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2020 Volume 48 Issue 2
2020, 48(2): 153-163
doi: 10.19756/j.issn.0253-3820.191399
Abstract:
G-quadruplex is a special secondary structure of nucleic acid. G-quadruplex can bind with hemin to form DNAzyme with peroxidase activity, and also it can enhance the fluorescence intensity of some dyes. As one of the functional nucleic acids, G-quadruplex is widely used in various biochemical analyses due to its high stability, specificity and flexibility. The application of G-quadruplex in biochemical analysis and its application prospects are briefly introduced.
G-quadruplex is a special secondary structure of nucleic acid. G-quadruplex can bind with hemin to form DNAzyme with peroxidase activity, and also it can enhance the fluorescence intensity of some dyes. As one of the functional nucleic acids, G-quadruplex is widely used in various biochemical analyses due to its high stability, specificity and flexibility. The application of G-quadruplex in biochemical analysis and its application prospects are briefly introduced.
2020, 48(2): 164-173
doi: 10.19756/j.issn.0253-3820.191562
Abstract:
Polydiacetylenes (PDAs) have received increasing attention as smart materials owing to their unique optical and electronic properties. PDAs are the most commonly synthesized as self-assembled bilayer vesicles via 1,4 addition polymerization. Vesicles are often perceived as closely mimicking the cell membrane, thus functioning as convenient biomimetic platforms. PDAs undergo colorimetric and fluorescent transition in response to external stimuli. Therefore, PDA vesicles can be used for medical detection, such as bacteria, viruses, proteins, and enzymes, etc. In this review, the preparation of PDA vesicle sensing systems and the mechanism of solid phase polymerization are summarized. The relative applications of PDA vesicle sensing systems in medical detection are introduced. Besides, the shortcomings and issues to be solved, and future directions are also discussed and prospected.
Polydiacetylenes (PDAs) have received increasing attention as smart materials owing to their unique optical and electronic properties. PDAs are the most commonly synthesized as self-assembled bilayer vesicles via 1,4 addition polymerization. Vesicles are often perceived as closely mimicking the cell membrane, thus functioning as convenient biomimetic platforms. PDAs undergo colorimetric and fluorescent transition in response to external stimuli. Therefore, PDA vesicles can be used for medical detection, such as bacteria, viruses, proteins, and enzymes, etc. In this review, the preparation of PDA vesicle sensing systems and the mechanism of solid phase polymerization are summarized. The relative applications of PDA vesicle sensing systems in medical detection are introduced. Besides, the shortcomings and issues to be solved, and future directions are also discussed and prospected.
2020, 48(2): 174-179
doi: 10.19756/j.issn.0253-3820.191502
Abstract:
A correlative super-resolution fluorescence and three-dimensional (3D) topography imaging microscopy was built, and the performance of the correlative system was evaluated with F-actin as an example. The experimental results suggested that the correlative instrument had the abilities to provide 3D overall topography and distribution of specific components at nanometer resolution, as well as to correlate the specific components' distribution with the overall topography. Thus, the correlative instrument is a powerful tool to investigate events regarding cytoskeleton and cellular substructure such as the composition and distribution characteristics of membrane protein clusters, and can also be widely applied to other fields such as life science and material science.
A correlative super-resolution fluorescence and three-dimensional (3D) topography imaging microscopy was built, and the performance of the correlative system was evaluated with F-actin as an example. The experimental results suggested that the correlative instrument had the abilities to provide 3D overall topography and distribution of specific components at nanometer resolution, as well as to correlate the specific components' distribution with the overall topography. Thus, the correlative instrument is a powerful tool to investigate events regarding cytoskeleton and cellular substructure such as the composition and distribution characteristics of membrane protein clusters, and can also be widely applied to other fields such as life science and material science.
2020, 48(2): 180-186
doi: 10.19756/j.issn.0253-3820.191580
Abstract:
Tumor microenvironment is a complex system, and it is of great importance in development, invasion and metastasis of tumor. In this work, an in vitro tumor hypoxic microenvironment model was constructed using an oxygen gradient microfluidic chip. The microfluidic chip was composed of two parts:one was a serpentine shape channel to generate oxygen gradient by a chemical reaction, the other was a three-parallel channel for the co-culture of cancer cells (Heap1-6 cells) and hepatic stellate cells (JS-1 cells). The oxygen gradient was made in cancer cells to simulate hypoxic tumor microenvironment, and then a drug-resistance study on paclitaxel and tirapazamine (TPZ) in hepa1-6 cells was performed and the potential reason of the drug resistance was analyzed by molecular mechanism. The result showed that the oxygen gradient was 2.3%-16.7% on chip. The Hepa1-6 cells expressed paclitaxel resistance in hypoxic condition, but cell viability was significant down due to the effect of TPZ. The results of immunofluorescence assay showed hypoxia and co-culture could promote the expression of TIMP-1 and TGF-β, induce the activation of JS-1 cell and then enhance the drug resistance of Hepa1-6 cell to paclitaxel.
Tumor microenvironment is a complex system, and it is of great importance in development, invasion and metastasis of tumor. In this work, an in vitro tumor hypoxic microenvironment model was constructed using an oxygen gradient microfluidic chip. The microfluidic chip was composed of two parts:one was a serpentine shape channel to generate oxygen gradient by a chemical reaction, the other was a three-parallel channel for the co-culture of cancer cells (Heap1-6 cells) and hepatic stellate cells (JS-1 cells). The oxygen gradient was made in cancer cells to simulate hypoxic tumor microenvironment, and then a drug-resistance study on paclitaxel and tirapazamine (TPZ) in hepa1-6 cells was performed and the potential reason of the drug resistance was analyzed by molecular mechanism. The result showed that the oxygen gradient was 2.3%-16.7% on chip. The Hepa1-6 cells expressed paclitaxel resistance in hypoxic condition, but cell viability was significant down due to the effect of TPZ. The results of immunofluorescence assay showed hypoxia and co-culture could promote the expression of TIMP-1 and TGF-β, induce the activation of JS-1 cell and then enhance the drug resistance of Hepa1-6 cell to paclitaxel.
