基于环糊精的荧光传感器*

基于环糊精的荧光传感器研究倍受人们关注,近年来得到了迅速发展。本文阐述了环糊精络合物的形成以及它们的分子识别机理,在此基础上详细综述了丹酰、芘、萘等荧光基团修饰的环糊精单体及含多个主体的荧光传感器的制备及应用研究现状。此外,对基于环糊精超分子络合物的荧光传感器2003年以后的发展也进行了介绍,同时展望了该领域的发展趋势。     

Complexation of Ketoconazole by Native and Modified Cyclodextrins

Complexation of ketoconazole (KET), a broad-spectrum antifungal drug, with β- and γ-cyclodextrins (CDs), heptakis (2,6-di-O-methyl)-β-CD (2,6-DM-β-CD), heptakis (2,3,6-tri-O-methyl)-β-CD (TM-β-CD), 2-hydroxypropyl-β-CD (2HP-β-CD) and carboxymethyl-β-CD (CM-β-CD) was studied. The stability constants were determined by the solubility method at pH = 6 and for 2,6-DM-β-CD and CM-β-CD at pH = 5. At pH = 6, the stability constants increased in the order: TM-β-D < γ-CD < 2HP-β-CD < β-CD < CM-β-CD < 2,6-DM-β-CD. At pH = 5, due to the increased ionization of KET, the stability constant with CM-β-CD increased and with 2,6-DM-β-CD decreased. For complexes of KET with 2HP-β-CD and 2,6-DM-β-CD, the thermodynamic parameters of complexation were determined from the temperature dependence of the corresponding stability constants. For β–γ and TM-β-CD complexes, calculations using HyperChem 6 software by the Amber force field were carried out to gain some insight into the host–guest geometry.     

Interactions with Charged Cyclodextrins and Chiral Recognition

Anionic N-acetylated -aminoacids (AcTrp^-, AcPhe^-, AcLeu^- and AcVal^-) are bound to protonatedheptakis(6-amino-6-deoxy)--cyclodextrin(per-NH₃ ⁺--CD) by a cooperative work of inclusion and Coulomb interactions.Such complexation occurs enantioselectively ((S)-selective)and is accompanied bypositive entropy changes. Similar (S)-selectivecomplexation occurs in the oppositelycharged system. Namely, cationic -aminoacid methyl esters are enantioselectivelybound to dissociatedheptakis(6-carboxymethylthio-6-deoxy)--cyclodextrin(per-COO^---CD). In order to obtain thegeneral mechanism for complexationof a charged host with an oppositely charged guest,we examined the ¹H NMR spectra oncomplexation of simple carboxylate anions suchas p-methylbenzoate anion andalkanoate anions with per-NH₃ ⁺--CD.Both Coulomb interactions andinclusion are essential to form stable complexesof these carboxylate anions. In allcases, positive entropy changes promote thecomplexation between the carboxylateanions and per-NH₃ ⁺--CD. Dehydrationfrom both charged host and guestis the origin of entropic gains. The mechanism forcomplexation of a charged host withan oppositely charged guest involving the cooperativework of inclusion and Coulombinteractions and positive entropy change due todehydration upon complexation isgenerally applied for related systems such asenantioselective complexation ofRu(phen)₃ ²⁺ with per-COO^---CDand of Ru(phen)₃ ²⁺with DNA.     

Molecular Recognition of Aliphatic Alcohols and Carboxylic Acid by Chromophoric Cyclodextrins

