Simple pulse-acquire NMR methods for the quantitative analysis of calcium, magnesium and sodium in human serum

Simple pulse-acquire NMR methods are presented for accurate quantitation of calcium (Ca(2+)), magnesium (Mg(2+)) and sodium (Na(+)) in human serum. Ca(2+) and Mg(2+) can be determined simultaneously by (1)H NMR via their EDTA (ethylenediaminetetraacetic acid) complexes. Spectra are acquired before and after addition of EDTA, and the difference spectrum is used for integration of the signals from the complexes relative to an internal reference. Serum sodium can also be determined through pulse-acquire (23)Na NMR by integration of the free sodium signal relative to a reference signal (Na(+)+EDTA in a coaxial capillary tube). The method shows excellent accuracy and precision for all three metal ions. Slow chemical exchange between complexed and free EDTA at the natural pH of serum does not limit the accuracy of the determination of Ca(2+) and Mg(2+), and the large errors associated with spin-echo spectral editing methods are avoided.     

Sequence diversity, metal specificity, and catalytic proficiency of metal-dependent phosphorylating DNA enzymes

Although DNA has not been found responsible for biological catalysis, many artificial DNA enzymes have been created by "in vitro selection." Here we describe a new selection approach to assess the influence of four common divalent metal ions (Ca(2+), Cu(2+), Mg(2+), and Mn(2+)) on sequence diversity, metal specificity, and catalytic proficiency of self-phosphorylating deoxyribozymes. Numerous autocatalytic DNA sequences were isolated, a majority of which were selected using Cu(2+) or Mn(2+) as the divalent metal cofactor. We found that Cu(2+)- and Mn(2+)-derived deoxyribozymes were strictly metal specific, while those selected by Ca(2+) and Mg(2+) were less specific. Further optimization by in vitro evolution resulted in a Mn(2+)-dependent deoxyribozyme with a k(cat) of 2.8 min(-1). Our findings suggest that DNA has sufficient structural diversity to facilitate efficient catalysis using a broad scope of metal cofactor utilizing mechanisms.     

A metallic cobalt electrode for the indirect potentiometric determination of calcium and magnesium in natural waters using flow injection analysis

A flow injection analysis of Ca(2+) and Mg(2+) using indirect potentiometric detection in natural waters is proposed, where Ca(2+) or Mg(2+) are injected into a buffer carrier containing phosphate, resulting in the formation of Ca(3)(PO(4))(2) or Mg(3)(PO(4))(2). The consequent reduction in free phosphate in the carrier solution is detected using a metallic cobalt wire electrode. Indirect electrode response was used and the experimental conditions affecting electrode response were optimized. Responses were linear in the concentration range 5x10(-4) to 5x10(-3) M with a detection limit of 1x10(-5) M in 20 mM phosphate buffer at pH 8.0. The relative standard derivation at 1 mM of Ca(2+) and Mg(2+) were 3.9 and 3.7% (n=10), respectively. EGTA and 8-hydroxyquinoline were used as the masking agents for Ca(2+) and Mg(2+), respectively. Concentrations of Ca(2+) and Mg(2+) in natural waters were successfully determined by the proposed method.     

PVC ion-selective electrodes based on calcium bis-[dialkyl- and di-(4-alkylphenyl) phosphates] and mixed solvent mediators

PVC calcium ion-selective electrodes based on either calcium bis-di(n-decyl)phosphate or calcium bis-di[4-(1,1,3,3-tetramethylbutyl)phenyl]phosphate with various solvent mediators (alone or in pairs) have been evaluated with particular respect to interference from Na(+), K(+), Mg(2+), Sr(2+), Ba(2+), Mn(2+), Cu(2+), Ni(2+) and Zn(2+) ions. PVC calcium ion-selective electrodes based on calcium bis-di[4-(n-octyl)phenyl]phosphate with varied amounts of decan-l-ol plus di-n-octyl phenylphosphonate showed a continuous gradation in selectivity coefficients on going from a high fraction of decan-l-ol to a high fraction of the second mediator. Thus, k(pot)(Ca,Mg) changed from 1.6 for an electrode based exclusively on decan-l-ol to 4.9 x 10(-4) for one based completely on di-n-octyl phenylphosphonate. The corresponding k(pot)(Ca,Na) values were 7.0 x 10(-2) and 1.1 x 10(-3).     