2020, 48(2): 187-196
doi: 10.19756/j.issn.0253-3820.191612
Abstract:
Trastuzumab resistance is one of the principal causes of failure in tumor chemotherapy. Previous studies show that the DNA-binding activity of GATA6, a transcription factor, has a remarkable enhancement in trastuzumab resistant gastric cancer cells (NCI N87/R), while its correlation to resistance remains unclear. In this study, Crispr/Cas9 was employed to establish a GATA6 knock-out cell line (NCI N87R/GATA6). The signaling pathways regulated by GATA6 and related to trastuzumab resistance were investigated based on label-free quantitative proteomics combined with bioinformatics. The extracted proteins were alkylated, and digested using filter aided sample preparation (FASP), and the peptides were separated via high performance liquid chromatography and thereafter quantified by LC-MS/MS. Differentially expressed proteins were screened by fold change and student's t-test between NCI N87/R and NCI N87R/GATA6 cells. WebGestalt website was adopted for Gene ontology analysis, and GeneAnalytics was utilized for pathway enrichment analysis. The results demonstrated that GATA6 knock-out enhanced the antiproliferative effect of trastuzumab on NCI N87/R cells and suppressed their invasion ability. A total of 5792 proteins were quantified by LC-MS/MS, among which 305 proteins were up-regulated in NCI N87R/GATA6 cells while 182 ones down-regulated. Pathway enrichment analysis revealed that mitochondrial transport, apoptosis, DNA damage, glucose metabolism, pyruvate metabolism and TCA cycle and Wnt/β-catenin degradation pathways exhibited significant changes. Western blot manifested that the expression of mitochondrial dyneins OPA1 and DNM1L, apoptosis protein caspase-9, TCA metabolic enzymes SUCLG2 and MDH1, and glycogen metabolic enzyme PYGL changed significantly in NCI N87R/GATA6 cells, manifesting that GATA6 knock-out gave rise to mitochondrial dysfunction and abnormal energy metabolism, and therefore inducing the apoptosis of NCI N87R/GATA6 cells. The result implicated that inhibiting the transcriptional activity of GATA6 could be an effective strategy to reverse trastuzumab resistance in gastric cancer.
Trastuzumab resistance is one of the principal causes of failure in tumor chemotherapy. Previous studies show that the DNA-binding activity of GATA6, a transcription factor, has a remarkable enhancement in trastuzumab resistant gastric cancer cells (NCI N87/R), while its correlation to resistance remains unclear. In this study, Crispr/Cas9 was employed to establish a GATA6 knock-out cell line (NCI N87R/GATA6). The signaling pathways regulated by GATA6 and related to trastuzumab resistance were investigated based on label-free quantitative proteomics combined with bioinformatics. The extracted proteins were alkylated, and digested using filter aided sample preparation (FASP), and the peptides were separated via high performance liquid chromatography and thereafter quantified by LC-MS/MS. Differentially expressed proteins were screened by fold change and student's t-test between NCI N87/R and NCI N87R/GATA6 cells. WebGestalt website was adopted for Gene ontology analysis, and GeneAnalytics was utilized for pathway enrichment analysis. The results demonstrated that GATA6 knock-out enhanced the antiproliferative effect of trastuzumab on NCI N87/R cells and suppressed their invasion ability. A total of 5792 proteins were quantified by LC-MS/MS, among which 305 proteins were up-regulated in NCI N87R/GATA6 cells while 182 ones down-regulated. Pathway enrichment analysis revealed that mitochondrial transport, apoptosis, DNA damage, glucose metabolism, pyruvate metabolism and TCA cycle and Wnt/β-catenin degradation pathways exhibited significant changes. Western blot manifested that the expression of mitochondrial dyneins OPA1 and DNM1L, apoptosis protein caspase-9, TCA metabolic enzymes SUCLG2 and MDH1, and glycogen metabolic enzyme PYGL changed significantly in NCI N87R/GATA6 cells, manifesting that GATA6 knock-out gave rise to mitochondrial dysfunction and abnormal energy metabolism, and therefore inducing the apoptosis of NCI N87R/GATA6 cells. The result implicated that inhibiting the transcriptional activity of GATA6 could be an effective strategy to reverse trastuzumab resistance in gastric cancer.
2020, 48(2): 197-205
doi: 10.19756/j.issn.0253-3820.191615
Abstract:
Bovine serum albumin-mediated copper sulfide nanocomplex (BSA/CuS) was modified on mesoporous silica (MSN) loaded with chemotherapeutic drug doxorubicin (DOX) by amide bond to prepare a new drug carrier, MSN-DOX@BSA/CuS. The disulfide bond of blocking agent BSA would break down after redox reaction with glutathione, resulting in MSN pore exposure and release of DOX. CuS, as a photothermal reagent, could convert light energy into heat for photothermal therapy. UV-Vis-NIR absorption spectra were used to investigate the absorption properties of the carrier. The results showed that the carrier had strong absorption in the range of 800-1100 nm and could realize the photothermal conversion under the near infrared light. In addition, MSN@BSA/CuS was incubated with cancer cells and showed no cytotoxicity. When MSN-DOX@BSA/CuS was added, the relative survival rate of the cells decreased to 49%. After the extra laser, the cell viability was only 18%. The above results showed that the synthesized nano drug carrier could realize the synergistic treatment of photothermal therapy and chemotherapy at the same time, and effectively improved the treatment efficiency of cancer.
Bovine serum albumin-mediated copper sulfide nanocomplex (BSA/CuS) was modified on mesoporous silica (MSN) loaded with chemotherapeutic drug doxorubicin (DOX) by amide bond to prepare a new drug carrier, MSN-DOX@BSA/CuS. The disulfide bond of blocking agent BSA would break down after redox reaction with glutathione, resulting in MSN pore exposure and release of DOX. CuS, as a photothermal reagent, could convert light energy into heat for photothermal therapy. UV-Vis-NIR absorption spectra were used to investigate the absorption properties of the carrier. The results showed that the carrier had strong absorption in the range of 800-1100 nm and could realize the photothermal conversion under the near infrared light. In addition, MSN@BSA/CuS was incubated with cancer cells and showed no cytotoxicity. When MSN-DOX@BSA/CuS was added, the relative survival rate of the cells decreased to 49%. After the extra laser, the cell viability was only 18%. The above results showed that the synthesized nano drug carrier could realize the synergistic treatment of photothermal therapy and chemotherapy at the same time, and effectively improved the treatment efficiency of cancer.
