AuNPs colloid dispersed in 2.5 mL B-R buffer solution was respectively added to each of the six test tubes and incubated for 15 min at room temperature. Secondly, 60 \u0026micro;L of the newly prepared creatinine solution with different concentrations (0.5 mM, 1 mM, 5 mM, 10 mM, 15 mM, and 20 mM) were respectively added to the above AuNPs solutions. After incubating at room temperature for 233 min, the mixture solution was subjected to measure UV-Vis absorption spectrum, and the visual observation photographs were taken at the same time.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.5 Creatinine Detection in Urine Sample\u003c/h2\u003e \u003cp\u003eTo prevent the influence of proteins that may be present in the urine, the artificial urine samples were filtered through a 0.22 \u0026micro;m membrane and then diluted 100 times with B-R buffer solution for immediate testing. Subsequently, 60 \u0026micro;L of creatinine solution (0.5 mM, 10 mM, 20 mM) prepared with the diluted artificial urine was added to the AuNPs solution, respectively. Then, the detection of creatinine was performed as described above.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results and Discussion","content":"\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e3.1 Working Principle of Creatinine Detection\u003c/h2\u003e \u003cp\u003eFigure\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea shows the UV-Vis absorption spectra of citrate-stabilized AuNPs colloid in the wavelength range of 400\u0026ndash;800 nm before and after addition of creatinine at room temperature. The AuNPs colloid prepared by the citrate reduction method displays characteristic LSPR band located at 521 nm. After the addition of creatinine, the LSPR peak of AuNPs at 521 nm was weakened and slightly red-shifted to 525 nm. And it was noted that a new absorption band emerged at 663 nm due to the aggregation of AuNPs induced by linkage with creatinine, and meanwhile a red-to-blue colour changes could be observed (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea, inset). It is demonstrated that the introduction of creatinine can induce the aggregation of AuNPs, thereby triggering the colour change of AuNPs colloid.\u003c/p\u003e \u003cp\u003eMoreover, to further confirm that the introduction of creatinine could cause the AuNPs aggregation, the particle size distribution and morphology of AuNPs before and after introduction of creatinine were performed by using DLS and SEM (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb-\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed), respectively. The hydrodynamic diameter distribution of the citrate-stabilized AuNPs in the absence and presence of creatinine were illustrated in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb. The AuNPs without creatinine show the mono-dispersity with an average hydrodynamic diameter of 29 nm. Upon the addition of creatinine, the aggregated AuNPs exhibit two major peaks at 37 nm and 251 nm. The former peak corresponds to the average size of citrate-stabilized AuNPs with a slight increase in size, which is consistent with the slightly red-shifted UV-Vis spectrum. The latter peak was attributed to the aggregated AuNPs, whose particle size increase significantly. In addition, the SEM im-ages of AuNPs before and after incubation with creatinine are shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed. It can be seen that the citrate-stabilized AuNPs were uniformly distributed with an average particle size of 23\u0026thinsp;\u0026plusmn;\u0026thinsp;2 nm by using Nano Measurer, according to the SEM images (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec). After incubation with creatinine, significant aggregation of AuNPs is observed in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed. These results suggested that creatinine can induce the aggregation of citrate-stabilized AuNPs.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTo explore the origins of the AuNPs aggregation induce by creatinine, the surface charge and structural properties of AuNPs before and after introduction of creatinine was analysed. The principle of creatinine detection using citrate-stabilized AuNPs is illustrated in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e. Before introducing of creatinine, the structure of citrate-stabilized AuNP consists a central carboxylate group interacting with AuNP surface and the remaining two terminal carboxylate group (see Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea) [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e]. Under neutral conditions (B-R buffer solution, pH 7.0), the electrostatic repulsion between the negatively charged carboxylate group of citrate anions adsorbed on AuNPs maintains the homogeneously dispersion of the AuNPs. It has been reported that the creatinine has two tautomeric (amino tautomer and imino tautomer) and can be interconverted between two tautomeric under neutral condition (see Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb) [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. After addition of creatinine, the citrate adsorbed on the AuNPs can be replaced by the creatinine ligands, due to the coordinated interaction between the nitrogen containing functional group (creatinine) and AuNPs is much stronger than that of citrate ions [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e, \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. After ligand-exchanged, creatinine and its two tautomeric are neutral charged at pH 7.0, which greatly reduces the number of surface negative charge of AuNPs, resulting in the aggregation of AuNPs (see Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea and \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec). Besides, creatinine and its two neutral tautomers can disrupt the electrostatic balance of citrate-stabilized AuNPs by forming hydrogen bonds with the exposed carboxylate groups of the citrate molecule, thereby causing the aggregation of AuNPs (see Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec) [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Thus, creatinine could induce the aggregation of AuNPs and accompanying a colour change through the synergistic effect of ligand-exchanged and hydrogen bonds with the citrate ions adsorbed on the AuNPs, thereby achieving colorimetric detection.