The characterization for the binding of calcium and terbium to Euplotes octocarinatus centrin

Centrin is a member of the EF-hand superfamily that plays critical role in the centrosome duplication and separation. In the present paper, we characterized properties of metal ions binding to Euplotes octocarinatus centrin (EoCen) by fluorescence spectra and circular dichroism (CD) spectra. Changes of fluorescence spectra and alpha-helix contents of EoCen proved that Tb(3+) and Ca(2+) induced great conformational changes of EoCen resulting in exposing hydrophobic surfaces. At pH 7.4, Ca(2+) (and Tb(3+)) bond with EoCen at the ratio of 4:1. Equilibrium experiment indicated that Ca(2+) and Tb(3+) exhibited different binding capabilities for C- and N-terminal domains of protein. C-terminal domain bond with Ca(2+) or Tb(3+) approximately 100-fold more strongly than N-terminal. Aromatic residue-sensitized Tb(3+) energy transfer suggested that site IV bond to Tb(3+) or Ca(2+) more strongly than site III. Based on fluorescence titration curves, we reckoned the conditional binding constants of EoCen site IV quantitatively to be K(IV)=(1.23+/-0.51)x10(8)M(-1) and K(IV)=(6.82+/-0.33)x10(5)M(-1) with Tb(3+) and Ca(2+), respectively. Metal ions bond to EoCen in the order of IV>III>II, I.     

Synthesis, photochromic properties, and light-controlled metal complexation of a naphthopyran derivative

A light-controlled reversible binding switch based on photochromic 3H-naphtho[2,1-b]pyran is under development for studying cellular oscillatory calcium signals. The binding affinities of the closed and open forms of substituted naphthopyran 1 for Ca(2+), Mg(2+), and Sr(2+) in buffer were determined. The photochemically ring-opened form of the receptor exhibited increased affinity compared to the thermally stable closed form of the receptor. The binding affinity difference for Ca(2+) was approximately 77-fold at pH 7.6.     

Highly sensitive, selective, and rapid fluorescence Hg2+ sensor based on DNA duplexes of poly(dT) and graphene oxide

Coupling T base with Hg(2+) to form stable T-Hg(2+)-T complexes represents a new direction in detection of Hg(2+). Here a graphene oxide (GO)-based fluorescence Hg(2+) analysis using DNA duplexes of poly(dT) that allows rapid, sensitive, and selective detection is first reported. The Hg(2+)-induced T(15)-(Hg(2+))(n)-T(15) duplexes make T(15) unable to hybridize with its complementary A(15) labelled with 6'-carboxyfluorescein (FAM-A(15)), which has low fluorescence in the presence of GO. On the contrary, when T(15) hybridizes with FAM-A(15) to form double-stranded DNA because of the absence of Hg(2+), the fluorescence largely remains in the presence of GO. A linear range from 10 nM to 2.0 μM (R(2) = 0.9963) and a detection limit of 0.5 nM for Hg(2+) were obtained under optimal experimental conditions. Other metal ions, such as Al(3+), Ag(+), Ca(2+), Ba(2+), Mg(2+), Zn(2+), Mn(2+), Co(2+), Pb(2+), Ni(2+), Cu(2+), Cd(2+), Cr(3+), Fe(2+), and Fe(3+), had no significant effect on Hg(2+) detection. Moreover, the sensing system was used for the determination of Hg(2+) in river water samples with satisfactory results.     

A mass spectrometric study of metal binding to osteocalcin

Electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry was used to investigate Ca(2+), Mg(2+), and La(3+) binding to bovine bone osteocalcin (OCN). OCN was shown to bind 3 mol Ca(2+) per mol protein. There was also evidence for the presence of four additional metal binding sites. Ca(2+) increased the formation of the OCN dimer. Mg(2+) bound to OCN to the same extent as Ca(2+) but did not induce the dimerization of OCN. La(3+) bound to a lesser extent than either Ca(2+) or Mg(2+) to OCN and, like Mg(2+), did not influence dimerization. Each Gla residue of OCN participates in Ca(2+) binding, whereas Mg(2+) binding may occur preferentially at sites other than Gla residues. This implies that the different natures of Ca(2+)- and Mg(2+)-containing OCN complexes influence the tendency of OCN to form a dimer.     