Abstract

Molecular recognition behavior of eight cyclodextrin derivatives, i.e. mono(6-pyridinio-6-deoxy)-α-cyclodextrin (1α), mono(6-pyridinio-6-deoxy)-β-cyclodetrin (1β), mono(6-pyridinio-6-deoxy)-γ-cyclodextrin (1γ), mono[6-(p-picolinio)-6-deoxy]-β-cyclodextrin (2β), mono(6-anilino-6-deoxy)-β-cyclodextrin (3β), mono[6-(m-toluidino)-6-deoxy]-β-cyclodextrin (4β), mono[6-O-(8-quinolyl)]-β-cyclodextrin (5β), and novel mono[6-(2-naphthylamino)-6-deoxy]-β-cyclodextrin (6β), with a series of aliphatic alcohols and carboxylic acid has been investigated spectroscopically. Using the appended aromatic group as a spectral probe, spectroflurometric or spectropolarimetric titrations have been performed at 25°C in aqueous phosphate buffer solution (pH 7.20, 0.1 M) to determine the complex stability constants (K_s ) and Gibbs free energy changes (-δG°) for the stoichiometric 1:1 inclusion complexation of cyclodextrin derivatives with the guests. The results obtained demonstrate that the modified cyclodextrins are highly sensitive to the size/shape and hydrophobicity of guest molecules, and particularly 5β gives an excellent molecular selectivity up to 215 for 1-adamantanol/cyclohexanol. The binding ability and selectivity of the modified cyclodextrins (1α, 1β, and 1β-6β) are discussed from the view points of size/shape-fit concept, induced-fit interaction, and the multiple recognition mechanisms.     

Release Characteristics of Iodine Encapsulated in Cyclodextrins

The effect of cyclodextrins (CDs) on water solubility of iodine was investigated. Modified CDs greatly enhanced the solubility of iodine. On the contrary, enhancement by natural CDs was rather moderate whereby the solubility was only doubled at the highest β-CD concentration examined. Desorption experiment of iodine from solution was carried out with addition of various CDs to study the effect of CDs on iodine retention. α-CD was the most efficient in retarding iodine desorption. Later, various concentrations of α-CD were used in the desorption experiment to observe its volatile suppression effect and determine the stability constant of iodine/α-CD complexation. At α-CD concentration of 10.3 mM, no lost of iodine from the solution was detected. A model was developed for desorption of iodine from the solution based on mass transfer theory. The stability constant K given by this model was 3.28×10⁴ M⁻¹ which was in the same order as the value estimated in this study by solubility method and as well those reported by other authors. In release experiments of solid state inclusion complexes, stability of inclusion complex powders decreased in the order of α-CD>β-CD>randomly methylated β-CD (RM-β-CD).     

The Driving Forces in the Inclusion Complexation of Cyclodextrins

The driving forces leading to the inclusion complexation of cyclodextrins were reviewed, which included the electrostatic interaction, van der Waalsinteraction, hydrophobic interaction, hydrogen bonding, release of conformational strain,exclusion of cavity-bound high-energy water, and charge–transferinteraction. It was shown that except for the release of conformation strain and exclusion of cavity-bound water, the otherinteractions were indeed contributive to the complex formation. However, it was concludedthat the enthalpy and entropy changes of the complexation were not good criteria to be used injudging whether a particular driving force was present or important, mainly because of theoccurrence of enthalpy-entropy compensation. On the other hand, the multivariate quantitativestructure-activity relationship analyses usually could illustrate which driving forces wereimportant in certain inclusion complexation systems.     

Bile Acids-Selective Chemosensors Based on NBD-amine-Modified Cyclodextrins

A new type of fluorescent chemosensor, based on modified cyclodextrins bearing the fluorophore unit NBD-amine, was prepared. One of these new chemosensors, NC0γCD is very sensitive to bile acids, but is not sensitive to other guests (e.g. adamantane and borneol derivatives). The response of the new type of chemosensor to a guest was an increase in the fluorescence intensity and the sensitivity parameter (ΔI/I ₀ ) dose not correlate to the binding affinity of NC0γCD.     

A Spectroscopic and Molecular Modelling Study of the Nature of the Association Complexes of Nile Red with Cyclodextrins

The polarity-sensitive fluorescent dye Nile Red forms association complexes with various cyclodextrins in aqueous solution. The formation of such association complexes has a significant effect on the Nile Red fluorescence, with the largest effect being observed in -cyclodextrin solution. When -cyclodextrin is used to increase the Nile Red concentration in solution, the absorption spectrum shows a large blue shift indicating the formation of an inclusion complex, but surprisingly the Nile Red fluorescence is strongly suppressed. A proposed explanation for this observation involves the formation of 1:2 host:guest complexes, in which the Nile Red guests are included as relatively non-fluorescent dimers. When the solutions were prepared by adding -cyclodextrinto a near-saturated aqueous solution of Nile Red (so that 1:1 or 2:1 complexation shouldbe favoured), significant fluorescence enhancement was observed. Analysis of thefluorescence enhancement as a function of host concentration indicated the formationof 2:1 host:guest complexes in these solutions. However, electrospray mass spectroscopic studies show no evidence for the formation of any such inclusion complexes. Furthermore, molecular modelling shows that the formation of a complex involving full insertion of Nile Red in the -cyclodextrin cavity is not stable, and will quickly eject the Nile Red guest molecule. These modelling results suggest that an association complex involving capping (via the association of Nile Red parallel to the cavity opening, or by partial insertion into the cavity) of the -cyclodextrin cavity by one or two Nile Red molecules is much more likely.     