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003e3.2 Optimization of Detection Conditions\u003c/h2\u003e \u003cdiv id=\"Sec11\" class=\"Section3\"\u003e \u003ch2\u003e3.2.1 Effect of pH\u003c/h2\u003e \u003cp\u003eThe pH value of AuNPs colloid is a crucial factor affecting the sensitivity and accuracy of creatinine detection. Here, the media samples were prepared using B-R buffer solutions with pH values of 4.0 to 10.0, and AuNPs colloid dispersed in 5 mL B-R buffer solution at different pH values. To evaluate the influence of pH on the stability of AuNPs colloid, we studied the effect of pH value of AuNPs over the absorbance (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea). With the increasing of pH value from 4.0 to 7.0, the absorbance at 521 nm progressively increases (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb), accompanied by a change in colour from purple to red (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec). This is because at pH\u0026thinsp;\u0026lt;\u0026thinsp;7, citrate is fully protonated and the number of surface negative charge is greatly reduced, resulting in the aggregation of AuNPs [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. While, with the increasing of pH value from 7.0 to 10.0, the absorbance at 521 nm progressively decreases (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb), accompanied by a change in colour from red to purple (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec). It can be interpreted that the two free terminal carboxylic acid groups of citrates adsorbed on the surface of AuNPs can be induced to bind to the surface of AuNPs through an increase of pH [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e], leading to a decrease in surface negative charge and thus causing the aggregation of AuNPs. At neutral pH, the citrate is deprotonated, the negative surface charge result in electrostatic repulsion between AuNPs and maintained the red wine colour indicating no aggregation took place. It was noticed that a new absorption band in the longwave direction appeared at higher or lower pH due to the aggregation of AuNPs, accompanied by a visible colour change from red to purple. Therefore, pH value has a significant impact on the stability of AuNPs, especially at higher or lower pH value. We choose pH 7.0 for the following experiments.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section3\"\u003e \u003ch2\u003e3.2.2 Effect of AuNPs concentration\u003c/h2\u003e \u003cp\u003eThe AuNPs concentrations are another significant factor affecting the limit of detection and sensitivity of creatinine. Here, different concentrations of AuNPs (0.59 to 1.37 nM) were prepared using B-R buffer solution (pH 7.0) at room temperature. The absorbance of various concentrations of AuNPs before and after the addition of creatinine was determined and shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea. As anticipated, with the increasing of AuNPs concentrations from 0.59 to 1.37 nM, the absorbance at 521 nm in the absence and presence of creatinine gradually increases (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea). While, with the increasing of AuNPs concentrations from 0.59 to 0.98 nM, the ΔA at peak at 521 nm progressively increases and then decreases from AuNPs concentrations of 0.98 to 1.37 nM (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb). It can be interpreted that higher AuNPs concentrations require more creatinine to compete with citrate ions for binding sites, and a small amount of creatinine cannot effectively induce the aggregation of higher concentrations of AuNPs, resulting in an insignificant change in ΔA at 521 nm after the addition of creatinine [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e]. Therefore, AuNPs concentration of 0.98 nM was chosen for the following experiments.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section3\"\u003e \u003ch2\u003e3.2.3 Effect of reaction time\u003c/h2\u003e \u003cp\u003eFigure\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea displays the UV-Vis spectra of AuNPs in the presence of 0.5 mM creatinine at different incubation time. With the increasing of incubation time, the absorption band at 521 nm decreased, and remained unchanged until about 103 min, but meanwhile, the new peak at 650 nm red-shifted and increased. To determine the effect of creatinine concentration on reaction time, we also studied the UV-Vis absorption spectra of AuNPs in the presence of 10 mM and 20 mM creatinine with different reaction times (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eb and \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ec). Similarly, upon reaction time, the absorption band at 521 nm decreased, and remain unchanged at about 125 min and 135 min, respectively. And meanwhile the absorption band at 650 nm red-shifted and increased. Figure\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ed shows the relative absorbance as a function of the incubation time in the presence of different concentrations of creatinine. The time constant τ for AuNPs reaction with creatinine of different concentration was further derived by using the exponential decay formula \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:y={y}_{0}+A\\text{*}{e}^{-x/t}\\)\u003c/span\u003e\u003c/span\u003e. From our study, it can be found that τ is 14\u0026thinsp;\u0026plusmn;\u0026thinsp;1 min, 24\u0026thinsp;\u0026plusmn;\u0026thinsp;4 min and 24\u0026thinsp;\u0026plusmn;\u0026thinsp;2 min for the concentration of 0.5 mM, 10 mM and 20 mM creatinine, respectively. The reaction time when the reaction reached equilibrium were calculated from the first order reaction kinetic equation to be 133\u0026thinsp;\u0026plusmn;\u0026thinsp;13 min, 233\u0026thinsp;\u0026plusmn;\u0026thinsp;13 min and 226\u0026thinsp;\u0026plusmn;\u0026thinsp;40 min, separately. As consequence, the optimal incubation time was selected to be 233 min.