Investigate Complex Formation Research Articles

APP family member dimeric complexes are formed predominantly in synaptic compartments

BackgroundThe amyloid precursor protein (APP), a key player in Alzheimer’s disease (AD), is part of a larger gene family, including the APP like proteins APLP1 and APLP2. They share similar structures, form homo- and heterotypic dimers and exhibit overlapping functions.ResultsWe investigated complex formation of the APP family members via two inducible dimerization systems, the FKBP-rapamycin based dimerization as well as cysteine induced dimerization, combined with co-immunoprecipitations and Blue Native (BN) gel analyses. Within the APP family, APLP1 shows the highest degree of dimerization and high molecular weight (HMW) complex formation. Interestingly, only about 20% of APP is dimerized in cultured cells whereas up to 50% of APP is dimerized in mouse brains, independent of age and splice forms. Furthermore, we could show that dimerized APP originates mostly from neurons and is enriched in synaptosomes. Finally, BN gel analysis of human cortex samples shows a significant decrease of APP dimers in AD patients compared to controls.ConclusionsTogether, we suggest that loss of full-length APP dimers might correlate with loss of synapses in the process of AD.

Heptakis(2,6-di-O-methyl)-β-CD as a host of olanzapine: Experimental and computational study

Olanzapine (OLN) is an atypical antipsychotic drug used for the treatment of schizophrenia and bipolar depression which suffers from low aqueous solubility. We employ phase solubility diagrams (PSDs), pH-dependent UV absorption profiles, FT-IR, 1HNMR and molecular dynamics (MD) simulations to investigate complex formation of OLN with Heptakis(2,6-di-O-methyl)-β-cyclodextrin (DM-β-CD) and β-CD and its effect on OLN solubility. pKa1 and pKa2 of OLN were found to be 4.79 and 7.72, respectively, both of which underwent a shift by ∼0.4–0.5 upon complexation with β-CD, while only pKa1 exhibited a shift by -0.5 in complex with DM-β-CD. FT-IR and 1HNMR spectra demonstrated the formation of the OLN/DM-β-CD inclusion complex, with PSDs indicating 1:1 stoichiometry and a complex formation constant for DM-β-CD greater than for β-CD. MD simulations and the computed free energies showed that the neutral OLN complexes are more stable than the protonated complexes and the preference of both hosts to include the benzodiazepine moiety. Furthermore, the more favorable ∆G for DM-β-CD is in accord with experiment.

Preparation and Characterization of Inclusion Complexes of β-Cyclodextrin and Phenolics from Wheat Bran by Combination of Experimental and Computational Techniques.

Bitterness often associated with whole wheat products may be related to phenolics in the bran. Cyclodextrins (CDs) are known to form inclusion complexes. The objective was to form inclusion complexes between β-CD and wheat phenolics. Pure phenolic acids (trans-ferulic acid (FA), caffeic acid (CA), and p-coumaric acid (CO)) and phenolic acids from wheat bran were used to investigate complex formation potential. Complexes were characterized by spectroscopy techniques, and a computational and molecular modeling study was carried out. The relative amount of complex formation between β-CD and wheat bran extract was CA > CO > FA. The phenolic compounds formed inclusion complexes with β-CDs by non-covalent bonds. The quantum-mechanical calculations supported the experimental results. The most stable complex was CO/β-CD complex. The ΔH value for CO/β-CD complex was −11.72 kcal/mol and was about 3 kcal/mol more stable than the other complexes. The QSPR model showed good correlation between binding energy and 1H NMR shift for the H5 signal. This research shows that phenolics and β-CD inclusion complexes could be utilized to improve the perception of whole meal food products since inclusion complexes have the potential to mask the bitter flavor and enhance the stability of the phenolics in wheat bran.

Investigating Complex Formation of C99 with Gamma-Secretase, Using Atomistic MM-MD Simulations and Free Energy Calculations

Investigating Complex Formation of C99 with Gamma-Secretase, Using Atomistic MM-MD Simulations and Free Energy Calculations

Using bioluminescence resonance energy transfer (BRET) and super-resolution microscopy to investigate complex formation between VEGFR2, its co-receptor Neuropilin-1 and G protein coupled receptors

Using bioluminescence resonance energy transfer (BRET) and super-resolution microscopy to investigate complex formation between VEGFR2, its co-receptor Neuropilin-1 and G protein coupled receptors

Supramolecular self-assembly of water-soluble cavitands: investigated by molecular dynamics simulation