2020, 48(2): 206-214
doi: 10.19756/j.issn.0253-3820.191394
Abstract:
A simple and sensitive method for quantification of transferrin (Tf) in human serum by high performance liquid chromatography (HPLC) coupled with inductively coupled plasma mass spectrometer (ICP-MS) with gold nanoparticles (AuNPs) labeling was established. Accurate quantification of standard Tf and Tf in human serum were achieved by measuring the concentration of AuNPs labeled on Tf. Tf could be incubated with most amount of AuNPs to form AuNPs-Tf in phosphate buffer (pH 6.8). Since gold atoms labeled to the Tf molecule could enhance the signals, accurate quantification of Tf in human serum could be achieved by gold signal measurement with ICP-MS after AuNPs-Tf was separated by HPLC. Good linear correlation coefficient (R2=0.9959) was obtained between Au peak area on AuNPs-Tf and concentration of Tf with a wide linear response over 3 orders of magnitude, and the limit of detection was 6 ng/mL. The developed method was successfully applied for detection of Tf in human serum certified reference material (ERM-DA470/IFCC) after being verified by standard Tf. Good agreement was achieved and the quantitative recovery was in the range of 95.2%-102.6%. Due to signal amplification of AuNPs and ultra-sensitive detection of ICP-MS, the sensitivity and accuracy were improved significantly compared to traditional enzyme-linked immunosorbent assay. Moreover, the measurement results could be directly traced to SI unit.
A simple and sensitive method for quantification of transferrin (Tf) in human serum by high performance liquid chromatography (HPLC) coupled with inductively coupled plasma mass spectrometer (ICP-MS) with gold nanoparticles (AuNPs) labeling was established. Accurate quantification of standard Tf and Tf in human serum were achieved by measuring the concentration of AuNPs labeled on Tf. Tf could be incubated with most amount of AuNPs to form AuNPs-Tf in phosphate buffer (pH 6.8). Since gold atoms labeled to the Tf molecule could enhance the signals, accurate quantification of Tf in human serum could be achieved by gold signal measurement with ICP-MS after AuNPs-Tf was separated by HPLC. Good linear correlation coefficient (R2=0.9959) was obtained between Au peak area on AuNPs-Tf and concentration of Tf with a wide linear response over 3 orders of magnitude, and the limit of detection was 6 ng/mL. The developed method was successfully applied for detection of Tf in human serum certified reference material (ERM-DA470/IFCC) after being verified by standard Tf. Good agreement was achieved and the quantitative recovery was in the range of 95.2%-102.6%. Due to signal amplification of AuNPs and ultra-sensitive detection of ICP-MS, the sensitivity and accuracy were improved significantly compared to traditional enzyme-linked immunosorbent assay. Moreover, the measurement results could be directly traced to SI unit.
2020, 48(2): 215-223
doi: 10.19756/j.issn.0253-3820.191554
Abstract:
Taxifolin (TFL), quercetin (QCT) and morin hydrate (MH) are common flavonoids in nature. They have many biological activities, such as antioxidant, anti-inflammatory, anti-allergic, anti-tumor and so on, but these compounds have poor water solubility and low oral utilization rate, which hinder the pharmacological action of these compounds. The bridged bis(β-cyclodextrin)s (bis(β-CD)s) has been known as it can significantly alter the molecular binding ability and the water solubility toward drug molecules, through the cooperative binding of drug molecule by two hydrophobic cavities located in close vicinity. In this work, the inclusion behavior of TFL, QCT and MH with diaminopropane bridged bis (β-CD)s) (H2) in solution and solid phase was studied. The three inclusion complexes (TFL/H2, QCT/H2 and MH/H2) were characterized by ultraviolet visible spectrum (UV-VIS), 1H nuclear magnetic resonance spectrum (1H NMR), two-dimensional nuclear magnetic resonance spectrum (2D NMR), scanning electron microscope (SEM), X-Ray diffraction spectrum (XRD) and Fourier transform infrared spectroscopy (FT-IR). The results showed that the A ring and C ring of the three flavonoids were separately included in two β-CD cavities from narrow side of H2, forming a 1:1 cooperative sandwich binding mode. The order of stability constant (KS) of the three inclusion complexes was KMH/H2 > KTFL/H2 > KQCT/H2. The inclusion complex stability depended greatly on the host-guest size, shape matching relation and hydrogen bond interaction force. After the formation of inclusion complex, the solubility of TFL, QCT and MH in water increased by 130.2, 180.4 and 210.1 times, respectively. In addition, the antioxidant activities were determined by 1-diphenyl-2-trinitrophenylhydrazide (DPPH) radical scavenging method. Excitingly, the antioxidant activities of TFL/H2and MH/H2 inclusion complexes presented a satisfactory antioxidant activity, which was even higher than that of free drugs. The study provided an important reference for the development of flavonoids with high water solubility and good antioxidant activity.
Taxifolin (TFL), quercetin (QCT) and morin hydrate (MH) are common flavonoids in nature. They have many biological activities, such as antioxidant, anti-inflammatory, anti-allergic, anti-tumor and so on, but these compounds have poor water solubility and low oral utilization rate, which hinder the pharmacological action of these compounds. The bridged bis(β-cyclodextrin)s (bis(β-CD)s) has been known as it can significantly alter the molecular binding ability and the water solubility toward drug molecules, through the cooperative binding of drug molecule by two hydrophobic cavities located in close vicinity. In this work, the inclusion behavior of TFL, QCT and MH with diaminopropane bridged bis (β-CD)s) (H2) in solution and solid phase was studied. The three inclusion complexes (TFL/H2, QCT/H2 and MH/H2) were characterized by ultraviolet visible spectrum (UV-VIS), 1H nuclear magnetic resonance spectrum (1H NMR), two-dimensional nuclear magnetic resonance spectrum (2D NMR), scanning electron microscope (SEM), X-Ray diffraction spectrum (XRD) and Fourier transform infrared spectroscopy (FT-IR). The results showed that the A ring and C ring of the three flavonoids were separately included in two β-CD cavities from narrow side of H2, forming a 1:1 cooperative sandwich binding mode. The order of stability constant (KS) of the three inclusion complexes was KMH/H2 > KTFL/H2 > KQCT/H2. The inclusion complex stability depended greatly on the host-guest size, shape matching relation and hydrogen bond interaction force. After the formation of inclusion complex, the solubility of TFL, QCT and MH in water increased by 130.2, 180.4 and 210.1 times, respectively. In addition, the antioxidant activities were determined by 1-diphenyl-2-trinitrophenylhydrazide (DPPH) radical scavenging method. Excitingly, the antioxidant activities of TFL/H2and MH/H2 inclusion complexes presented a satisfactory antioxidant activity, which was even higher than that of free drugs. The study provided an important reference for the development of flavonoids with high water solubility and good antioxidant activity.