Fluorescence Signal Enhancement Using Double Pass Cell Configurations

Abstract

A simple and reproducible method for increasing the sensitivity of fluorescence measurements by a factor of 2.0–2.5 is presented. Data are presented to explain: the deviation from the expected theoretical increase in sensitivity of 4.0; wavelength dependency and subsequent problems arising from enhanced inner filtering effects.     

Complexation of Budesonide in Cyclodextrins and Particle Aerodynamic Characterization of the Complex Solid Form for Dry Powder Inhalation

The purpose of this study is to evaluate the effect ofbudesonide-cyclodextrins (CDs) complex formation in thein-vitro aerodynamic properties of the dry powder producedfor pulmonary delivery. Phase-solubility studies were performedusing budesonide and -CD, DM--CD and HP--CD.The complex budesonide:DM--CD revealed the highest stabilityconstant (K_s = 3339.7 ± 4.76%; n = 3) and the solid powder was prepared by spray-drying.Complexation was evidenced by Differential Scanning Calorimetry (DSC). A physical mixture of budesonide and DM--CD was prepared for use as reference. The fine particle fraction and particle size distribution of both powders were assessed using Twin Stage Liquid Impinger (TSLI) and Aerosizer®LD, respectively. The content uniformity of the capsules filled (sd); (n) was 191.8 (± 2.74) g; (10) for the budesonide:DM--CD solid complex and 204.9 (± 9.35) g; (10) for the physical mixture. The emitted dose (rsd); (n) was 68.0% (± 26.1%); (5) of the nominal dose (solid complex) and 70.6% (± 12.6%); (5) (physical mixture). The fine particle fraction was 67.7% (± 18.9%); (5) of the emitted dose (solid complex) and 39.8% (± 16.9%); (5) (physical mixture). While no statistically significant difference was observed between the emitted dose means of both the solid complex and physical mixture, a statistically significant higher fine particle fraction mean was obtained for the solid complex. The results suggest that using a spray-dried CD complex powder for pulmonary drug delivery may increase the drug's respirable fraction and consequently its therapeutic efficacy.     

Electroanalytical Method of Acid Blue 120 and its Supramolecular System with Cyclodextrins

In this paper, a simple, rapid, sensitive and accurate electroanalytical method of Acid Blue 120 (AB120) has been established by polarography. In a supporting electrolyte of 0.01 mol l⁻¹ Na₂HPO₄–KH₂PO₄ (pH 7.04) solution, a sensitive first derivative reduction peak (ip′) of AB120 was found by Linear Sweep Voltammetry (LSV). The peak potential is −820 mV (versus SCE). The peak current (ip′) is proportional to the concentration over the range 2.0 × 10⁻⁷–5.0 × 10⁻⁵ mol l⁻¹ (r=0.9961–0.9991) and the limit of detection (LOD) is 1.0 × 10⁻⁷ mol l⁻¹. The recovery of AB120 varied from 95.3 to 103.0% and the relative standard deviation (RSD) was 2.2% (n=8). The method has been expected to determination of wastewater in dye industry. In addition, the supramolecular system of AB120 with cyclodextrins has been studied. It can form 1:1 inclusion complex with six CDs. The inclusion constants were calculated and the inclusion ability of different kinds of CDs was compared. Furthermore, the inclusion mechanism was also preliminarily discussed, which provided some valuable information for further application of AB120 and CDs.     

A Theoretical Study on the Inclusion Complexation of Cyclodextrins with Inorganic Cations and Anions

PM3 and B3LYP/3-21+g(d) calculations were performed on theinclusion complexation of- and -cyclodextrin with inorganic cationsand anions includingLi⁺, Na⁺, F^-, and Cl^-. Both the gas-phase interaction and solvent effect weretaken into consideration. The CD complex with an anionwas more stable than that with acation, which was in agreement with the experimentalfindings. It was proposed thathydrogen bonding between the anion and the cyclodextrincavity was the physical origin ofsuch behavior.     