In this study, we have examined supramolecular self-assembly process of a hydrophobic guest with a water-soluble host known by the trivial name octa acid (OA). Two octa acids form a capsular assembly only in presence of a nonpolar guest(s). Size and shape of the guest control the stoichiometry of the capsular complex. Here, all atom molecular dynamics simulation has been utilized to investigate complex formation mechanisms of a nonpolar guest (nonylbenzene) with two OA cavitands. Nonylbenzene was encapsulated into the nonpolar cavity of OA capsule owing to solvophobic interactions. Upon encapsulation it was twisted and bent due to lack of free space within the capsule. These unusual forms obtained from the simulation study were in accord with experimental findings. The post-complexation attributes of the guest were regulated by the available free space within the OA and favorable non-covalent interactions between the guest and the walls of the OA capsule. In the identical simulation condition two OA cavitands did not form a capsule without a guest, thus indicating requirement of a guest during the self-assembly of OA cavitands.

Mixed-ligand complexes of copper (II) with ditiolfenols and heterocyclic diamines

By the methods spectrophotometric methods was investigated complex formation of copper with 2,6-ditiolphenol and its derivatives (2,6-ditiol-4-methylphenol, 2,6-ditiol-4-ethylphenol and 2,6-ditiol-4-tert-buthylphenol) in the presence of heterocyclic diamines. As heterocyclic diamines phenantroline (Fen), batophenantroline and dipyridile were used. It was established, that mixedligand complex were formed in weak acidic medium (pHop=6,0-8,5). Maximum of light absorption observed at (=522–550 nm. Molar coefficient of light absorption was ( = (3.08–4.92)?104. The stoichiometry of complexes had been discovered using shift the equilibrium and relative yield methods. During the extraction of copper(II) complexes with ditiol-phenols with the presence of heterocyclic diamines there was discovered that there is no chlorophorm molecules in the composition of the extracted complexes. IR spectra shows that hydroxyl group is involved to the formation of a bond with the metal and that phenantroline is included in the composition of the copper complex. In the organic phase mixed-ligand complexes have the monomeric form and can’t polymerize; that fact can be proved by calculations. By dint of spectrophotometric, IR-spectroscopic, thermogravimetric experiments and some published data we were able to determine the structural formula of complexes. Also had been determined main spectrophotometric characteristics of copper(II) mixed-ligand complexes: pH of the beginning of the deposition, optimal pH, detection limit and sensivity. For separation and finding copper, we studied the effect of interfering foreign ions: alkali, alkaline earth and rare earth elements, as well as acid anions selected masking reagents. On the base of the results was offered photometric method for determination of copper in steel different trade-marks and in food. This method characterized by good reproduction (relatively standard deviation no more 0.05) and low limit of discovery.

Column chromatographic characterization of complex formation of pro-IGF-II isoforms with acid labile subunit and IGF-binding proteins associated with non-islet cell tumour induced hypoglycaemia

Objective and designNon-islet cell tumour induced hypoglycaemia (NICTH) is a paraneoplastic phenomenon that is associated with the formation of several isoforms of pro-insulin like growth factor 2 (pro-IGF-II), or so called “big” IGF-II. Disturbance of ternary complex formation by big IGF-II is assumed to be a crucial early event in the pathogenic cascade of hypoglycaemia.By size-exclusion chromatography, we investigated complex formation by adding different naturally occurring isoforms of pro-IGF-II to pooled normal adult serum. Results were compared with the analysis of the serum from a patient with NICTH. ResultsGel filtration experiments with the serum of a patient with NICTH demonstrated that ternary complex formation was severely compromised.The various forms of pro-IGF-II did not induce a shift of IGF-binding protein 3 (IGFBP-3) from 150kD towards smaller binary complexes in the normal adult serum, suggesting that they did not interfere with the interaction between the acid labile subunit and IGFBP-3. Instead, unglycosylated recombinant pro-IGF-II[1–104] was capable of forming a 150kD complex. In contrast, predominantly glycosylated and unglycosylated pro-IGF-II[1–87] eluted in the free unbound form. We showed that mature IGF-II and isoforms of pro-IGF-II were able to phosphorylate the IGF-I receptors of MC7 cells, albeit to a markedly lesser extent than IGF-I. When the patient's serum was tested in this system, the IGF-I receptor phosphorylation activity was considerably less than that in sera from age matched healthy individuals. ConclusionWe postulate that, alongside the presence of big IGF-II in the circulation, additional steps are required to stimulate the release of IGF-II and pro-IGF-II isoforms from IGFBPs in vivo. These factors may be proteases, that are present in the local environment of the tumour and in insulin-sensitive tissues.

