A rapid UHPLC-QqQ-MS/MS method for the simultaneous qualitation and quantitation of coumarins, furocoumarins, flavonoids, phenolic acids in pummelo fruits.

The impact of secondary metabolites on fruit quality, plant growth and human health has led to an increased demand for analytical methods to characterize and quantify these metabolites in recent years. A versatile, sensitive and rapid method based on UHPLC-QqQ-MS/MS was developed for simultaneous qualitation and quantitation of coumarins, furocoumarins, flavonoids and phenolic acids. The chromatographic elution and multiple reaction monitoring mode transitions were optimized to achieve good separation and accurate quantitation of 47 analytes, including 13 groups of isomers, during a single 13 min chromatographic run. This method was validated with good precision and recoveries, wide linear ranges and low limits of detection and quantitation (0.014-1.50 µg L-1). The validated method was further applied to quantify the analytes in flavedo, albedo and pulp from two pummelo varieties, C. grandis 'Shatianyu' and C. grandis 'Guanximiyu'. This method combines high sensitivity, good selectivity, and short chromatographic run time.

Analysis of phytochemical contributors to antioxidant capacity of the peel of Chinese mandarin and orange varieties.

The phytochemicals in the peel of six oranges and ten mandarins including seven wild varieties and three cultivars were systematically characterised using UHPLC-Q-TOF-MS, and the correlation analysis was performed between phytochemicals and antioxidant capacity in order to investigate the phytochemical contributors to antioxidant capacity. The gradient elution was completed within 16 min and 92 compounds were undoubtedly or tentatively identified. Furthermore, the antioxidant capacities were determined using ABTS, DPPH and FRAP methods. The number of compounds, their contents and the antioxidant capacities were sequenced in the same order of the wild mandarins > cultivated mandarins > oranges. The correlation analysis that showed five compounds were significantly correlated with the antioxidant capacity and can act as main contributors to the citrus varieties with high antioxidant capacities. This study is systematic for the metabolites identification of mandarins and oranges and provides valuable information for effective utilisation of citrus peel and their bioactive compounds.

Efficient analysis of phytochemical constituents in the peel of Chinese wild citrus Mangshanju (Citrus reticulata Blanco) by ultra high performance liquid chromatography-quadrupole time-of-flight-mass spectrometry.

An efficient ultra high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry method was developed for separation and profiling of phytochemical constituents of Chinese wild mandarin Mangshanju (Citrus reticulata Blanco). All constituents were well separated within 16 min. Based on retention times, accurate mass, MSE fragments, and/or reference standards as well as databases, a total of 81 compounds were unambiguously identified or tentatively assigned including flavonoid glycosides, acylated flavonoid glycosides, flavones, polymethoxylated flavonoids, and limonoids as well as four other compounds. Among them, 22 polymethoxylated flavones and ten polymethoxylated flavanones/chalcones were identified in Mangshanju, more types than other citrus reported before. A basic procedure for identifying flavonoid-O-glycosides and the aglycones including polymethoxylated flavonoids was proposed. In addition, this method was successfully used to analyze another four mandarin germplasms, Cenxi suan ju, Xipi gousi gan, Nanfeng miju, and Or, showing that Mangshanju contained two characteristic compounds distinct from the other four citrus species. This study systematically profiled phytochemical constituents of Mangshanju, which was helpful for further utilization of Mangshanju owing to its abundant bioactive compounds.

Fast Separation and Sensitive Quantitation of Polymethoxylated Flavonoids in the Peels of Citrus Using UPLC-Q-TOF-MS.

A rapid, sensitive, and efficient ultraperformance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) method has been developed to analyze polymethoxylated flavonoids (PMFs) in 14 Citrus peels, including 7 Citrus reticulata (C. reticulata) and 7 Citrus sinensis (C. sinensis). In this study, fast separation can be achieved within 12 min and 42 PMFs have been identified including 33 flavones and 9 flavanones. Most C. reticulata were shown to contain more than 20 PMFs, except Guangxihongpisuanju (GX) containing only 12 PMFs, while most C. sinensis contained fewer than 20 PMFs, except Edangan (EG) containing as many as 32 PMFs. To our knowledge, there are few reports about the quantitation of PMFs using the MS response. Here, a MS quantitative method was established and systematically validated in linearity, precision, and recovery. The linearity was from 1.25 ng/mL to 1.0 µg/mL with the limit of detection (LOD) as low as 75 pg/mL and the limit of quantitation (LOQ) as low as 0.25 ng/mL. Up to 13 PMFs, more types than ever before, were undoubtedly identified and quantitated according to the PMF standards. The results showed that the contents of PMFs in the C. reticulata were generally higher than those in the C. sinensis. This study is systematic for analyzing PMFs and is of great significance because it can provide guidance on utilization of both PMFs and citrus germplasm resources in the future.

