Study on interaction of mangiferin to insulin and glucagon in ternary system

Spectrochimica Acta Part A 75 (2010) 1584–1591

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Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy journal homepage: www.elsevier.com/locate/saa

Study on interaction of mangiferin to insulin and glucagon in ternary system Hui Lin, Rui Chen, Xiaoyan Liu, Fenling Sheng, Haixia Zhang ∗ State Key Laboratory of Applied Organic Chemistry, Lanzhou University, Lanzhou 730000, China

a r t i c l e

i n f o

Article history: Received 16 September 2009 Received in revised form 28 January 2010 Accepted 11 February 2010 Keywords: Mangiferin Insulin Glucagon Spectroscopy Binding parameters

a b s t r a c t The binding of mangiferin to insulin and glucagon was investigated in the presence and absence of another Peptide by optical spectroscopy. Fluorescence titration experiments revealed that mangiferin quenched the intrinsic fluorescence of insulin and glucagon by static quenching. The ratios of binding constants of glucagon-mangiferin to insulin-mangiferin at different temperatures were calculated in “pure” and ternary system, respectively. The results indicated that the Peptides were competitive with each other to act on mangiferin. Values of the thermodynamic parameters and the experiments of pH effect proved that the key interacting forces between mangiferin and the Peptides were hydrophobic interaction. In addition, UV–vis absorption, synchronous fluorescence and Fourier transform infrared measurements showed that the conformation of insulin and glucagon were changed after adding mangiferin. © 2010 Elsevier B.V. All rights reserved.

1. Introduction A lot of studies related with interaction between small drugs and transport proteins have been widely investigated in the past few years [1–4]. The binding between small molecules and proteins can elucidate the properties of small molecule–protein complex, provide useful information of the delivery or structural features of small molecules, and influence on the concentration, distribution and conformation of target protein in body synchronously [5]. However, few studies were related with the interaction of small drugs and active Peptides in ternary system. In blood, there are a lot of active Peptides which form transient biological interactions with other substances including drugs, protein and so on [6]. These transient biological interactions are dynamic and control the local concentrations of interacting biological pair. Owing to the low concentrations of the Peptides, the transient biological interactions can affect not only the concentrations and forms of Peptides but also the delivery of foreign compounds such as drugs. Thus, it is significant equally to study the interaction of small drugs and Peptides. Diabetes mellitus is found increasing rapidly all over the world. People suffering from diabetes are not able to produce or properly use insulin in the body and have a high content of blood glucose. Both of insulin and glucagon are important Peptides in blood glucose regulation and diabetes treatment. Insulin plays important roles in lowering blood glucose, regulating lipid metabolism, inhibiting of protein degradation, promoting protein synthesis, and affecting growth [7]. Glucagon possess significant roles in

∗ Corresponding author. Tel.: +86 931 4165997; fax: +86 931 8912582. E-mail addresses: [email protected], [email protected] (H. Zhang). 1386-1425/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.saa.2010.02.023

increasing blood glucose, promoting ketogenesis, regulating lipid metabolism, and affecting protein metabolism [8]. Both Peptides have the adversed functions and their concentrations should keep the relatively stable. Up to now, there is no report related with the interaction between small drug molecule and the two Peptides. Most papers related with interaction of small molecule and protein investigated that in a “pure” system [9–12], in which only the studied protein existed in the solution. Here, we firstly try to study the interaction of small molecule and Peptide under the coexistance of another Peptide. Mangiferin (MA) was chosen as the model small molecule, which is the active component of Chinese medicine Anemarrhena asphodeloides Bge., and has a lot of beneficial functions including curing Diabetes. Its binding to bovine serum albumin (BSA) and human serum albumin (HSA) had been studied [13,14]. Spectral analysis was chosen here as a tool to study the interaction because of the convenient manipulation, low spending and abundant theory foundation [15]. 2. Materials and methods 2.1. Materials Insulin (from bovine pancreas) was obtained from Roche Diagnostics Co. (Indianapolis, USA). Glucagon was purchased from Chengdu Kaijie Bio-pharmaceutical Co. Ltd. (Chengdu, China). Stock solution of insulin and glucagon were prepared in buffer solution and kept in the dark at 277 K. MA (90%) was obtained from Chengdu tianfutang Economy Co. Ltd. (Chengdu, China). Stock solution of MA (8 × 10−5 mol L−1 ) was prepared in buffer solution for FT-IR experiments and stock solution of MA (1.0 × 10−3 mol L−1 ) was prepared in ethanol for UV–vis and fluorescence experiments.