Abstract 234: Cigarette Smoke Induction of Mmp9 in Aortic Vascular Smooth Muscle Cells is Mediated by the Jak/stat Pathway

Introduction Cigarette smoke has a strong correlation with abdominal aortic aneurysm (AAA) formation and is characterized by increased levels of MMP9 in the aorta. We hypothesized that smoke-induced MMP9 regulation is mediated by the Jak/Stat pathway thereby contributing to AAA formation. Methods Aqueous extract of cigarette smoke (AEC) was applied to male rat aortic vascular smooth muscle cells (RASMC) for 24 hours (h) in serum free medium (SFM), using SFM alone for controls. Media were collected for MMP2 and MMP9 zymography and proteins extracted from cells for phospho-Stat3 (pStat3), total Stat3 (T-Stat3), phospho-Jak2 (pJak2) and total Jak2 (T-Jak2) western blots. RASMC were transfected by siRNAs for Jak2 and Stat3 before AEC treatment to evaluate induction of MMPs. Coimmunoprecipitation and immunofluorescence microscopy investigated complex formation and cellular distribution of Jak2 and Stat3 following AEC treatment. Results MMP9 was increased in the medium of the AEC treated RASMC (P=0.005) and pro-MMP2 slightly elevated (P=0.055) at 24h compared to controls. By western blot protein extracts from AEC treated RASMC showed up regulation of pStat3, pJak2 and T-Jak2 and stable T-Stat3 compared with controls. siRNA transfection of RASMC for Jak2, Stat3 and a combination of Jak2 and Stat3 reduced MMP9 (P<0.005) and pro-MMP2 (P<0.05) in medium of AEC treated RASMC compared with AEC-treated cells transfected with control siRNA. Coimmunoprecipitation of total proteins from RASMC with total Jak2 antibody revealed increased levels of pStat3 in the AEC treated cells compared to controls, while separation of nuclear and cytoplasmic components followed by coimmunoprecipitation revealed more abundant Jak2/T-Stat3 complexes in the nuclear fraction than the cytoplasm following AEC treatment. Immunofluorescence microscopy revealed increased presence of pJak2, T-Jak2, pStat3 and T-Stat3 in the cytoplasm and nucleus of the AEC treated cells compared to the control cells. Conclusion This study suggests that cigarette smoke may result in AAA formation through Jak/Stat-mediated MMP9 production. siRNA inhibition of the Jak/Stat pathway greatly reduced AEC-induced MMP9 by RASMC and suggests a potential therapeutic target for the treatment of AAA.

Investigating complex formation of palladium in desorption eluates from VP-1P anionite

Complex forms of palladium that are formed in ammonia-nitrate solutions during the desorption of platinum group metals (PGMs) from the VP-1P anionite are investigated. The possibility in principle of using the latter for the removal of PGMs from electrolytes in the refining technology of silver is confirmed.

Effect of Serum on Transfection by Polyethylenimine/Virus-Like Particle Hybrid Gene Delivery Vectors

Murine leukemia virus-like particles (M-VLP) complexed with polymers to promote cellular uptake and endosomal escape represent a new class of effective gene delivery vectors. Building upon recent studies of viral-synthetic hybrid vectors, we report the effects of serum on the formation, activity and stability of PEI/M-VLP complexes. M-VLP were produced by cells grown in serum-supplemented media (M-VLP-S), serum-free media (M-VLP-SF) or serum-free Opti-MEM® I (M-VLP-OM). PEI/M-VLP stoichiometry was varied to investigate complex formation and optimal transfection conditions. The effects of prolonged storage, freeze-thaw cycles, and ultracentrifugation of M-VLP on the stability of vector transduction efficiency were also observed. M-VLP-S required more PEI to form infective complexes than M-VLP-SF and M-VLP-OM. The stoichiometry of PEI/M-VLP-S was dependent on total PEI concentration (7-8 μg/100 μL M-VLP supernatant), while optimal infectivity of PEI/M-VLP-SF and PEI/M-VLP-OM depended on PEI/M-VLP ratios (12-17 μg and 10-14 μg PEI/10(9) M-VLP, respectively). PEI/M-VLP-SF and PEI/M-VLP-OM complexes were significantly more efficient than PEI/M-VLP-S. Stability of the hybrid vectors was not significantly affected by serum. PEI/M-VLP complexes exhibiting increased efficiency were constructed by producing M-VLP in serum-free media. M-VLP could be stored by freezing or refrigeration and concentrated by ultracentrifugation without unacceptable loss of infectivity.