Lanosterol reverses protein aggregation in cataracts.

The human lens is comprised largely of crystallin proteins assembled into a highly ordered, interactive macro-structure essential for lens transparency and refractive index. Any disruption of intra- or inter-protein interactions will alter this delicate structure, exposing hydrophobic surfaces, with consequent protein aggregation and cataract formation. Cataracts are the most common cause of blindness worldwide, affecting tens of millions of people, and currently the only treatment is surgical removal of cataractous lenses. The precise mechanisms by which lens proteins both prevent aggregation and maintain lens transparency are largely unknown. Lanosterol is an amphipathic molecule enriched in the lens. It is synthesized by lanosterol synthase (LSS) in a key cyclization reaction of a cholesterol synthesis pathway. Here we identify two distinct homozygous LSS missense mutations (W581R and G588S) in two families with extensive congenital cataracts. Both of these mutations affect highly conserved amino acid residues and impair key catalytic functions of LSS. Engineered expression of wild-type, but not mutant, LSS prevents intracellular protein aggregation of various cataract-causing mutant crystallins. Treatment by lanosterol, but not cholesterol, significantly decreased preformed protein aggregates both in vitro and in cell-transfection experiments. We further show that lanosterol treatment could reduce cataract severity and increase transparency in dissected rabbit cataractous lenses in vitro and cataract severity in vivo in dogs. Our study identifies lanosterol as a key molecule in the prevention of lens protein aggregation and points to a novel strategy for cataract prevention and treatment.

Sodium hydroxide-mediated hydrogel of citrus pectin for preparation of fluorescent carbon dots for bioimaging.

The citrus process industry produces annually a huge amount of pomace, which is a rich source of citrus pectin. Here, we report the hydrogel of citrus pectin mediated by sodium hydroxide can be used to prepare fluorescent carbon dots (CDs). The introduction of hydrogel can not only make the temperature of the hydrothermal reaction down to 100 °C, but also avoid visually carbonized precipitates in the synthesis process even up to 180 °C. The as-synthesized CDs are well dispersed in water with an average size of 2.7 nm and show cyan fluorescence with high photostability, good biocompatibility. Furthermore, the CDs can act as a potential fluorescent probe for cell imaging. Citrus pectin as a non-toxic carbonaceous precursor for preparation of fluorescent CDs provides a new approach for the efficient utilization of citrus germplasm in future.

DNA-templated Ag nanoclusters as fluorescent probes for sensing and intracellular imaging of hydroxyl radicals.

We have developed a simple, rapid and label-free sensor for the essential biological OH radicals based on the fluorescence quenching of DNA-templated Ag nanoclusters (DNA-Ag NCs). The OH radicals generated from the Fenton reagent attack and cleave the DNA template, which disturbs the microenvironments around Ag NCs, resulting in spontaneous aggregation due to the lack of stabilization and further the quenching of the Ag NCs fluorescence. These changes in fluorescence intensity allow sensing of OH radicals with good sensitivity and selectivity under optimal conditions. The sensor can be also applied for quantifying the radical scavenging action of antioxidants. Various characterizations including absorption spectra, fluorescence lifetimes, light scattering (LS) spectra, transmission electron microscopy (TEM), dark field light scattering imaging, and circular dichroism (CD) spectrometry have been employed to illustrate the proposed sensing mechanism. Further investigations demonstrate that the fluorescent probe could penetrate into intact cell membranes to selectively detect intracellular OH radicals induced by the phorbol myristate acetate (PMA) stimulation. These advantageous characteristics make the fluorescent DNA-Ag NCs potentially useful as a new candidate to monitor OH in broad biosystems.

Metal-organic framework MIL-101 as a low background signal platform for label-free DNA detection.