2020, 48(2): 224-232
doi: 10.19756/j.issn.0253-3820.191600
Abstract:
A non-enzymatic superoxide anion (O2·-) electrochemical sensor (Co3[Fe(CN)6]2-RGO/GCE) was fabricated by co-depositing cobalt ferricyanide (Co3[Fe(CN)6]2) prepared by coprecipitation method and reduced graphene oxide (RGO) prepared by Hummers method on a glassy carbon electrode (GCE). The characteristics of Co3[Fe(CN)6]2-RGO nanocomposites and the possibility for electrochemical detection of O2·- were studied using scanning electron microscopy (SEM), X-ray diffraction (XRD), electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The mechanism of Co3[Fe(CN)6]2-RGO for O2·- detection was investigated in this work. The sensor had a sensitivity of 91.8 μA L/(μmol cm2) and a detection limit of 0.071 μmol/L (S/N=3). Besides, it had excellent selectivity and could avoid the interference of hydrogen peroxide, oxygen and other electrochemical active substances in vivo. It was successfully applied to the electrochemical detection of superoxide anions in stimulated MCF-7 cells. The sensor had potential application for quantifying superoxide anions in the field of pathology and biology.
A non-enzymatic superoxide anion (O2·-) electrochemical sensor (Co3[Fe(CN)6]2-RGO/GCE) was fabricated by co-depositing cobalt ferricyanide (Co3[Fe(CN)6]2) prepared by coprecipitation method and reduced graphene oxide (RGO) prepared by Hummers method on a glassy carbon electrode (GCE). The characteristics of Co3[Fe(CN)6]2-RGO nanocomposites and the possibility for electrochemical detection of O2·- were studied using scanning electron microscopy (SEM), X-ray diffraction (XRD), electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The mechanism of Co3[Fe(CN)6]2-RGO for O2·- detection was investigated in this work. The sensor had a sensitivity of 91.8 μA L/(μmol cm2) and a detection limit of 0.071 μmol/L (S/N=3). Besides, it had excellent selectivity and could avoid the interference of hydrogen peroxide, oxygen and other electrochemical active substances in vivo. It was successfully applied to the electrochemical detection of superoxide anions in stimulated MCF-7 cells. The sensor had potential application for quantifying superoxide anions in the field of pathology and biology.
2020, 48(2): 233-239
doi: 10.19756/j.issn.0253-3820.191681
Abstract:
Protein phosphorylation is one of the most common and important post-translational modifications, which plays key regulatory roles in biological processes, such as cell signaling and apoptosis. However, due to the low abundance and negative charge of phosphorylated proteins, in the conventional positive ion mode of bio-mass spectrometry, phosphoproteins are susceptible to be suppressed by high-abundance non-phosphorylated proteins, making them difficult to be directly analyzed. Therefore, the enrichment of phosphorylated proteins/peptides prior to mass spectrometry is important in biological research. For the first time, in this study, a novel zirconium ion-immobilized affinity magnetic nanoparticle (Zr4+-ATP-MNP) was prepared using adenosine triphosphate (ATP) as the ligand. The ATP ligand in Zr4+-ATP-MNP contains three phosphate groups, which can simultaneously provide multiple zirconium ion chelating sites, thus form stable network compounds with zirconium ions and provide high activity towards phosphorylated peptide. In addition, the hydrophilic pentose (ribose) and purine (adenine) parts in the ATP structure can significantly increase the hydrophilicity of the material and reduce the non-specific adsorption from non-phosphorylated peptides. The results of scanning electron microscopy and elemental analysis indicated that Zr4+-ATP-MNP was successfully prepared. Using β-casein digest as the sample, 9 phosphopeptides were identified. From digest mixture of β-casein and BSA with molar ratio as low as 1:100, five phosphopeptides were effectively identified with significantly improved signal and the non-phosphorylated peptides were effectively removed. The above results demonstrated the high specificity of Zr4+-ATP-MNP for both monophosphorylated peptides and multiphosphorylated peptides. The Zr4+-ATP-MNP was further applied to capture phosphopeptides in milk digest. All these results indicated the promising potential of Zr4+-ATP-MNP to capture mono- and multi-phosphopeptides in complex biological samples.
Protein phosphorylation is one of the most common and important post-translational modifications, which plays key regulatory roles in biological processes, such as cell signaling and apoptosis. However, due to the low abundance and negative charge of phosphorylated proteins, in the conventional positive ion mode of bio-mass spectrometry, phosphoproteins are susceptible to be suppressed by high-abundance non-phosphorylated proteins, making them difficult to be directly analyzed. Therefore, the enrichment of phosphorylated proteins/peptides prior to mass spectrometry is important in biological research. For the first time, in this study, a novel zirconium ion-immobilized affinity magnetic nanoparticle (Zr4+-ATP-MNP) was prepared using adenosine triphosphate (ATP) as the ligand. The ATP ligand in Zr4+-ATP-MNP contains three phosphate groups, which can simultaneously provide multiple zirconium ion chelating sites, thus form stable network compounds with zirconium ions and provide high activity towards phosphorylated peptide. In addition, the hydrophilic pentose (ribose) and purine (adenine) parts in the ATP structure can significantly increase the hydrophilicity of the material and reduce the non-specific adsorption from non-phosphorylated peptides. The results of scanning electron microscopy and elemental analysis indicated that Zr4+-ATP-MNP was successfully prepared. Using β-casein digest as the sample, 9 phosphopeptides were identified. From digest mixture of β-casein and BSA with molar ratio as low as 1:100, five phosphopeptides were effectively identified with significantly improved signal and the non-phosphorylated peptides were effectively removed. The above results demonstrated the high specificity of Zr4+-ATP-MNP for both monophosphorylated peptides and multiphosphorylated peptides. The Zr4+-ATP-MNP was further applied to capture phosphopeptides in milk digest. All these results indicated the promising potential of Zr4+-ATP-MNP to capture mono- and multi-phosphopeptides in complex biological samples.