A highly selective turn-on colorimetric, red fluorescent sensor for detecting mobile zinc in living cells

We describe ZRL1, a turn-on colorimetric and red fluorescent zinc ion sensor. The Zn(2+)-promoted ring opening of the rhodamine spirolactam ring in ZRL1 evokes a 220-fold fluorescence turn-on response. In aqueous media, ZRL1 turn-on luminescence is highly selective for Zn(2+) ions, with no significant response to other competitive cations, including Na(+), K(+), Ca(2+), Mg(2+), Mn(2+), Fe(2+), Co(2+), Ni(2+), Cu(2+), Cd(2+), or Hg(2+). In addition to these characteristics, preliminary results indicate that ZRL1 can be delivered to living cells and can be used to monitor changes in intracellular Zn(2+) levels.     

Structure-related variable responses of calcium sensitive MRI probes

A new series of Gd(3+) complexes based on DO3A (GdL(1)-GdL(4)) was synthesized and investigated. They possess side chains with different structures which determine their varying binding properties and response towards endogenous metal ions, measured by changes in the longitudinal relaxivity (r(1)). GdL(4) exhibits the highest selectivity toward Ca(2+) in comparison to the other complexes, with up to a 63% increase of the r(1). GdL(2) and GdL(3) also respond to different Ca(2+) concentration ranges, however with a lower selectivity since the r(1) changes are also observed in the presence of other cations such as Mg(2+), Zn(2+) or Cu(2+). Assessment of the hydration number (q) via luminescence lifetime measurements confirmed that the change in q is responsible for the r(1) response for all the complexes.     

Determination of lead in phosphate ore and phosphogypsum

Laser induced breakdown spectroscopy (LIBS) is applied to analyze aqueous solutions of Li(+), Na(+), Ca(2+), Ba(2+), Pb(2+), Cd(2+), Hg(2+) and Er(3+) and suspensions of ErBa(2)Cu(3)O(x) particles (d=0.2 microm). An excimer (308 nm) pumped dye laser with laser pulse at 500 nm and pulse energy at 22+/-2 mJ is used to produce plasma in aqueous solution. Plasma emission lines of the elements are detected by a photodiode array detector. Detection limits of the metal ions are 500 mg/l for Cd(2+), 12.5 mg/l for Pb(2+), 6.8 mg/l for Ba(2+), 0.13 mg/l for Ca(2+), 13 microg/l for Li(+) and 7.5 microg/l for Na(+). No mercury and erbium emission can be detected, even at Hg(2+) and Er(3+) concentrations of up to the g/l range. On the other side, for Er in suspensions of ErBa(2)Cu(3)O(x) particles a more than 10(3) times higher sensitivity is found than for dissolved Er(3+). This result gives a possibility to analyze colloid-borne metal ions with an increased sensitivity.     

Mn2+在铝酸盐中的发光

用金属硝酸盐、稀土氧化物和乙酸锰为原料,用燃烧法合成了Ce³⁺、Tb³⁺、Mn²⁺共激活的铝酸盐绿色荧光粉,在Ce³⁺和Tb³⁺共激活的铝酸盐体系中掺入Mn²⁺后,发射峰中出现锰的特征峰.通过对其结构的分析,对Mn²⁺发光和最佳掺杂量给出了合理的解释.同时研究了不同碱金属和碱土金属离子代替Mg²⁺时,对Mn²⁺发光的影响.     

Dual-mode unsymmetrical squaraine-based sensor for selective detection of Hg2+ in aqueous media

A novel chemosensor based on unsymmetrical squaraine dye (USQ-1) for the selective detection of Hg(2+) in aqueous media is described. USQ-1 in combination with metal ions shows dual chromogenic and "turn-on" fluorogenic response selectivity toward Hg(2+) as compared to Li(+), Na(+), K(+), Mg(2+), Ca(2+), Ba(2+), Al(3+), Cu(2+), Cd(2+), Mn(2+), Fe(3+), Ag(+), Pb(2+), Zn(2+), Ni(2+) and Co(2+) due to the Hg(2+)-induced deaggregation of the dye molecule. A recognition mechanism based on the binding mode is proposed based on the absorption and fluorescence changes, (1)H NMR titration experiments, ESI-MS study, and theoretical calculations.     