A label-free and sensitive fluorescence method for recognition of sequence-specific DNA using DNA-intercalating dye and metal-organic frameworks (MOFs) is developed. Here, MIL-101 (Cr3F(H2O)2O[(O2C)-C6H4-(CO2)]3·nH2O) is introduced as a quenching platform to decrease the high background fluorescence of SYBR Green I (SG)/probe DNA complex. Mechanism investigations show that MIL-101 can strongly adsorb the SG/probe DNA complex through π-π stacking and electrostatic interactions, and as a consequence, the fluorescence of the SG dye is greatly quenched. While in the presence of target DNA, the as-formed rigid double-stranded (ds) structure of DNA will be far away from the surface of MIL-101; meanwhile, the SG dye can be bound with the dsDNA in the mode of intercalation and minor groove binding, resulting in enhancement of the SG dye fluorescence. The increased signal-to-background ratio has a linear relationship with the concentration of target DNA in the range of 0.1-14 nM. It is confirmed that the detection limit is 73 pM (3σ), which is much lower than that based on the carbon nanotubes and graphene oxide platform. Moreover, one-base-mismatched target DNA can be discriminated effectively. With the introduction of MIL-101, the signal-to-background ratio has been improved ∼8-fold, demonstrating that MIL-101 is an efficient low-background signal platform.

A nanosized metal-organic framework of Fe-MIL-88NH2 as a novel peroxidase mimic used for colorimetric detection of glucose.

In this paper, a nanosized porous metal-organic framework, Fe-MIL-88NH2, was facilely prepared with a uniform octahedral shape by the addition of acetic acid, and for the first time was demonstrated to possess intrinsic peroxidase-like activity. Kinetic analysis and electron spin resonance measurements indicated that the catalytic behavior was consistent with typical Michaelis-Menten kinetics and follows a ping-pong mechanism. As a novel peroxidase mimic material, Fe-MIL-88NH2 shows the advantages of high catalytic efficiency, ultrahigh stability and high biocompatibility in aqueous medium compared with natural enzymes and other peroxidase nanomimetics. Here, Fe-MIL-88NH2 was used to quickly catalyze oxidation of the peroxidase substrate 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of H2O2 to produce a colored product, which provided a simple, sensitive and selective method for the colorimetric detection of glucose. Glucose could be linearly detected in the range from 2.0 × 10⁻6 to 3.0 × 10⁻4 M with a detection limit of 4.8 × 10⁻7 M, and the color variation for glucose response was also obvious by visual observation at concentrations as low as 2.0 × 10⁻6 M. More importantly, the colorimetric method could be successfully applied to the determination of glucose in diluted serum samples.

Visual detection of tetracycline antibiotics with the turned on fluorescence induced by a metal-organic coordination polymer.

Here, a rapid, simple and sensitive method for visual detection of tetracycline antibiotics (TCs) was developed based on the turned on fluorescence induced by a metal-organic coordination polymer of Zn(bix) [bix=1,4-bis(imidazol-1-ylmethyl)benzene]. Tetracycline antibiotics, such as tetracycline hydrochloride (TC), doxycycline hyclate (DC), chlortetracycline hydrochloride (CTC) and oxytetracyline hydrochloride (OTC), showed very weak fluorescence in Tris-HCl buffer of pH 8.68-9.40. However, the fluorescence intensities of these TCs were greatly enhanced in the presence of Zn(bix) and were found to be proportional to the concentrations of TCs. Accordingly, TCs could be quantitatively determined with the detection limits of 12.0, 6.0, 19.0 and 19.0nM for TC, DC, CTC and OTC, respectively. Furthermore, this fluorimetric method was successfully applied to the determination of DC contents in doxycycline hyclate tablets from three factories with satisfactory results compared with the HPLC method. Also, the interaction mechanism between TCs and Zn(bix) was investigated by the absorption spectra, fluorescence spectra and circular dichroism spectra. The results showed that TCs could interact with Zn(bix) to form TCs-Zn(bix) ternary complexes and the turned on fluorescence was mainly ascribed to the restricted conformational rotation of TCs molecules through coordination with Zn(II) from the polymeric networks of Zn(bix).

Switching on fluorescence for selective visual recognition of naringenin and morin with a metal-organic coordination polymer of Zn(bix) [bix=1,4-bis(imidazol-1-ylmethyl)benzene].