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The buffer solution (pH 7.40) consisted of Tris (0.1 mol L−1 ) and HCl (0.1 mol L−1 ). pH was checked with a PHSJ-3F pH-meter (Shanghai Precision Scientific Instruments Co. Ltd., Shanghai, China). Other chemicals were of analytical grade and the double-distilled water was used throughout the experiments. 2.2. Apparatus and methods The UV–vis absorption spectra were recorded on a TU-1810 spectrophotometer (Purkinje, China) at 298 K in the range of 250–400 nm equipped with 1.0 cm path length quartz cells. Fluorescence measurements were recorded on RF-5301PC fluorophotometer (Shimadzu, Japan) using 5 nm/5 nm bandwidths at excitation and emission wavelength of 276 and 300–500 nm, respectively. Synchronous fluorescence spectra were recorded with the excitation and emission slit widths of 5 nm/5 nm, and the range of synchronous scanning were:  = 15,  = 60 nm. Fluorescence titration experiment was accomplished as following: 3.0 mL Peptides (insulin, glucagon or mixture of them) solution at concentration of 1.5 × 10−6 mol L−1 , was titrated by successive additions of 10 ␮L stock solution of MA (1.0 × 10−3 mol L−1 ). Titrations were done manually by using micro-injector, and the fluorescence spectra were then measured at three temperatures (296, 303, and 310 K). FT-IR measurements were carried out at room temperature on a Nicolet Nexus 670 FT-IR spectrometer (USA) equipped with a germanium attenuated total reflection (ATR) accessory, a DTGS KBr detector and a KBr beam splitter. All spectra were taken via the attenuated total reflection (ATR) method with a resolution of 4 cm−1 and 60 scans. Spectra processing procedures: The absorbance of reference solution from the spectra of sample solution was subtracted to get the FT-IR spectra of proteins. The subtraction criterion was that the original spectrum of protein solution between 2200 and 1800 cm−1 was featureless [16]. 3. Results and discussion 3.1. Effect of MA on Peptides fluorescence spectra The fluorescence spectra of insulin, glucagon and the mixture of insulin and glucagon in the presence of different concentrations of MA were shown in Fig. 1(a)–(c), respectively. Insulin shows a fluorescence emission with a peak at 307 nm and the excitation wavelength of 276 nm, and MA is almost none fluorescent under the present experiment conditions [13]. With the increasing concentration of MA, the fluorescence intensity of insulin decreased remarkably. However, the maximum emission wavelength was hardly changed. The results indicated the formation of a complex between MA and insulin. Glucagon shows a strong fluorescence emission with a peak at 346 nm at the excitation wavelength of 276 nm. With the increasing concentration of MA, the fluorescence intensities of glucagon decreased remarkably too, and there was obvious em blue shift of maximum emission wavelength with the addition of MA. It implied formation of a complex between MA and glucagon and changes in Peptide conformation. The fluorescence intensity of the mixture of insulin and glucagon decreased at 307 and 344 nm, respectively, which indicated the information of both Peptide. The quenching phenomena were similar to those in Fig. 1(a) and (b). 3.2. Quenching mechanism A variety of molecular interactions can result in quenching, including excited-state reactions, molecular rearrangements, energy transfer, ground-state complex formation, and collisional