2020, 48(2): 240-247
doi: 10.19756/j.issn.0253-3820.191559
Abstract:
Nitrogen-doped graphene quantum dots (NGQDs) were synthesized by hydrothermal method with citric acid and ammonia as raw materials. A solid-state electrochemiluminescence (ECL) sensor was thus constructed by dropping NGQDs/CdS nanocluster (NCs) composites. The properties of NGQDs and CdS NCs were characterized by UV-Vis absorption spectroscopy and fluorescence spectroscopy. The electrochemiluminescent and electrochemical behaviors of the sensor were studied. The results showed that NGQDs could enhance electrochemiluminescence signal of CdS NCs and the stability of NGQDs/CdS NCs increased when H2S was used as the co-reaction reagent of CdS NCs. In the presence of H2S, S2- bonded with excess Cd2+, and the ECL intensity decreased. Under the optimal conditions, the decrease in electrochemiluminescence intensity varied proportionally with the logarithmic concentration of H2S from 2.0×10-10-2.0×10-5 mol/L, and the detection limit was estimated to be 6.7×10-11 mol/L. The sensor was used for the determination of H2S in bovine serum with recoveries of 92.7%-103.8%, which indicated that the proposed sensor had the potential for detection of H2S in practical complex samples.
Nitrogen-doped graphene quantum dots (NGQDs) were synthesized by hydrothermal method with citric acid and ammonia as raw materials. A solid-state electrochemiluminescence (ECL) sensor was thus constructed by dropping NGQDs/CdS nanocluster (NCs) composites. The properties of NGQDs and CdS NCs were characterized by UV-Vis absorption spectroscopy and fluorescence spectroscopy. The electrochemiluminescent and electrochemical behaviors of the sensor were studied. The results showed that NGQDs could enhance electrochemiluminescence signal of CdS NCs and the stability of NGQDs/CdS NCs increased when H2S was used as the co-reaction reagent of CdS NCs. In the presence of H2S, S2- bonded with excess Cd2+, and the ECL intensity decreased. Under the optimal conditions, the decrease in electrochemiluminescence intensity varied proportionally with the logarithmic concentration of H2S from 2.0×10-10-2.0×10-5 mol/L, and the detection limit was estimated to be 6.7×10-11 mol/L. The sensor was used for the determination of H2S in bovine serum with recoveries of 92.7%-103.8%, which indicated that the proposed sensor had the potential for detection of H2S in practical complex samples.
2020, 48(2): 248-254
doi: 10.19756/j.issn.0253-3820.191491
Abstract:
A fluorescence sensor for Pb2+ detection was developed on the basis of a DNAzyme cleavage and exonuclease Ⅲ assistant amplification strategy. In the presence of Pb2+, the DNAzyme hybridized with substrate strand and catalyzed the hydrolytic cleavage of the substrate strand, and then the DNAzyme released from the substrate strand and bound another substrate strand to trigger another cycle of hydrolytic cleavage. The DNAzymes were used as catalysts for amplified sensing through multiple turnover reactions. The substrate probe was cleavaged and broken to form a Y-shaped probe which could hybridize and open the molecular beacon, resulting in the increase of the fluorescence signal. At the same time, exonuclease Ⅲ catalytically digested the molecular beacon from 3'-end and released the Y-shaped substrate strand. The released Y-shaped substrate strand could directly hybridize with another molecular beacon to generate fluorescence signal, and thus was further recognized and cleaved by exonuclease Ⅲ from the second step of cyclic signal amplification. Accompanying with each cleavage toward molecular probe by exonuclease Ⅲ, the fluorescence signal was accumulated, which resulted in a cyclic amplification format for the fluorescence response toward Pb2+ detection. The fluorescence response was detected in a 200 μL reaction system that was incubated at 37℃ for 60 min. The linear range for detection of Pb2+ was 0.05-200 nmol/L with a detection limit of 0.01 nmol/L, and the recoveries of environmental water samples were 96.3%-108.3%. This method had many advantages such as simple operation, rapid detection, high selectivity and high sensitivity, and showed great application potential in Pb2+ detection.
A fluorescence sensor for Pb2+ detection was developed on the basis of a DNAzyme cleavage and exonuclease Ⅲ assistant amplification strategy. In the presence of Pb2+, the DNAzyme hybridized with substrate strand and catalyzed the hydrolytic cleavage of the substrate strand, and then the DNAzyme released from the substrate strand and bound another substrate strand to trigger another cycle of hydrolytic cleavage. The DNAzymes were used as catalysts for amplified sensing through multiple turnover reactions. The substrate probe was cleavaged and broken to form a Y-shaped probe which could hybridize and open the molecular beacon, resulting in the increase of the fluorescence signal. At the same time, exonuclease Ⅲ catalytically digested the molecular beacon from 3'-end and released the Y-shaped substrate strand. The released Y-shaped substrate strand could directly hybridize with another molecular beacon to generate fluorescence signal, and thus was further recognized and cleaved by exonuclease Ⅲ from the second step of cyclic signal amplification. Accompanying with each cleavage toward molecular probe by exonuclease Ⅲ, the fluorescence signal was accumulated, which resulted in a cyclic amplification format for the fluorescence response toward Pb2+ detection. The fluorescence response was detected in a 200 μL reaction system that was incubated at 37℃ for 60 min. The linear range for detection of Pb2+ was 0.05-200 nmol/L with a detection limit of 0.01 nmol/L, and the recoveries of environmental water samples were 96.3%-108.3%. This method had many advantages such as simple operation, rapid detection, high selectivity and high sensitivity, and showed great application potential in Pb2+ detection.