Flavonoids such as naringenin and morin are ubiquitous in a wide range of foods isolated from plants, and have diverse effects on plants even on human health. Here, we establish a selective visual method for recognition of aringenin and morin based on the "switched on" fluorescence induced by a metal-organic coordination polymer of Zn(bix) [bix=1,4-bis(imidazol-1-ylmethyl)benzene]. Owing to the coordination interaction of aringenin and morin with Zn(II) from the polymeric structure of Zn(bix), the conformational free rotation of naringenin and morin is restricted leading to relatively rigid structures. And as a consequence, the fluorescence is switched on. While luteolin and quercetin, holding a very similar structure with naringenin and morin, have no such fluorescence enhancement most likely owing to the 3'-hydroxy substitution in the B ring. Under 365 nm UV lamp light, we can visually recognize and discriminate naringenin and morin from them each other and luteolin as well as quercetin based on the colors of their emission. With this recognition system, the detection of naringenin and morin in human urine was made with satisfactory results.

Highly selective visual distinction of pyrophosphate from other phosphate anions with 4-[(5-chloro-2-pyridyl)azo]-1,3-diaminobenzene in the presence of copper(II) ions.

Pyrophosphate (PPi), as a biologically related phosphate anion, plays very important roles in organisms. Here, a highly selective visual method for distinction of PPi was made with commercial available 4-[(5-chloro-2-pyridyl)azo]-1,3-diaminobenzene (5-Cl-PADAB) in the presence of copper(II). The yellow solution of 5-Cl-PADAB exhibits strong absorption at 450.5 nm, and addition of Cu(II) results in a red solution with a new absorption band at 506.0 nm. Upon titration with PPi, the absorption band at 506.0 nm decreases with blueshift, while another new absorption band in the region from 562.0 nm to 750.0 nm appears which gradually splits into two peaks, and the color accordingly changes from red to cyan. Further addition of PPi, the new absorption peaks gradually disappear, and the mixture shows the absorption of 5-Cl-PADAB and recovers to yellow from cyan. This process is highly selective for PPi since other phosphate anions such as nucleotides cannot induce such spectral and color changes. With this method, the detection of PPi concentration in human urine was made with satisfactory results.

A terbium(III)-organic framework for highly selective sensing of cytidine triphosphate.

Highly selective sensing of cytidine triphosphate (CTP) against other triphosphate nucleosides including ATP, GTP and UTP is successfully achieved with a luminescent terbium(III)-organic framework (TbOF) of [Tb(2)(2,3-pzdc)(2)(ox)(H(2)O)(2)](n) (2,3-pzdc(2-) = 2,3-pyrazinedicarboxylate, ox(2-) = oxalate).

A ratiometric fluorescence recognition of guanosine triphosphate on the basis of Zn(II) complex of 1,4-bis(imidazol-1-ylmethyl) benzene.

As one vital member among the family of phosphates, guanosine triphosphate (GTP) not only plays a very important role in many critical biological processes but also closely associates with definite pathological states. Based on the ratiometric fluorescence response of the zinc complex of 1,4-bis(imidazol-1-ylmethyl) benzene (bix) in this contribution, a highly selective recognition of GTP has been successfully developed. The fluorescence of bix-Zn(II) at 289 nm decreased in the presence of GTP with the appearance of one new emission band at 341 nm, resulting in ratiometric fluorescence changes with the concentration of GTP. With that, ratiometric fluorescence recognition for GTP could be effectively established, and so GTP could be successfully discriminated from other structurally similar anions, including ATP and PPi. Furthermore, bix-Zn(II) also has a ratiometric fluorescence response to DNA sequences containing guanine.

Photophysical and electrochemical properties of platinum(II) complexes bearing a chromophore-acceptor dyad and their photocatalytic hydrogen evolution.