Phase transition from hexagonal LnF3 (Ln = La, Ce, Pr) to cubic Ln0.8M0.2F2.8 (M = Ca, Sr, Ba) nanocrystals with enhanced upconversion induced by alkaline-earth doping

Complete phase transition from hexagonal LnF(3) (Ln(3+) = La(3+), Ce(3+), Pr(3+)) to monodisperse ultrasmall (～7 nm) cubic Ln(0.8)M(0.2)F(2.8) (M(2+) = Ca(2+), Sr(2+), Ba(2+)) disordered solid solution nanocubes was successfully achieved through alkaline-earth doping, which induced great intensification of the near-infrared to visible upconversion emissions of the optically active rare earth ions.     

Complexes of bivalent metal cations in electrospray mass spectra of common organic compounds

In the standard electrospray ionization mass spectra of many common, low molecular mass organic compounds dissolved in methanol, peaks corresponding to ions with formula [3M + Met](2+) (M = organic molecule, Met = bivalent metal cation) are observed, sometimes with significant abundances. The most common are ions containing Mg(2+), Ca(2+) and Fe(2+). Their presence can be easily rationalized on the basis of typical organic reaction work-up procedures. The formation of [3M + Met](2+) ions has been studied using N-FMOC-proline methyl ester as a model organic ligand and Mg(2+), Ca(2+), Sr(2+), Ba(2+), Fe(2+), Ni(2+), Mn(2+), Co(2+) and Zn(2+) chlorides or acetates as the sources of bivalent cation. It was found that all ions studied form [3M + Met](2+) complexes with N-FMOC-proline methyl ester, some of them at very low concentrations. Transition metal cations generally show higher complexation activity in comparison with alkaline earth metal cations. They are also more specific in the formation of [3M + Met](2+) complexes. In the case of alkaline earth metal cations [2M + Met](2+) and [4M + Met](2+) complex ions are also observed. It has been found that [3M + Met](2+) complex ions undergo specific fragmentation at relatively low energy, yielding fluorenylmethyl cation as a major product. [M + Na](+) ions are much more stable and their fragmentation is not as specific.     

Transient, sparsely populated compact states of apo and calcium-loaded calmodulin probed by paramagnetic relaxation enhancement: interplay of conformational selection and induced fit

Calmodulin (CaM) is the universal calcium sensor in eukaryotes, regulating the function of numerous proteins. Crystallography and NMR show that free CaM-4Ca(2+) exists in an extended conformation with significant interdomain separation, but clamps down upon target peptides to form a highly compact structure. NMR has revealed substantial interdomain motions in CaM-4Ca(2+), enabled by a flexible linker. In one instance, CaM-4Ca(2+) has been crystallized in a compact configuration; however, no direct evidence for transient interdomain contacts has been observed in solution, and little is known about how large-scale interdomain motions contribute to biological function. Here, we use paramagnetic relaxation enhancement (PRE) to characterize transient compact states of free CaM that are too sparsely populated to observe by traditional NMR methods. We show that unbound CaM samples a range of compact structures, populated at 5-10%, and that Ca(2+) dramatically alters the distribution of these configurations in favor of states resembling the peptide-bound structure. In the absence of Ca(2+), the target peptide binds only to the C-terminal domain, and the distribution of compact states is similar with and without peptide. These data suggest an alternative pathway of CaM action in which CaM remains associated with its kinase targets even in the resting state. Only CaM-4Ca(2+), however, shows an innate propensity to form the physiologically active compact structures, suggesting that Ca(2+) activates CaM not only through local structural changes within each domain but also through more global remodeling of interdomain interactions. Thus, these findings illustrate the subtle interplay between conformational selection and induced fit.     

Lower rim 1,3-di{4-antipyrine}amide conjugate of calix[4]arene: synthesis, characterization, and selective recognition of Hg2+ and its sensitivity toward pyrimidine bases