2020, 48(2): 255-261
doi: 10.19756/j.issn.0253-3820.191346
Abstract:
A method for rapid detection of 11 kinds of bisphenols in plastic food contact materials based on ultra-performance convergence chromatography (UPC2) was established. Plastic food contact material sample was extracted with methanol by ultrasonic treatment, followed by clean-up through a 0.22 μm organic membrane. The separation was conducted on ACQUITY UPC2 Torus DIOL chromatographic column (150 mm×3.0 mm, 1.7 mm) with supercritical carbon dioxide as the main mobile phase and methanol as a modified co-solvent for gradient elution. The 11 kinds of bisphenols were analyzed by UV detector, and quantified by external standard method. The 11 kinds of bisphenols were separated within 5 min under the optimum conditions, and exhibited a good linearity with the standard curve correlation coefficients (R2) of not less than 0.998. The recoveries at three standard levels (1.0, 25.0 and 50.0 mg/kg) were 75.7%-122.0%, the relative standard deviation was 0.6%-7.3%, and the detection limits (LODs) were 0.1-0.3 mg/kg. This method is convenient, accurate, green and efficient, and can be used for the rapid detection of 11 kinds of bisphenols in plastic food contact materials.
A method for rapid detection of 11 kinds of bisphenols in plastic food contact materials based on ultra-performance convergence chromatography (UPC2) was established. Plastic food contact material sample was extracted with methanol by ultrasonic treatment, followed by clean-up through a 0.22 μm organic membrane. The separation was conducted on ACQUITY UPC2 Torus DIOL chromatographic column (150 mm×3.0 mm, 1.7 mm) with supercritical carbon dioxide as the main mobile phase and methanol as a modified co-solvent for gradient elution. The 11 kinds of bisphenols were analyzed by UV detector, and quantified by external standard method. The 11 kinds of bisphenols were separated within 5 min under the optimum conditions, and exhibited a good linearity with the standard curve correlation coefficients (R2) of not less than 0.998. The recoveries at three standard levels (1.0, 25.0 and 50.0 mg/kg) were 75.7%-122.0%, the relative standard deviation was 0.6%-7.3%, and the detection limits (LODs) were 0.1-0.3 mg/kg. This method is convenient, accurate, green and efficient, and can be used for the rapid detection of 11 kinds of bisphenols in plastic food contact materials.
2020, 48(2): 262-268
doi: 10.19756/j.issn.0253-3820.191570
Abstract:
High-performance liquid chromatography time-of-flight mass spectrometry (HPLC-TOF-MS) was employed to analyze the extract of total flavones of Abelmoschus manihot L. Medic (TFA) in both positive and negative ion mode. The raw LC-MS data were filtered by a well-established mass defect filter (MDF) approach to screen the flavone-related ions, and then, each screened ion was assigned by TOF-MS accurate mass measurement to certain elemental composition and ion type. Next, the co-eluted ions from one single compound were attempted to be combined, and the presence of each compound was confirmed by at least two quasi-molecular ions. Aided by authentic standards comparison and searching literature and databases, the targeted compounds were tentatively identified or assigned according to their parent and fragment ions. Finally, 49 flavones were screened and tentatively identified or assigned from the TFA extract. The results showed that, glycosylation, methylation, formylation, acetylation and glucuronidation were considered to be the major conversion involved in the formation of the TFA flavones. The current study not only provided the sufficient chemical substance for further studies of TFA, such as quality control and pharmacological activity, but also demonstrated the potential of the combination of LC-TOF-MS and MDF for systematically screening the constituents of herbal medicines.
High-performance liquid chromatography time-of-flight mass spectrometry (HPLC-TOF-MS) was employed to analyze the extract of total flavones of Abelmoschus manihot L. Medic (TFA) in both positive and negative ion mode. The raw LC-MS data were filtered by a well-established mass defect filter (MDF) approach to screen the flavone-related ions, and then, each screened ion was assigned by TOF-MS accurate mass measurement to certain elemental composition and ion type. Next, the co-eluted ions from one single compound were attempted to be combined, and the presence of each compound was confirmed by at least two quasi-molecular ions. Aided by authentic standards comparison and searching literature and databases, the targeted compounds were tentatively identified or assigned according to their parent and fragment ions. Finally, 49 flavones were screened and tentatively identified or assigned from the TFA extract. The results showed that, glycosylation, methylation, formylation, acetylation and glucuronidation were considered to be the major conversion involved in the formation of the TFA flavones. The current study not only provided the sufficient chemical substance for further studies of TFA, such as quality control and pharmacological activity, but also demonstrated the potential of the combination of LC-TOF-MS and MDF for systematically screening the constituents of herbal medicines.
2020, 48(2): 269-274
doi: 10.19756/j.issn.0253-3820.191400
Abstract:
The core-shell nanoparticles (i.e., Au-core@4-mercaptobenzoic acid@Ag-shell) were combined with spectral shape deformation (SSD) quantitative model for quantification of 6-thioguanine (6-TG) in urine and red blood cell lysate samples. Experimental results showed that the combination of core-shell nanoparticles with SSD model could effectively remove the detrimental multiplicative effects on samples' surface enhanced Raman spectroscopy (SERS) signals caused by the variations of the number of "hot spots" and their distribution near/on the surfaces of SERS enhancing substrates, and hence realized the accurate quantitative determination of 6-TG in urine and red blood cell lysate samples with recovery of 95.6%-106.7%. The limit of detection (3σ) and limit of quantification (10σ) for 6-TG were estimated to be about 1 and 3 nmol/L, respectively. Compared with conventional methods for analysis of 6-TG such as high performance liquid chromatography (HPLC), the proposed method had many advantages such as simplicity, rapidity and high sensitivity, and had great potential in routine quantitative analysis of 6-TG in complex samples.