Two-column ion chromatography for investigating equilibrium processes in solutions of complex compounds

Two-column ion chromatography with conductometric detection was used for investigating complex formation processes in the reactions of Cu(II), Ni(II), Co(II), and Pb(II) ions with ethylenediaminetetraacetic acid in aqueous solutions. The ion-chromatographic retention parameters of complex anions were used for determining the charge and composition of the complex particles as the concentration and pH of the carbonate eluent were varied. It was found that, depending on sample acidity, the peak heights of deprotonated chelate anions changed because of a change in the composition of the anions. A method was suggested for calculating the quantitative characteristics of complex formation reactions and the stability constants of the deprotonated chelate compounds using ion chromatography parameters.

Protein Interactions in the Human Methionine Synthase−Methionine Synthase Reductase Complex and Implications for the Mechanism of Enzyme Reactivation

Methionine synthase (MS) is a cobalamin-dependent enzyme. It transfers a methyl group from methyltetrahydrofolate to homocysteine forming methionine and tetrahydrofolate. On the basis of sequence similarity with Escherichia coli cobalamin-dependent MS (MetH), human MS comprises four discrete functional modules that bind from the N- to C-terminus, respectively, homocysteine, methyltetrahydrofolate, cobalamin, and S-adenosylmethionine (AdoMet). The C-terminal activation domain also interacts with methionine synthase reductase (MSR), a NADPH-dependent diflavin oxidoreductase required for the reductive regeneration of catalytically inert cob(II)alamin (which is formed every 200-1000 catalytic cycles of MS) to cob(I)alamin. We have investigated complex formation between the (i) MS activation domain and MSR and (ii) MS activation domain and the isolated FMN-binding domain of MSR. We show that the MS activation domain interacts directly with the FMN-binding domain of MSR. Binding is weakened at high ionic strength, emphasizing the importance of electrostatic interactions at the protein-protein interface. Mutagenesis of conserved lysine residues (Lys1071 and Lys987) in the human activation domain weakens this protein interaction. Chemical cross-linking demonstrates complex formation mediated by acidic residues (FMN-binding domain) and basic residues (activation domain). The activation domain and isolated FMN-domain form a 1:1 complex, but a 1:2 complex is formed with activation domain and MSR. The midpoint reduction potentials of the FAD and FMN cofactors of MSR are not perturbed significantly on forming this complex, implying that electron transfer to cob(II)alamin is endergonic. The kinetics of electron transfer in MSR and the MSR-activation domain complex are similar. Our studies indicate (i) conserved binding determinants, but differences in protein stoichiometry, between human MS and bacterial MetH in complex formation with redox partners; (ii) a substantial endergonic barrier to electron transfer in the reactivation complex; and (iii) a lack of control on the thermodynamics and kinetics of electron transfer in MSR exerted by complex formation with activation domain. The structural and functional consequences of complex formation are discussed in light of the known crystal structure of human activation domain and the inferred conformational heterogeneity of the multidomain MSR-MS complex.

Metal-Complex Formation in Continuous-Flow Ligand-Exchange Reactors Studied by Electrospray Mass Spectrometry

Electrospray ionization mass spectrometry was used to investigate complex formation of different metal complexes in a continuous-flow ligand-exchange reactor. A computer program was developed based on normal equilibrium calculations to predict the formation of various metal-ligand complexes. Corresponding to these calculations, infusion electrospray mass spectrometric experiments were performed to investigate the actual complex formation in solution. The data clearly show good correlation between the theoretically calculated formation of metal-ligand complexes and the experimental mass spectrometric data. Moreover, the approach demonstrates that the influence of the pH can be investigated using a similar approach. Indirectly, these infusion experiments provide information on relative binding constants of different ligands towards a metal-ion. To demonstrate this, a continuous-flow ligand-exchange detection system with mass spectrometric detection was developed. Injection of ligands, with different affinity for the metal-ion, into the reactor shows good correlation between binding constants and the response in the ligand-exchange detection system. Additional information on the introduced ligand, and the complexes formed after introduction of the ligand, can be obtained from interpretation of the mass spectra.