Fig. 1. Fluorescence quenching spectra of insulin (a), glucagon (b) and the mixture of them (c) at different concentrations of MA; Cprotein = 1.5 ␮M; CMA from a to g: 0 (a), 3.33 (b), 6.66 (c), 9.99 (d), 13.32 (e), 16.65 (f), and 19.98 ␮M (g). ex = 276 nm.

quenching [17]. Fluorescence quenching can proceed via different mechanisms usually classified as dynamic quenching, static quenching or simultaneous static and dynamic quenching. For dynamic quenching, fluorescence quenching can be described by Stern–Volmer equation [18]: F0 = 1 + kq 0 [Q ] = 1 + Ksv [Q ] F

(1)

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Fig. 2. Stern–Volmer plots for quenching of insulin and glucagon by MA in “pure” and ternary system at 296, 303 and 310 K.

F0 and F are fluorescence intensities of biomolecule before and after addition of quencher, respectively. kq is the quenching rate constant of the biomolecule (kq: L mol−1 s−1 ), Ksv is the Stern–Volmer dynamic quenching constant (Ksv: L mol−1 ),  0 is the average lifetime of the biomolecule without quenching ( 0: 10−8 s) and [Q] is the concentration of quencher. In order to investigate the quenching mechanism, the procedure of the fluorescence quenching was first assumed to be a dynamic quenching progress. Stern–Volmer

plots of insulin and glucagon with MA were shown in Fig. 2. The corresponding Stern–Volmer dynamic quenching constants at different temperatures were summarized in Table 1. It can be found the Stern–Volmer dynamic quenching constants of insulin–MA in “pure” and ternary system were almost same within error limit, which indicated that the quenching rate was independent of temperature and was not influenced by the existence of glucagon. The same conclusion was obtained for the system of

Table 1 Stern–Volmer quenching constants of the insulin–MA and glucagon–MA in “pure” and ternary system at different temperatures. Systems

T (K)

Ksv (×104 L mol−1 )

kq (×1012 L mol−1 s−1 )

SDa

Rb

Insulin–MA in “pure” system

296 303 310

2.975 3.145 2.922

2.975 3.145 2.922

0.0063 0.0190 0.0077

0.9995 0.9963 0.9993

Insulin–MA in ternary system

296 303 310

3.158 3.090 3.125

3.158 3.090 3.125

0.0115 0.0194 0.0111

0.9986 0.9960 0.9987

Glucagon–MA in “pure” system

296 303 310

5.613 5.420 4.998

5.613 5.420 4.998

0.0300 0.0226 0.0316

0.9971 0.9982 0.9959

Glucagon–MA in ternary system

296 303 310

5.771 5.462 5.060

5.771 5.462 5.060

0.0223 0.0115 0.0290

0.9985 0.9995 0.9966

a b

The standard deviation for the Ksv values. The correlation coefficient.

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Fig. 3. Plots of log[(F0 − F)/F] vs. log[Q] in “pure” and ternary system at 296, 303 and 310 K.

glucagon and MA. For dynamic quenching, the maximum scatter collision quenching constant kq of various quenchers with the biopolymer is 2 × 1010 L mol−1 S−1 [19]. The values of kq in all systems studied at three temperatures were much greater than it, which indicated that the quenching of insulin and glucogon are not initiated by dynamic quenching, but probably by static quenching resulting from the formation of insulin–MA complex or glucagon–MA complex [20].

Comparing with the Ksv of the complex of MA–BSA [13], to those of insulin–MA and glucagon–MA, it was found MA could quench the fluorescence of BSA much more effectively. For investigating the relationship between Ksv and the molecular volumes of Peptides, the molecular volumes were calculated roughly with hyperchem software (Hyperchem 7.0). The results showed that the Ksv has no relationship with Peptide volumes (insulin: 10805.69 Å3 , glucogon: 7396.42 Å3 ).