The core-shell nanoparticles (i.e., Au-core@4-mercaptobenzoic acid@Ag-shell) were combined with spectral shape deformation (SSD) quantitative model for quantification of 6-thioguanine (6-TG) in urine and red blood cell lysate samples. Experimental results showed that the combination of core-shell nanoparticles with SSD model could effectively remove the detrimental multiplicative effects on samples' surface enhanced Raman spectroscopy (SERS) signals caused by the variations of the number of "hot spots" and their distribution near/on the surfaces of SERS enhancing substrates, and hence realized the accurate quantitative determination of 6-TG in urine and red blood cell lysate samples with recovery of 95.6%-106.7%. The limit of detection (3σ) and limit of quantification (10σ) for 6-TG were estimated to be about 1 and 3 nmol/L, respectively. Compared with conventional methods for analysis of 6-TG such as high performance liquid chromatography (HPLC), the proposed method had many advantages such as simplicity, rapidity and high sensitivity, and had great potential in routine quantitative analysis of 6-TG in complex samples.
2020, 48(2): 275-281
doi: 10.19756/j.issn.0253-3820.191299
Abstract:
The contents of four oxides in cement raw meal were determined by near infrared spectroscopy. In this method, the near infrared spectrum was collected by diffuse reflectance method, and X-ray fluorescence spectroscopy analysis was used to determine the reference values of oxides content. The outliers were eliminated according to the Mahalanobis distance, and then the samples were divided into calibration subset and validation subset by sample set partitioning based on joint X-Y distance (SPXY) method. The optimal wavelength variables were selected by backward interval partial least squares and genetic algorithm. Quantitative calibration models of four oxides were established by partial least squares algorithm, which showed good prediction performance. The correlation coefficients of validation subsets of CaO, SiO2, Al2O3 and Fe2O3 models were 0.9411, 0.9337, 0.8612 and 0.7351, the root mean square error of prediction were 0.0994, 0.1044, 0.0693 and 0.0387, the average absolute errors were 0.075%, 0.083%, 0.051% and 0.025%, respectively. By comparing with prompt gamma-ray neutron activation analysis and laser-induced breakdown spectroscopy analysis, the near infrared spectroscopy analysis has short time-consuming, highest accuracy, and best effect. It is suitable for rapid and accurate determination of contents of four oxides in cement raw meal, and provides a new idea for quality monitoring of cement raw meal.
The contents of four oxides in cement raw meal were determined by near infrared spectroscopy. In this method, the near infrared spectrum was collected by diffuse reflectance method, and X-ray fluorescence spectroscopy analysis was used to determine the reference values of oxides content. The outliers were eliminated according to the Mahalanobis distance, and then the samples were divided into calibration subset and validation subset by sample set partitioning based on joint X-Y distance (SPXY) method. The optimal wavelength variables were selected by backward interval partial least squares and genetic algorithm. Quantitative calibration models of four oxides were established by partial least squares algorithm, which showed good prediction performance. The correlation coefficients of validation subsets of CaO, SiO2, Al2O3 and Fe2O3 models were 0.9411, 0.9337, 0.8612 and 0.7351, the root mean square error of prediction were 0.0994, 0.1044, 0.0693 and 0.0387, the average absolute errors were 0.075%, 0.083%, 0.051% and 0.025%, respectively. By comparing with prompt gamma-ray neutron activation analysis and laser-induced breakdown spectroscopy analysis, the near infrared spectroscopy analysis has short time-consuming, highest accuracy, and best effect. It is suitable for rapid and accurate determination of contents of four oxides in cement raw meal, and provides a new idea for quality monitoring of cement raw meal.
2020, 48(2): 282-288
doi: 10.19756/j.issn.0253-3820.191352
Abstract:
The silver active substrates with average particle size of about 20 nm for surface-enhanced Raman spectroscopic analysis were prepared using cyanide-free electrochemical deposition method. The electrochemical deposition behavior of the system was studied by cyclic voltammetry at different scanning rates. The results showed that the electrodeposition process of silver was controlled by diffusion. With the increase of scanning rate, the oxidation peak slightly shifted positively, whereas the reduction peak had a significant negative shift. The result of scanning electron microscopy showed that the silver particles were fine, uniform, and regular in shape, and their size was (20±10) nm. Using methyl orange as a probe, the enhancement of Raman spectral signal with the silver-deposited substrate was investigated, and an enhancement factor of 5.2×105 was obtained. The linear range for detection of rhodamine 6G was 1×10-12-1×10-6 mol/L, the correlation coefficient was 0.98, and the limit of the detection was lower than 1×10-12 mol/L. Raman spectroscopy mapping also showed that the silver-deposited substrate had a uniform distribution.
The silver active substrates with average particle size of about 20 nm for surface-enhanced Raman spectroscopic analysis were prepared using cyanide-free electrochemical deposition method. The electrochemical deposition behavior of the system was studied by cyclic voltammetry at different scanning rates. The results showed that the electrodeposition process of silver was controlled by diffusion. With the increase of scanning rate, the oxidation peak slightly shifted positively, whereas the reduction peak had a significant negative shift. The result of scanning electron microscopy showed that the silver particles were fine, uniform, and regular in shape, and their size was (20±10) nm. Using methyl orange as a probe, the enhancement of Raman spectral signal with the silver-deposited substrate was investigated, and an enhancement factor of 5.2×105 was obtained. The linear range for detection of rhodamine 6G was 1×10-12-1×10-6 mol/L, the correlation coefficient was 0.98, and the limit of the detection was lower than 1×10-12 mol/L. Raman spectroscopy mapping also showed that the silver-deposited substrate had a uniform distribution.