Cyclam cored luminescent dendrimers as ligands for Co(II), Ni(II) and Cu(II) ions

We have investigated the photophysical properties of two dendrimers containing a cyclam core decorated with 4 naphthyl units ( G0), 12 dimethoxybenzene and 16 naphthyl units ( G2). These dendrimers show fluorescence bands that can be assigned to naphthyl localized excited states ( λ max = 337 nm), naphthyl-amine exciplexes ( λ max = 470 nm) and, for G2, naphthyl excimers ( λ max ca. 400 nm). Cyclam is a very good ligand for transition metal ions and we have investigated complex formation between these dendritic ligands and Ni(II), Co(II) and Cu(II), added as nitrate salts. This process can be monitored by the strong changes, both in shape and intensity, observed in the emission spectra of these dendrimers. Complexation with Cu(II) causes not only changes in the relative intensities of the fluorescence bands, but also the appearance of a new absorption band in the near UV spectral region. An analysis of the titration curves has allowed us to obtain clear evidence for the formation of not only 1:1 species, but also 1:2 metal to ligand species. G2 shows a clear preference, compared to G0, in forming complexes with a 1:2 metal-to-ligand stoichiometry, although it possesses very bulky dendrons appended to the cyclam central unit.

The initial step of DNA hairpin folding: a kinetic analysis using fluorescence correlation spectroscopy

Conformational fluctuations of single-stranded DNA (ssDNA) oligonucleotides were studied in aqueous solution by monitoring contact-induced fluorescence quenching of the oxazine fluorophore MR121 by intrinsic guanosine residues (dG). We applied fluorescence correlation spectroscopy as well as steady-state and time-resolved fluorescence spectroscopy to analyze kinetics of DNA hairpin folding. We first characterized the reporter system by investigating bimolecular quenching interactions between MR121 and guanosine monophosphate in aqueous solution estimating rate constants, efficiency and stability for formation of quenched complexes. We then studied the kinetics of complex formation between MR121 and dG residues site-specifically incorporated in DNA hairpins. To uncover the initial steps of DNA hairpin folding we investigated complex formation in ssDNA carrying one or two complementary base pairs (dC–dG pairs) that could hybridize to form a short stem. Our data show that incorporation of a single dC–dG pair leads to non-exponential decays for opening and closing kinetics and reduces rate constants by one to two orders of magnitude. We found positive activation enthalpies independent of the number of dC–dG pairs. These results imply that the rate limiting step of DNA hairpin folding is not determined by loop dynamics, or by mismatches in the stem, but rather by interactions between stem and loop nucleotides.

Energetics and self-compensation ofLiinGaAs

The structural and electronic properties of lithium substitutional acceptors and interstitial donors in crystalline $\mathrm{GaAs}$ are investigated using a density-functional theory-based method. In $p$-type $\mathrm{GaAs}$, $\mathrm{Li}$ tetrahedral interstitial $({\mathrm{Li}}_{TAs}^{+})$ donors have a low formation energy, which is comparable to that of the acceptors $({\mathrm{Li}}_{Ga}^{\ensuremath{-}2})$ in $n$ type, namely, a substitutional $\mathrm{Li}$ on the $\mathrm{Ga}$ site. The donors enhance the solubility of the acceptors and vice versa. In thermodynamic equilibrium, the incorporation of large amounts of $\mathrm{Li}$ will result in severe self-compensation. To examine the kinetics of Li, diffusion mechanisms of ${\mathrm{Li}}_{TAs}^{+}$ and ${\mathrm{Li}}_{Ga}^{\ensuremath{-}2}$ are explored. ${\mathrm{Li}}_{TAs}^{+}$ is highly mobile with a barrier of $0.43\phantom{\rule{0.3em}{0ex}}\mathrm{eV}$, compared to a large barrier of $2.73\phantom{\rule{0.3em}{0ex}}\mathrm{eV}$ for relatively immobile ${\mathrm{Li}}_{Ga}^{\ensuremath{-}2}$. We have also investigated complex formation and dissociation between interstitial donors and substitutional acceptors. The mechanism of lithium self-compensation involving two diffusive ${\mathrm{Li}}_{TAs}^{+}$ cations in two stages to neutralize a ${\mathrm{Li}}_{Ga}^{\ensuremath{-}2}$ anion can be resolved. A comparison with available experiments confirms our findings. The expectation of the results is also consistent with lithium compensation in both strongly extrinsic $p$-type and $n$-type $\mathrm{GaAs}$.