A series of platinum(II) complexes bearing a chromophore-acceptor dyad obtained by reacting 4-(p-bromomethylphenyl)-6-phenyl-2,2'-bipyridine or 4'-(p-bromomethylphenyl)-2,2':6',2''-terpyridine with pyridine, 4-phenylpyridine, 4,4'-bipyridine, 1-methyl-4-(pyridin-4'-yl)pyridinium hexafluorophosphate respectively, were synthesized. Their photophysical properties, emission quenching studies by Pt nanoparticles and methyl viologen, electrochemical properties and photoinduced electron-transfer reactions in a photocatalytic hydrogen-generating system containing triethanolamine and colloidal Pt without an extra electron relay, were investigated. A comparison of the rates of hydrogen production for the two photocatalytic systems, one containing a metal-organic dyad and the other comprising a 1:1 mixture of the parental platinum(II) complexes and the corresponding electron relay, showed that intramolecular electron transfer improves the photocatalytic efficiency. Compared with cyclometalated platinum(II) complexes, the related platinum(II) terpyridyl complexes exhibited poor performance for photocatalytic hydrogen evolution. An investigation into the amount of hydrogen generated by three platinum(II) complexes containing cyclometalated ligands with methyl groups located on different phenyl rings revealed that the efficiency of hydrogen evolution was affected by a subtle change of functional group on ligand, and the hydrogen-generating efficiency in the presence or absence of methyl viologen is comparable, indicating electron transfer from the excited [Pt(C^N^N)] chromophore to colloidal Pt. (1)H NMR spectroscopy of the metal-organic dyads in an aqueous solution in the presence of excess triethanolamine revealed that the dyad with a viologen unit was unstable, and a chemical reaction in the compound occurred prior to irradiation by visible light under basic conditions.

Zn(II) complex of terpyridine for the highly selective fluorescent recognition of pyrophosphate.

Pyrophosphate ion (PPi) is crucial in varieties of biological processes and industrial applications, and thus it is very important how to recognize it with high selectivity. In this contribution, one terpyridine (tpy)-based fluorescent molecule, 4-(methylphenyl)-2,2':6',2''-terpyridine (mptpy), has been reported to display a highly selective recognition for PPi in the presence of Zn(II). After exposure toward the Zn(II) ion, the characteristic emission of mptpy at 376 nm red-shifted to 406 nm with a strong enhancement upon an excitation at 280 nm, and then blue-shifted to 388 nm with the further addition of PPi. Absorption and fluorescence measurements showed that other phosphates including phosphate (Pi) as well as nucleotide triphosphates could not induce the spectral changes similar to PPi, demonstrating the unique binding effect between mptpy-Zn(II) and PPi. This process could also discriminate PPi from other inorganic anions. Therefore, a tpy-based fluorescence method for the highly selective recognition of PPi could be developed.

Selective fluorometric detection of pyrophosphate and stringent alarmone with copper(II)-2,6-bis(2-benzimidazolyl)pyridine complex.

Pyrophosphate (PPi) is involved in lots of anabolism and bioenergetic processes in organisms and possesses important biological functions so that its detection is very significant. Here, we developed a selective fluorometric detection method for PPi with a copper(II) complex of 2,6-bis(2-benzimidazolyl)pyridine (bbimp), and then applied it to the detection of bacterial alarmone ppGpp. bbimp has the fluorescence emission at 395 nm, but the bbimp-Cu(2+) complex is hardly fluorescent because the intrinsic fluorescence of bbimp is effectively quenched by Cu(2+). With the addition of PPi, however, the fluorescence emission of bbimp turns on with a 2 nm red-shift, and has a linear relationship with PPi in the range of 3-90 µmol/L. This method has good selectivity for PPi over other anions especially those phosphate-containing anions such as ATP, UTP, CTP, GTP, GDP, and PO(4)(3-).

Highly selective detection of phosphate in very complicated matrixes with an off-on fluorescent probe of europium-adjusted carbon dots.

A simple method for phosphate (Pi) detection is established by developing an off-on fluorescence probe of europium-adjusted carbon dots (CDs), which has been successfully applied to the detection of Pi in very complicated matrixes such as artificial wetlands system.

A one-pot strategy for biomimetic synthesis and self-assembly of gold nanoparticles.

A simple, one-pot and controllable strategy is reported in this contribution for biomimetic synthesis and self-assembly of gold nanoparticles (Au-NPs). It involves our synthesized polyaldehyde dextran (PAD), which has been proved to be a biomacromolecule with excellent biocompatibility and biodegradability, acting as both a reducing agent and a stabilizer. The morphology of the as-prepared Au-NP assemblies can be controlled by adjusting the reaction conditions, such as the concentration of aldehyde in PAD, the reaction time and the temperature. Investigations of the mechanism suggest that stabilizers may distribute on different crystal facets of NPs non-uniformly owing to the different binding forces, and dipole-dipole interaction of NPs could be the main driving force for the assembly of Au-NPs. In addition, intermolecular hydrogen bonding interaction of stabilizers could also act as a possible driving force. The excellent biocompatibility of the Au-NP assemblies makes them promising candidates for fabricating future optical nanodevices and application in biological systems.