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e3.3 Detection of Creatinine\u003c/h2\u003e \u003cp\u003eUnder the optimized conditions, the colorimetric determination of creatinine was performed. As the creatinine concentration increased, the colour of AuNPs solution gradually changed from red to purple and then to blue (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ea), which could be visible to naked eyes above 0.5 mM. Thus, it can be visually detected by the naked eye when the creatinine concentration is higher than 0.5 mM. Furthermore, the colour change of AuNPs in presence of creatinine was quantitatively determined by using UV-Vis spectrophotometry. Figure\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eb shows the UV-Vis spectra of AuNPs with different concentrations of creatinine. As the creatinine concentration increased, the absorption bands at 521 nm progressively decreased and red-shifted to 525 nm and 529 nm, while the absorption bands of 600 to 700 nm emerged and gradually increased. The differences in absorbance at 521 nm of AuNPs with and without creatinine was used to compare the aggregation of AuNPs caused by different concentrations of creatinine. Figure\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ec shows the relationship between ΔA and creatinine concentration. The corresponding standard dose-response curve was a linear relationship described by the fitting curve \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\text{y}=0.00746\\text{x}+0.0519\\)\u003c/span\u003e\u003c/span\u003e (see Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003ec), and the limit of detection (LOD) of creatinine was determined to be 0.16 mM (S/N\u0026thinsp;=\u0026thinsp;3), which was calculated via the linear regression method (LOD\u0026thinsp;=\u0026thinsp;3σ/slope) [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e, \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. The reproducibility of this method was further evaluated by a series of six parallel experiments of three different concentrations of creatinine (0.5 mM, 10 mM, 20 mM), corresponding to a relative standard deviation (RSD) of 3.4%, 1.8% and 4.8%, respectively. Therefore, our colorimetric methods based on citrate-stabilized AuNPs has high sensitivity and better reproducibility to reliably detect the trace creatinine level.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTable\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e presents the summary of certain current methods for creatinine determination and their detection capabilities. It can be found that the LOD of our method is lower than that of the commonly used Jaffe-based method in the clinic, indicating that the established method can meet the requirements for clinical detection. Compared with some of other AuNPs-based creatinine assays, our method has a wider linear range or higher linear correlation, and allows for creatinine detection without requiring functionalized modification of AuNPs or pretreatment of creatinine. Thus, our suggested AuNPs-based colorimetric detection of creatinine strategy has a LOD equivalent to or lower than that of common methods, and our strategy is remarkable for its simplicity and effectivity.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComparison of the present method with some previously reported methods for urinary creatinine detection\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMethods\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eLimit of detection\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eAnalytical range (mM)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eR\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eReference\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSmartphone-based method\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.084 \u0026micro;M\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.001\u0026ndash;0.05\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.975\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e[\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eElectrochemical method\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.04 mM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u0026ndash;1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.98\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e[\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e]\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAuNPs aggregation-based colorimetric method\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.08 mM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.1\u0026ndash;20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.99\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e[\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eJaffe-based method\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.72 mM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0\u0026ndash;6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.991\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e[\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e]\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePolyol functionalized AuNPs methods\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.7 nM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0-0.002\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.994\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e[\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAmperometric method\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.21 mM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.5-2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e-\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e[\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e]\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSolid phase extraction method\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.12 