2020, 48(2): 289-297
doi: 10.19756/j.issn.0253-3820.191329
Abstract:
The visible-near-infrared (Vis-NIR) transmission spectroscopy technique was used to analyze the content of S-ovalbumin (S-ova), which had high correlation with egg freshness, and to establish a nondestructive prediction model. The visible/near-infrared fiber spectroscopy were used to collect the transmission spectrum of two varieties of eggs at 349-1000 nm, and the S-ovalbumin content of 270 eggs was measured by wet chemistry method. By comparing the average spectra of eggs of different varieties during storage, it was found that the spectral absorption peaks of different varieties of eggs had the same position, and only the spectral energy values in the visible range differed. The original spectrum was preprocessed by standard normal variate (SNV), and 67 characteristic wavelengths were extracted from the full spectrum of 500-950 nm using uninformative variables elimination (UVE). It was concluded that partial least squares (PLS) regression model based on 67 characteristic wavelengths could predict the S-ovalbumin content. To further eliminate the multi-collinearity between the characteristic wavelengths, the stepwise regression algorithm was used to perform secondary screening on the characteristic wavelengths, and finally 16 characteristic wavelengths were selected. By using the 16 characteristic wavelengths to establish a multivariate regression model, the coefficient of determination (R2) of the training set was 0.9511, the root mean square error (RMSE) was 0.0478, and the R2 of the prediction set was 0.8380. Besides, the RMSE was 0.1116, and the residual predictive deviation (RPD) was 2.2620. The general predictive model was used to predict the S-ovalbumin content of 50 eggs with Roman pink shell and 40 eggs with sea blue brown shell in the prediction set. The R2 of the predicted and measured values were 0.8119 and 0.9116, respectively, and the RMSEs were 0.1298 and 0.0834, respectively. Therefore, the general model could perform nondestructive testing on the S-ovalbumin content of these two different varieties of eggs better, and the model was more applicable. The results showed that the visible/near-infrared spectroscopy could accurately detect the S-ovalbumin content of eggs in different varieties, and the established general prediction model laid a foundation for the development of portable non-destructive testing device for protein content.
The visible-near-infrared (Vis-NIR) transmission spectroscopy technique was used to analyze the content of S-ovalbumin (S-ova), which had high correlation with egg freshness, and to establish a nondestructive prediction model. The visible/near-infrared fiber spectroscopy were used to collect the transmission spectrum of two varieties of eggs at 349-1000 nm, and the S-ovalbumin content of 270 eggs was measured by wet chemistry method. By comparing the average spectra of eggs of different varieties during storage, it was found that the spectral absorption peaks of different varieties of eggs had the same position, and only the spectral energy values in the visible range differed. The original spectrum was preprocessed by standard normal variate (SNV), and 67 characteristic wavelengths were extracted from the full spectrum of 500-950 nm using uninformative variables elimination (UVE). It was concluded that partial least squares (PLS) regression model based on 67 characteristic wavelengths could predict the S-ovalbumin content. To further eliminate the multi-collinearity between the characteristic wavelengths, the stepwise regression algorithm was used to perform secondary screening on the characteristic wavelengths, and finally 16 characteristic wavelengths were selected. By using the 16 characteristic wavelengths to establish a multivariate regression model, the coefficient of determination (R2) of the training set was 0.9511, the root mean square error (RMSE) was 0.0478, and the R2 of the prediction set was 0.8380. Besides, the RMSE was 0.1116, and the residual predictive deviation (RPD) was 2.2620. The general predictive model was used to predict the S-ovalbumin content of 50 eggs with Roman pink shell and 40 eggs with sea blue brown shell in the prediction set. The R2 of the predicted and measured values were 0.8119 and 0.9116, respectively, and the RMSEs were 0.1298 and 0.0834, respectively. Therefore, the general model could perform nondestructive testing on the S-ovalbumin content of these two different varieties of eggs better, and the model was more applicable. The results showed that the visible/near-infrared spectroscopy could accurately detect the S-ovalbumin content of eggs in different varieties, and the established general prediction model laid a foundation for the development of portable non-destructive testing device for protein content.
2020, 48(2): 298-305
doi: 10.19756/j.issn.0253-3820.191256
Abstract:
Raman spectral data contain fingerprint spectral information corresponding to the components of the measured substances, which is an effective method to identify the components of mixtures. The traditional mixture component detection methods based on Raman spectrum have some issues such as the difficulty in extracting spectral features, the performance of search and peak matching algorithms is easily affected by the database, and the recognition accuracy is difficult to guarantee. To overcome these problems, a method of mixture components recognition using Raman spectrum based on sparse non-negative least squares algorithm is proposed. In this method, the spectral data of the mixture to be recognized is regarded as the linear representation of the spectral data of all kinds of pure substances. Considering the sparse characteristic of the mixture components quantity relative to the pure substance quantity in the database, the linear representation coefficient of the mixture spectrum in the pure substance spectrum data is obtained by the sparse non-negative least squares algorithm. And the suspected components are determined according to the statistical 2δ principle. On this basis, the iterative least squares algorithm combined with the T-distribution test method is used to realize the final identification of the mixture components. The Raman spectral data of 500 pure substances are used to build a standard database for identification of the experimental sample of 19 mixtures with equal volume ratio of components and 81 mixtures with different volume ratio of components. The results show that the precision is 90.24% and the recall is 88.10% under the condition of equal volume ratios, and the precision is 93.22% and the recall is 83.65% under the condition of different volume ratios, which proves the good stability and accuracy of the proposed algorithm.
Raman spectral data contain fingerprint spectral information corresponding to the components of the measured substances, which is an effective method to identify the components of mixtures. The traditional mixture component detection methods based on Raman spectrum have some issues such as the difficulty in extracting spectral features, the performance of search and peak matching algorithms is easily affected by the database, and the recognition accuracy is difficult to guarantee. To overcome these problems, a method of mixture components recognition using Raman spectrum based on sparse non-negative least squares algorithm is proposed. In this method, the spectral data of the mixture to be recognized is regarded as the linear representation of the spectral data of all kinds of pure substances. Considering the sparse characteristic of the mixture components quantity relative to the pure substance quantity in the database, the linear representation coefficient of the mixture spectrum in the pure substance spectrum data is obtained by the sparse non-negative least squares algorithm. And the suspected components are determined according to the statistical 2δ principle. On this basis, the iterative least squares algorithm combined with the T-distribution test method is used to realize the final identification of the mixture components. The Raman spectral data of 500 pure substances are used to build a standard database for identification of the experimental sample of 19 mixtures with equal volume ratio of components and 81 mixtures with different volume ratio of components. The results show that the precision is 90.24% and the recall is 88.10% under the condition of equal volume ratios, and the precision is 93.22% and the recall is 83.65% under the condition of different volume ratios, which proves the good stability and accuracy of the proposed algorithm.