Sweet Potato Acid Phosphatase Immobilized on Glutaraldehyde-Activated Aminopropyl Controlled-Pore Glass: Activation, Repeated Use and Enzyme Fatigue

Sweet potato acid phosphatase was covalently coupled with glutaraldehyde to aminopropyl controlled-pore glass, and used as a pre-column enzyme reactor. The immobilized enzyme reactor (IMER) was continuously operated using an automated chromatographic detection system we developed. Functional evaluation of the IMER was carried out by injecting ten samples on the same day at an injection amount of 1.25 nmol (62.5 nmol per ml) using riboflavin sodium phosphate (FMNs) as a substrate, and by prolonged use for ten months. The IMER exhibited decreased activity after repeated use for a total of 3000 samples, but about 75% of its original activity remained. The conversion rate of FMNs to riboflavin by IMER was increased from 89 to 97% by adding citrate, ethylenediaminetetraacetic acid disodium salt, etc., but especially by adding citrate. The increased conversion of FMNs to riboflavin due to the addition of citrate was probably not due to the chelation of heavy metal ions by citrate. We also investigated complex formation of acid phosphatase with the substrate FMNs using surface plasmon resonance to determine the effect of citrate on the processes of association and/or dissociation between the enzyme and substrate. Enzyme fatigue was also observed during the course of prolonged and repeated use.

Complex formation and degradation in poly(acrylonitrile-co-vinyl acetate) containing metal nitrates

Polymers containing metal-nitrates have been proposed as advantageous precursors for high temperature superconductors such as yttrium-barium-copper-oxide (YBCO). The advantage lies in using conventional polymer processing such as fiber spinning or microlithography before pyrolysis. This research investigated complex formation and degradation in poly(acrylonitrile-co-vinyl acetate) (P(AN-VA)) containing either yttrium nitrate (YN) or barium nitrate (BaN) and follows a similar investigation for copper nitrate. Complex formation was observed in P(AN-VA)/YN but not in P(AN-VA)/BaN. The exothermic nitrate degradation below the P(AN-VA)) cyclization temperature involved the release of NO 3, a reaction with the nitrile group that disrupted cyclization, and, for BaN, P(AN-VA) degradation. For a P(AN-VA)/nitrate ratio of 2/1 there was no cyclization and the degradation temperature was reduced by about 200 °C. The pyrolysis of P(AN-VA)/BaN yielded largely BaCO 3, which is likely to impede the formation of YBCO.

Methylene Blue를 이용한 역미셀에서 물의 세 가지 상태에 대한 분광학적 연구

Cyclohexane에 녹아있는 Triton X-100/n-hexanol/water 계의 비이온성 역미셀(reverse micelle)에 대한 미세 환경정보를 알아보기 위해 methylene blue(MB)를 이용하여 흡광 및 형광 분광법으로 고찰하였다. 역미셀의 극성 중심(polar core)에서 물의 미세 상태에 대한 정보는 MB와 Triton X-100 사이의 착물형성과 용매화변색(solvatochromic)거동을 조사해 봄으로써 얻을 수 있었으며, 역미셀의 안쪽 극성 중심에서 3가지 상태로 존재함을 알 수 있었다. 물의 세 가지 상태에 대한 <TEX>$W(=[H_2O]/[Surf])$</TEX>값은 0.71, 4.98, 7.26으로 측정되었으며, 그에 대한 MB의 형광수명은 <TEX>$15.45 ns{\pm}0.56$</TEX>, <TEX>$12.27 ns{\pm}0.79$</TEX>, 그리고 <TEX>$8.28 ns{\pm}0.82$</TEX>으로 나타났다. In order to find out the microscopic environmental information on the nonionic reverse micelle of Triton X-100/n-hexanol/water in cyclohexane, an absorption and fluorescence spectroscopic study has been conducted using a methylene blue(MB). The information on the microscopic states of water in the polar core of the reverse micelle has been found by investigating complex formation and solvatochromic behavior between MB and Triton X-100. As a result, it was found that there exist three states in the polar core of the reverse micelle. The measured values of <TEX>$W(=[H_2O]/[Surf])$</TEX> for the three states of water are 0.71, 4.98, and 7.26, and the corresponding lifetimes of MB are <TEX>$15.45 ns{\pm}0.56$</TEX>, <TEX>$12.27 ns{\pm}0.79$</TEX>, and <TEX>$8.28 ns{\pm}0.82$</TEX>, respectively.