mM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.13\u0026ndash;0.35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.998\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e[\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMonodisperse AuNPs-based method\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.16 mM\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0.5\u0026ndash;20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.999\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eThis work\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003e3.4 Assay Specificity\u003c/h2\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe presence of inorganic ions and other organic compounds in the urine sample may lead to a false-positive of creatinine estimation. In order to verify the specific recognition of creatinine in urine samples by the proposed sensor, some potential interfering substances, including glucose (Glu), glycine (Gly), ascorbic acid (AA), urea, uric acid (UA), sodium chloride (NaCl), ammonium chloride (NH\u003csub\u003e4\u003c/sub\u003eCl), potassium phosphate (K\u003csub\u003e3\u003c/sub\u003ePO\u003csub\u003e4\u003c/sub\u003e), sodium sulfate (Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e) and magnesium chloride (MgCl\u003csub\u003e2\u003c/sub\u003e) were detected under the same optimum conditions. Figure\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e shows the ΔA for interfering substances (10 mM) and creatinine (1 mM). Obviously, the ΔA in presence of the above interfering substances is much smaller than that of creatinine, indicating that these substances did not interfere with the recognition and detection of creatinine. The observed high selectivity in this case was attributed to the stronger affinity between the three nitrogen atoms of creatinine and AuNPs within the concentration range of this method under the specified experimental conditions. Moreover, the pKa of uric acid and glycine were 5.5 and 2.3, respectively, which were lower than the pH of medium in this assay (7.0). Therefore, uric acid and glycine were negatively charged. When they were adsorbed onto the surface of AuNPs through nitrogen atoms, they did not disrupt the electrostatic equilibrium of AuNPs particles. Consequently, the proposed sensor can be used to specifically detect creatinine.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e3.5 Detection of Creatinine in artificial urine Samples\u003c/h2\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTo evaluate the practical application, different amounts of creatinine were added into the diluted artificial urine samples (1:100) and then detected by the proposed sensor. As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e, it can be found that the ΔA of AuNPs is increased with increasing the concentrations of creatinine, which is similar to the situation in buffer solution. It is indicated that our proposed method can successfully detect the creatinine level in urine samples. Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e specifically presents the comparison of the detection results of creatinine in B-R buffer solution and diluted artificial urine. As can be seen in Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, the amount of creatinine added and the assay values were comparable in both buffer solution and diluted urine, respectively, indicating the high accuracy of our method. Compared with detection results of creatinine in buffer solution, we performed the same six sets of parallel measurements of 0.5 mM, 10 mM and 20 mM creatinine in diluted urine, and the corresponding relative standard deviations were 8.4%, 1.6% and 4.3%, respectively. It demonstrated that our method has good reproducibility even in diluted urine. Moreover, the spiked recoveries of 0.5 mM, 10 mM and 20 mM creatinine in diluted urine were calculated as 110%, 103.4% and 105.3%, respectively, indicating the high reliability of our method. Therefore, our method is suitable for the detection of creatinine in artificial urine.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComparison of the sensing results of creatinine in B-R solution and in diluted artificial urine\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSample\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eAdded (mM)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eDetected (mM)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRecovery (%)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eRSD (n\u0026thinsp;=\u0026thinsp;6, %)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eCreatinine in buffer\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.48\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e96\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e3.4\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e9.73\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e97.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e19.58\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e97.9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e4.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003eCreatinine in urine\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.55\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e110\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e8.4\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e10.34\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e103.4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e1.6\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e20\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e21.06\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e105.