多胺修饰β-环糊精对铝-铁试剂荧光体系的增敏作用

研究了β－环糊精（1）、单－[6-乙二胺－6－脱氧]-β－环糊精（2）、单-[6-(二乙烯三胺）-6-脱氧]-β-环糊精（3）和单-[6-(三乙烯四胺）－6－脱氧]-β-环糊精（4）对铝－铁试剂荧光体系的增敏作用。实验结果表明：加入β－环糊精及其衍生物后体系的荧光强度均增强，增敏率为4＞3＞2＞1，其中4给出最大的荧光增敏率。研究了pH值在3.0-7.5之间酸度对主体化合物增敏能力的影响，发现体系的pH值对β－环糊精的增敏率影响较小，而随着体系酸性的增强，多胺修饰β－环糊精的增敏作用显著增强。探讨了修饰环糊精引入边臂的长度和主－客体间的尺寸适合等因素对铝－铁试剂体系荧光增敏率的影响。     

中氮茚腈类化合物的合成及其荧光性质的研究

合成了一系列中氮茚化合物,研究了化合物3a～3k,4a～4b,5～6分子中不同取代基对其荧光光谱和荧光量子效率的影响.发现中氮茚环上的氰基具有较好的荧光助色作用,中氮茚环上的COPh,COMe或CHO对分子的荧光具有猝灭作用;发现中氮茚类有机化合物的分子结构与荧光性质之间的某些关系.化合物3a～3b,3g～3i具有较高的荧光量子效率,可以作为比较好的有机荧光分子用于探针化合物的结构修饰.     

加压毛细管电色谱-激光诱导荧光法检测4种黄曲霉毒素

建立了三氟乙酸( TFA)柱前衍生,加压毛细管电色谱-激光诱导荧光( pCEC-LIF)快速测定黄曲霉毒素B1、B2、G1、G2方法。使用粒径1.8μm的C18毛细管色谱柱,以甲醇-水(45：55, V/V,含0.05%甲酸)为流动相,泵流速为0.05 mL/min,分离电压为15 kV,激发波长为375 nm,发射波长为450 nm,黄曲霉毒素B1, B2, G1, G2达到基线分离。各组分的检出限(S/N=3)分别为0.02,0.016,0.008和0.01μg/L,在0.1~10μg/L,0.1~10μg/L,0.1~3.0μg/L,0.1~3.0μg/L 范围内分别呈线性相关,相关系数 R2分别为0.9999,1.0000,0.9995,0.9997。将本方法应用于花生酱的分析,加标回收率在90.0%~112.0%之间,RSD在0.5%~1.9%之间。     

The Interactions Between Nonionic Surfactants and Cyclodextrins Studied by Fluorescence Measurements

The complex formation between some nonionic surfactants and -, - and -cyclodextrin was studied by fluorescence measurements. The relative fluorescence intensity of a solute containing a nonionic surfactant at a constant concentration far below the critical micelle concentration (CMC) are enhanced by the addition of cyclodextrins. Non linear type equations were derived to obtain stability constants by fluorescence measurements for inclusion complexes formed between cyclodextrins and the nonionic surfactants. In most cases 1 : 1- and 2 : 1-complexes (ratio of cyclodextrin to surfactant) are formed.     

Fluorescence Enhancement of Levosulpiride Upon Coordination with Transition Metal Ions and Spectrophotometric Determination of Complex Formation

Abstract

Absorbance and fluorescence spectral pattern of levosulpiride in absence and presence of first row transition metal ions (Mn-Zn) has been studied at room temperature under physiological condition. The fluorescence spectra of the drug in presence of different concentrations of transition metal ions showed enhancement in fluorescence intensity of levosulpiride. The photophysical changes owing to the direct interaction between metal ion and the amide nitrogen of levosulpiride has been described in terms of CHEF (chelating enhancement fluorescence) effect. The absorption spectra of the drug at different pH exhibited two isosbestic points at 255 and 275 nm respectively, indicating the presence of three chemical species in solution. The ratio of the drug to metal ions is found to be 2:1 and the log K of the resulting complex was determined spectrophotometrically and potentiometrically. The apparent ionization constant of levosulpiride is found to be 8.98. The low value of stability constant suggests that complexes may dissolve and the drug can be absorbed.     