The structurally characterized lower rim 1,3-di{4-antipyrine}amide conjugate of calix[4]arene (L) exhibits high selectivity toward Hg(2+) among other biologically important metal ions, viz., Na(+), K(+), Ca(2+), Mg(2+), Mn(2+), Fe(2+), Co(2+), Ni(2+), Cu(2+), Zn(2+), Cd(2+), Hg(2+), Pb(2+), and Ag(+) as studied by fluorescence, absorption, and ESI MS. L acts as a sensor for Hg(2+) by switch-off fluorescence and exhibits a lowest detectable concentration of 1.87 ± 0.1 ppm. The complex formed between L and Hg(2+) is found to be 1:1 on the basis of absorption and fluorescence titrations and was confirmed by ESI MS. The coordination features of the mercury complex of L were derived on the basis of DFT computations and found that the Hg(2+) is bound through an N(2)O(2) extending from both the arms to result in a distorted octahedral geometry with two vacant sites. The nanostructural features such as shape and size obtained using AFM and TEM distinguishes L from its Hg(2+) complex and were different from those of the simple mercuric perchlorate. L is also suited to sense pyrimidine bases by fluorescence quenching with a minimum detection limit of 1.15 ± 0.1 ppm in the case of cytosine. The nature of interaction of pyrimidine bases with L has been further studied by DFT computational calculations and found to have interactions through a hydrogen bonding and NH-π interaction between the host and the guest.     

Development of a far-red to near-infrared fluorescence probe for calcium ion and its application to multicolor neuronal imaging

To improve optical imaging of Ca(2+) and to make available a distinct color window for multicolor imaging, we designed and synthesized CaSiR-1, a far-red to near-infrared fluorescence probe for Ca(2+), using Si-rhodamine (SiR) as the fluorophore and the well-known Ca(2+) chelator BAPTA. This wavelength region is advantageous, affording higher tissue penetration, lower background autofluorescence, and lower phototoxicity in comparison with the UV to visible range. CaSiR-1 has a high fluorescence off/on ratio of over 1000. We demonstrate its usefulness for multicolor fluorescence imaging of action potentials (visualized as increases in intracellular Ca(2+)) in brain slices loaded with sulforhodamine 101 (red color; specific for astrocytes) that were prepared from transgenic mice in which some neurons expressed green fluorescent protein.     

Toward optimized high-relaxivity MRI agents: thermodynamic selectivity of hydroxypyridonate/catecholate ligands

The thermodynamic selectivity for Gd(3+) relative to Ca(2+), Zn(2+), and Fe(3+) of two ligands of potential interest as magnetic resonance imaging (MRI) contrast agents has been determined by NMR spectroscopy and potentiometric and spectrophotometric titration. The two hexadentate ligands TREN-6-Me-3,2-HOPO (H(3)L2) and TREN-bisHOPO-TAM-EA (H(4)L3) incorporate 2,3-dihydroxypyridonate and 2,3-dihydroxyterephthalamide moieties. They were chosen to span a range of basicity while maintaining a structural motif similar to that of the parent ligand, TREN-1-Me-3,2-HOPO (H(3)L1), in order to investigate the effect of the ligand basicity on its selectivity. The 1:1 stability constants (beta(110)) at 25 degrees C and 0.1 M KCl are as follows. L2: Gd(3+), 20.3; Ca(2+), 7.4; Zn(2+), 11.9; Fe(3+), 27.9. L3: Gd(3+), 24.3; Ca(2+), 5.2; Zn(2+), 14.6; Fe(3+), 35.1. At physiological pH, the selectivity of the ligand for Gd(3+) over Ca(2+) increases with the basicity of the ligand and decreases for Gd(3+) over Fe(3+). These trends are consistent with the relative acidities of the various metal ions;- more basic ligands favor harder metals with a higher charge-to-radius ratio. The stabilities of the Zn(2+) complexes do not correlate with basicity and are thought to be more influenced by geometric factors. The selectivities of these ligands are superior to those of the octadentate poly(aminocarboxylate) ligands that are currently used as MRI contrast agents in diagnostic medicine.     

微波消解样品-离子色谱法测定卷烟纸中钠、钾、镁、钙的含量

提出了离子色谱法同时测定卷烟纸中钠、钾、镁和钙含量的方法。卷烟纸试样经硝酸-过氧化氢-氢氟酸微波消解,以IonPac CS16阳离子交换柱为固定相,用0.027 mol.L-1甲烷磺酸溶液作流动相。钠、钾、镁和钙4种元素在30 min内可完全分离;各离子的检出限(3S/N)分别为13,15,8.1,97 mg.L-1。方法的加标回收率在100.9%～108.8%之间,测定值的相对标准偏差(n=5)在0.87%～3.4%之间。