Table 2 Binding parameters and thermodynamic parameters of insulin–MA and glucagon–MA systems at different temperatures. ka (×104 L mol−1 )

S (J mol−1 K−1 )

G (kJ mol−1 )

31.94

193.54

−25.35 −26.70 −28.06

1.02 1.15 1.19

98.88

422.77

−26.26 −29.22 −32.18

4.509 14.79 30.66

0.98 1.10 1.17

104.627

443.12

−26.54 −29.64 −32.74

9.550 21.38 22.30

1.05 1.13 1.14

46.81

254.45

−28.51 −30.29 −32.07

Systems

T (K)

Insulin–MA in “pure” system

296 303 310

2.951 4.135 5.300

1.00 1.04 1.05

296 303 310

3.802 14.12 23.20

296 303 310 296 303 310

Insulin–MA in ternary system

Glucagon–MA in “pure” system

Glucagon–MA in ternary system

n

H (kJ mol−1 )

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Fig. 4. Plots of Ka1 (glucagon–MA)/Ka2 (insulin–MA) vs. T.

3.3. Analysis of binding constants and binding sites For static quenching, the relationship between fluorescence quenching intensity and the concentration of quenchers can be described by Eq. (2) [21,22]: lg

F − F  0 F

= lg Ka + n lg Q

Fig. 5. Plots of Ka vs. pH in “pure” and ternary system.

where Ka is the binding constant (Ka : L mol−1 ), and n is the number of binding sites per the biomolecule with fluorophore. Fig. 3 shows the double-logarithm curves of insulin–MA (a) and glucagon–MA (b) in “pure” and ternary system under different temperatures. The results of the all systems studied were summarized in Table 2. All correlation coefficients were larger than 0.995, which indicated that the interaction between Peptide (insulin or glucagon) and MA agreed well with the site-binding model underlined in Eq. (2). From Fig. 4, it could be found the ratios of binding constants of glucagon–MA to insulin–MA at 303 and 310 K in “pure” system were higher than those in ternary system, but lower a little at 296 K. Glucagon acted strongly with MA than insulin in “pure” system under 310 K, which is closed to the normal body temperature. In ternary system, the interaction of glucagon–MA became much weaker and the interaction of insulin–MA became much stronger until both had similar binding constants under 310 K. The solution of insulin is a dynamic equilibrium of monomers, dimers, tetramers, and so on, depending on the concentration, pH, ionic strength, temperature, and solvent composition [23–25]. The insulin may exist as more dimers on the condition of “pure” system than in ternary system. The binding site was possibly hidden in the core of dimers, which hindered the binding of insulin with small molecule MA. In the ternary system, glucagon may destroy some insulin dimers and

(2)

Fig. 6. UV absorption spectra of insulin–MA (a) and glucagon–MA (b): Cprotein = 15 ␮M; CMA from b to g: 0 (b), 3.33 (c), 6.66 (d), 9.99 (e), 13.32 (f), and 16.65 ␮M (g), the spectra of MA, CMA = 3.33 ␮M (a).

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some binding sites of insulin were exposed, which increased the possibility of insulin binding to MA. Meanwhile, owing to smaller molecular size than insulin, glucagon was maybe enwrapped by insulin and lost the competition to bind MA. According to Eq. (2), the binding sites of the insulin–MA and glucagon–MA were calculated to be 1.00 approximately. The binding sites in “pure” system were also measured by UV–vis experiments with mole ratio method [26]. It was found the measured results were consistent with the calculated above. 3.4. Thermodynamic parameters and nature of the binding forces The interaction forces between drug and biomolecule may involve hydrophobic force, electrostatic interactions, van der Waals interactions, hydrogen bonds, steric contacts within the antibodybinding site, etc. In order to elucidate the interaction of MA to insulin and glucagon, the thermodynamic parameters were calculated from Eqs. (3) and (4). If the temperature does not vary significantly, the enthalpy change (H) can be regarded as a constant and were calculated according to the van’t Hoff equation: ln k =

−H S + RT R

(3)