Determining Metal Ion Complexation Kinetics with Fluorescent Ligands by Using Fluorescence Correlation Spectroscopy

Fluorescence correlation spectroscopy (FCS) has been extensively used to measure equilibrium binding constants (K) or association and dissociation rates in many reversible chemical reactions across chemistry and biology. For the majority of investigated reactions, the binding constant was on the order of ∼100 M⁻¹, with dissociation constants faster or equal to 10³ s⁻¹, which ensured that enough association/dissociation events occur during the typical diffusion-determined transition time of molecules through the FCS detection volume. However, complexation reactions involving metal ions and chelating ligands exhibit equilibrium constants exceeding 10⁴ M⁻¹. In the present paper, we explore the applicability of FCS for measuring reaction rates of such complexation reactions, and apply it to binding of iron, europium and uranyl ions to a fluorescent chelating ligand, calcein. For this purpose, we exploit the fact that the ligand fluorescence becomes strongly quenched after binding a metal ion, which results in strong intensity fluctuations that lead to a partial correlation decay in FCS. We also present measurements for the strongly radioactive ions of ²⁴¹Am³⁺, where the extreme sensitivity of FCS allows us to work with sample concentrations and volumes that exhibit close to negligible radioactivity levels. A general discussion of the applicability of FCS to the investigation of metal-ligand binding reactions concludes our paper.     

Stabilization of All-trans-retinol by Cyclodextrins: A Comparative Study Using HPLC and Fluorescence Spectroscopy

Purpose: To formulate preparations incorporating cyclodextrins (CDs) which could be used for direct delivery to the retina of vitamin A (all-trans-retinol), while also protecting it from degradation in the aqueous environment. Vitamin A supplementation is being considered for treatment of several ophthalmic diseases characterised by progressive photoreceptor degeneration. Methods: The complexation between vitamin A and ten cyclodextrins, Captisol^®(sulfobutyl ether-7--cyclodextrin), hydroxypropyl--CD, 2-hydroxypropyl--CD, -CD, hydroxypropyl--CD, hydroxypropyl--CD, -CD, methyl--CD,Heptakis-(2,6-di-O-methyl)--CD andHeptakis-(2,3,6-tri-O-methyl)--CD,was investigated using bothhigh sensitivity fluorescence spectrometry and HPLC (high pressure liquid chromatography). Samples of retinol-CD complexes in phosphate buffer, pH 7.4 were analysed for up to 72 hours. Optimum conditions for formation of the Captisol-retinol complexes were investigated. Results: Using spectroscopic measurements and HPLC, the complexes formed between ten cyclodextrins and all-trans-retinol were evaluated. The results indicate that all cyclodextrins tested were able to form inclusion complexes as shown by the fluorescence signals which are considerably larger than those obtained in the absence of cyclodextrin. Only minimal degradation of retinol over 48 hours was observed with three of these cyclodextrins. Captisol was able to stabilise all-trans-retinol for up to 72 hours, as shown by HPLC, and the optimum ratio of Captisol to retinol was determined to be 50 to 1. Addition of glutathione and decrease in pH did not improve stability of the complex. Conclusions: This survey suggests that Captisol and other cyclodextrins could be used to stabilise and solubilise vitamin A in aqueous media and this establishes the basis for an ocular Captisol-retinol drug delivery system now under development in our laboratory.     

Instrumental HPTLC of Aflatoxins: Fluorescence Enhancement by Corn Free Fatty Acids

Abstract

Greater than 100% recoveries using instrumental HPTLC were observed for aflatoxin (AFT) analysis in spiked corn samples. I₂ detection of spots over-laying AFT B₁ and B₂ were identified by GLC as C₁₆ ^?c18 free fatty acids (FFA). These FFA were found to enchance the fluorescence of AFT B₁ from 13.7% to 35.7% greater than controls resulting in >100% recoveries. The inclusion of glacial acetic acid in the TLC mobile phase resulted in an increased mobility of the FFA which eliminated the positive interference on AFT fluorescence. Recoveries using the modified developing solvent then gave values in acceptable ranges.