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e4.3\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"4. Conclusions","content":"\u003cp\u003eIn summary, we proposed a strategy by studying the relative absorbance of citrate-stabilized AuNPs in absence and presence of creatinine to quantitively detect urine creatinine level. The experimental conditions that may affect the sensitivity and accuracy of creatinine detection, including the pH, the concentration of AuNPs and reaction time, were optimized. Subsequently, under optimal conditions, the relative absorbance of citrate-stabilized AuNPs with creatinine concentration range of 0.5 to 20 mM were obtained, and the LOD was calculated to be 0.16 mM. Moreover, the control experiments for detection of inorganic ions and other organic compounds in the urine sample exhibited that the proposed methods had superior selectivity for the detection of creatinine. Furthermore, the method was successfully applied to the detection of creatinine level in spiked artificial urine. The proposed method based on the relative absorbance of citrate-stabilized AuNPs has a great potential for detecting creatinine in resource-limited point-of-care settings sites.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eCompeting Interests\u003c/h2\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003ch2\u003eFunding\u003c/h2\u003e\n\u003cp\u003eThis work is supported by the National Science Foundation of China (No. 52305317), Natural Science Foundation of Shandong Province (Nos. ZR2022QB040, ZR2022QH006 and ZR20230B113).\u003c/p\u003e\n\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\n\u003cp\u003e\u0026ldquo;H.C. and L.Z. conceived the idea of the work. X.L., T.S., X.B. and Z.G. performed design, material preparation, data collection, and analysis. X.L. wrote the original manuscript text. R.J., H.C. and L.Z. edited the manuscript and reviewed the numerical analysis. All authors read and approved the final manuscript.\u0026rdquo;\u003c/p\u003e\n\u003ch2\u003eData availability\u003c/h2\u003e\n\u003cp\u003eThe data sets used and/or analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eDong Y, Liu Y, Lv J, Yang L, Cui Y et al (2023) Advancements in Amperometric biosensing instruments for creatinine detection: A critical review. IEEE Trans Instrum Meas 72: 1-15. https://doi.org/10.1109/TIM.2023.3279459\u003c/li\u003e\n\u003cli\u003eEne-Iordache B, Perico N, Bikbov B (2016) Chronic kidney disease and cardiovascular risk in six regions of the world (ISN-KDDC): a cross-sectional study. 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Electroanalysis 24: 2325-2331. https://doi.org/10.1002/elan.201200497\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"plasmonics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plas","sideBox":"Learn more about [Plasmonics](https://www.springer.com/journal/11468)","snPcode":"11468","submissionUrl":"https://submission.nature.com/new-submission/11468/3","title":"Plasmonics","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"citrate-stabilized gold nanoparticles (AuNPs), creatinine, colorimetric method, relative absorbance (ΔA)","lastPublishedDoi":"10.21203/rs.3.rs-4785879/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4785879/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eCreatinine level is a crucial indicator in the clinical assessment and diagnosis of renal diseases, achieving simple and accurate detection of urinary creatinine levels in resource-limited point-of-care settings is of great significant in the timely prevention and diagnosis of kidney diseases. As a popular zero-dimensional material, gold nanoparticles (AuNPs) exhibit intriguing optical properties and thus have become a promising material for many sensing detection applications. Here, we proposed a simple, efficient and sensitive quantitative detection of creatinine by studying the relative absorbance (ΔA) of AuNPs in absence and presence of creatinine. The method relies on the aggregation of AuNPs via ligand-exchanged of citrate ions and creatinine on the surface of AuNPs to achieve colorimetric detection. With this assay, the limit of detection for creatinine was as low as 0.16 mM, and the dynamic detection range was 0.5 to 20 mM under optimized conditions. In our experiments, the specificity of proposed method was investigated and successfully applied to detect creatinine in urine sample. It reveals that the proposed colorimetric protocol has demonstrated a high sensitivity and selectivity for creatinine, and has a potential practicability in clinical diagnostics.\u003c/p\u003e","manuscriptTitle":"A Simple and Efficient Colorimetric Detection of Creatinine Based on Citrate-Stabilized Gold Nanoparticles","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-08-20 11:18:39","doi":"10.21203/rs.3.rs-4785879/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-08-07T11:40:04+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-07-30T06:58:29+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"114185371837092271131687200331439225389","date":"2024-07-26T08:22:28+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-07-24T10:33:38+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-07-24T07:56:51+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-07-24T07:55:03+00:00","index":"","fulltext":""},{"type":"submitted","content":"Plasmonics","date":"2024-07-23T05:44:12+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"plasmonics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"plas","sideBox":"Learn more about [Plasmonics](https://www.springer.com/journal/11468)","snPcode":"11468","submissionUrl":"https://submission.nature.com/new-submission/11468/3","title":"Plasmonics","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"fec3183e-3778-4a27-99a5-36e90496f64a","owner":[],"postedDate":"August 20th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-09-02T16:01:52+00:00","versionOfRecord":{"articleIdentity":"rs-4785879","link":"https://doi.org/10.1007/s11468-024-02510-2","journal":{"identity":"plasmonics","isVorOnly":false,"title":"Plasmonics"},"publishedOn":"2024-08-27 15:57:35","publishedOnDateReadable":"August 27th, 2024"},"versionCreatedAt":"2024-08-20 11:18:39","video":"","vorDoi":"10.1007/s11468-024-02510-2","vorDoiUrl":"https://doi.org/10.1007/s11468-024-02510-2","workflowStages":[]},"version":"v1","identity":"rs-4785879","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4785879","identity":"rs-4785879","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}