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Thus, a plot of ln k vs. 1/T enabled the determination of H and S for the binding. The free energy change (G) was estimated from the following equation: G = H − TS

(4)

where k is the binding constant at the corresponding temperature and R is the gas constant. The thermodynamic parameters for the interaction of MA to insulin and glucagon were summarized in Table 2. The negative sign for G means that the interaction progress was spontaneous. The positive H and S values indicated that hydrophobic force may play a major role in the binding between MA and the both Peptides [27]. The effect of pH values on the binding constants of insulin–MA and glucagon–MA was studied (Fig. 5). The trend of change on Ka in ternary system was similar to that in “pure” system. The values of binding constants of Peptides–MA are biggest at pH 7.40 in “pure” and ternary systems. MA existed as ionic form and was negatively charged while pH 7.40 [28]. The isoelectric points of insulin and glucagons are 5.35 and 7.00, respectively [29], the charges of Peptides surface are negative at pH 7.40. Hence, electrostatic attraction force and hydrogen bond interaction were not the key roles at this environment, and hydrophobic force may play a major role in the binding between MA and Peptides (insulin or glucagon).

Fig. 7. Effect of MA on the synchronous spectra of insulin and glucagon in “pure” and ternary system. Cprotein = 15 ␮M; CMA from a to g: 0 (a), 3.33 (b), 6.66 (c), 9.99 (d), 13.32 (e), 16.65 (f), and 19.98 ␮M (g).  = 15 nm (a and c) (insulin, ex = 240 nm);  = 60 nm (b and d) (glucagon, ex = 240 nm).

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Table 3 Secondary structure of insulin and glucagon. Systems

␤-Turn (%)

␣-Helical (%)

Random coil (%)

␤-Sheet (%)

Insulin-free Insulin–MA Glucagon-free Glucagon–MA

12.88 16.31 29.37 25.09

17.15 31.59 10.00 22.86.

19.00 8.02 33.34 17.33

50.97 39.51 27.29 34.73

3.5. Conformation investigation To explore the structural change of insulin and glucagon, UV–vis absorption, synchronous fluorescence and FT-IR were performed. The UV–vis spectrum of MA, insulin, glucagon, insulin–MA and glucagon–MA were recorded as shown in Fig. 6. With the increasing concentration of MA, the UV intensities of insulin and glucagon increased at 280 nm and had a blue shift due to formation of complex between MA to proteins [14,30]. The results implied that the microenvironment around chromophore of insulin and glucagon was changed upon addition of MA. The synchronous fluorescence spectra offers the characteristics of tyrosine residues and the tryptophan residue of protein when the wavelength interval () is 15 and 60 nm, respectively ( = emission − excitation ) [31]. Insulin has no tryptophan residues, synchronous fluorescence spectra of insulin upon addition of MA gained at  = 15 nm are shown in Fig. 7(a) and (c). It can be found that the fluorescence intensity of tyrosine decreased and had a slightly red shift at maximum emission in “pure” system, which indicated that the tyrosine residues are placed in a more hydrophilic environment and their microenvironment is rearranged [31] thus resulted in the conformational changes of insulin. However, the fluorescence intensity of tyrosine had no shift in ternary system, the possible reason was that the synchronous fluorescence spectra of glucagon influenced the spectra of insulin. Glucagon has both of tyrosine residues and tryptophan residues. However, tryptophan residues have stronger fluorescence intensities. Fig. 7(b) and (d) gave the synchronous fluorescence spectra of glucagon upon addition of MA gained at  = 60 nm, which shows that the fluorescence intensity of tryptophan decreased and had a red shift at maximum emission in “pure” and ternary systems. It implied the conformation of glucagon is changed due to that tryptophan residues are exposed more to the aqueous phase [31]. Detailed information of insulin and glucagon conformation after adding MA was investigated with FT-IR difference spectra as shown in Fig. 8. The spectra of Fig. 8a (1) and b (1) were spectra of insulin before and after adding MA, respectively. The spectra of Fig. 8a (2) and b (2) were spectra of glucagon before and after adding MA, respectively. From Fig. 8, it concluded that the secondary structure of insulin and glucagon were changed after the addition of MA, because the peak position of amide I band (insulin: 1643.28 cm−1 , glucagon: 1651.65 cm−1 ) and amide II band (insulin: 1559.60 cm−1 , glucagon: 1546.85 cm−1 ) had small shifts and their peak shapes were also slightly changed [32–35]. The results confirmed that there was a reaction happened between MA and the both Peptides. A quantitative analysis of the protein secondary structure of insulin and glucagon before and after the interaction with MA in Tris–HCl buffer was finished. Based on the literature [36], the component bands of amide I was attributed according to the well-established assignment criterion. Then the percentages of each secondary structure of insulin and glucagon can be calculated and Table 3 shows the calculated results. It was found that the ␤-turn, ␣-helical, random coil and ␤-sheet structures in the secondary structure of insulin and glucagon were changed after adding MA. The results confirmed that the binding of MA to insulin

Fig. 8. FT-IR spectra of free insulin (a1) and glucagon (b1). Difference spectra [(protein solution + MA solution) − (MA solution)] in Tris–HCl buffer (pH 7.40), insulin (a2), glucagon (b2). (Cprotein = 10 ␮M CMA = 30 ␮M).

and glucagon had changed the secondary structure of insulin and glucagon. 4. Conclusion In this paper, the binding MA to insulin and glucagon has been investigated for the first time by fluorescence method combined with UV–vis and FT-IR spectroscopy techniques under simulative physiological condition. The results suggested that MA quenches the intrinsic fluorescence of insulin and glucagon through static quenching mode. Conformations of both Peptides were changed slightly after adding MA. Hydrophobic interactions played a key role in the binding progress of MA to insulin and glucagon. The experimental results indicated that insulin outvied glucagons to bind MA at 303 and 310 K when they are coexisted in the same system. Acknowledgements This work was supported by the National Natural Science Foundation of China (NSFC) Fund (No.20775029), Program for New Century Excellent Talents in University (NCET-07-0400) and the Central Teacher Plan in Lanzhou University. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.saa.2010.02.023.

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References [1] F.-Q. Cheng, Y.-P. Wang, Z.-P. Li, C. Dong, Spectrochim. Acta A 65 (2006) 1144–1147. [2] J.Q. Liu, J.N. Tian, Z.D. Hu, X.G. Chen, Bioorg. Med. Chem. 73 (2004) 443–450. [3] S.Y. Bi, D.Q. Song, Y.H. Kan, D. Xu, Y. Tian, X. Zhou, H.Q. Zhang, Spectrochim. Acta A 62 (2005) 203–212. [4] Y.H. Zhang, L.J. Dong, J.Z. Li, X.G. Chen, Talanta 76 (2008) 246–253. [5] D.H. Ran, X. Wu, J.H. Zheng, J.H. Yang, H.P. Zhou, M.F. Zhang, Y.J. Tang, J. Fluoresc. 17 (2007) 721–726. [6] S. Ohlson, S. Shoravi, T. Fex, R. Isaksson, Anal. Biochem. 359 (2006) 120–123. [7] M.Y. Xu, G.H. Lu, M.D. Chen, Diabetes Mellitus, first ed., Shanghai Scientific and Technical Publishers, Shanghai, 2003, pp. 55–56. [8] M.Y. Xu, G.H. Lu, M.D. Chen, Diabetes Mellitus, first ed., Shanghai Scientific and Technical Publishers, Shanghai, 2003, pp. 60–61. [9] M. Bogdan, A. Pirnau, C. Floare, C. Bugeac, J. Pharm. Biomed. Anal. 47 (2008) 981–984. [10] Y.-J. Hu, W. Li, Y. Liu, J.-X. Dong, S.-S. Qu, J. Pharm. Biomed. Anal. 39 (2005) 740–745. [11] X.-H. Liu, P.-X. Xi, F.-J. Chen, Z.-H. Xu, Z.-Z. Zeng, J. Photochem. Photobiol. B 92 (2008) 98–102. [12] P.B. Kandagal, S.S. Kalanur, D.H. Manjunatha, J. Seetharamappa, J. Pharm. Biomed. Anal. 47 (2008) 260–267. [13] H. Lin, J.F. Lan, M. Guan, F.L. Sheng, H.X. Zhang, Spectrochim. Acta A 73 (2009) 936–941. [14] Y.Y. Yue, X.G. Chen, J. Qin, X.J. Yao, J. Pharm. Biomed. Anal. 49 (2009) 753–759. [15] J.H. Li, C.L. Ren, Y.H. Zhang, X.Y. Liu, X.J. Yao, Z.D. Hu, J. Mol. Struct. 885 (2008) 64–69. [16] T. Yuan, A.M. Weljie, H.J. Vogel, Biochemistry 37 (1998) 3187–3195.

1591

[17] F.L. Cui, L.X. Qin, G.S. Zhang, Q.F. Liu, X.J. Yao, B.L. Lei, J. Pharm. Biomed. Anal. 48 (2008) 1029–1036. [18] J.R. Lakowicz, Principles of Fluorescence Spectroscopy, second ed., Plenum Press, New York, 1999, pp. 237–265. [19] W.R. Ware, J. Phys. Chem. 66 (1962) 455–458. [20] Y.-P. Wang, Y.-L. Wei, C. Dong, J. Photochem. Photobiol. A 177 (2006) 6–11. [21] X.Z. Feng, R.X. Jin, Y. Qu, X.W. He, Chin. Univ. 17 (1996) 866. [22] X.F. Wei, H.Z. Liu, J. Chin. Anal. Chem. 28 (2000) 699. [23] Y. Zhang, Y.J. Deng, M. Lu, J. Colloid Interf. Sci. 337 (2009) 322–331. [24] W.D. Lougheed, H. Woulfe-Flanagan, J.R. Clement, A.M. Albisser, Diabetologia 19 (1980) 1. [25] J. Brange, L. Langkjoer, Pharm. Biotechnol. 5 (1993) 315. [26] C.D. Chriswell, A.A. Schilt, Anal. Chem. 47 (1975) 1623–1629. [27] P.D. Ross, S. Subramanian, Biochemistry 20 (1981) 3096–3102. [28] B. Gómez-Zaleta, M.T. Ramírez-Silva, A. Gutiérrez, E. González-Vergara, M. Güizado-Rodríguez, A. Rojas-Hernández, Spectrochim. Acta A 64 (2006) 1002–1009. [29] A.S.R. Andrade, L. Vilela, H. Tunes, Braz. J. Med. Biol. Res. 30 (1997) 1421–1426. [30] K.H. Ulrich, Pharmacol. Rev. 33 (1981) 17–53. [31] C.-X. Wang, F.-F. Yan, Y.-X. Zhang, L. Ye, J. Photochem. Photobiol. A 192 (2007) 23–28. [32] S. Wi, P. Pancoka, T.A. Keiderling, Biospectroscopy 4 (1998) 93–106. [33] K. Rahmelow, W. Hubner, Anal. Biochem. 241 (1996) 5–13. [34] X.-H. Liu, P.-X. Xi, F.J. Chen, Z.H. Xu, Z.Z. Zeng, J. Photochem. Photobiol. B 92 (2008) 98–102. [35] Y. Li, W.Y. He, Y.M. Dong, F.L. Sheng, Z.D. Hu, Bioorg. Med. Chem. 14 (2006) 1431–1436. [36] M. Jiang, M.X. Xie, D. Zheng, Y. Liu, X.Y. Li, X. Chen, J. Mol. Struct. 692 (2004) 71–80.