Interaction of DMAPIP-b with different analyte

Molecular TRANSFER gate

6.2. Construction of molecular logic gate and keypad lock system by inducing the inter molecular proton transfer

6.3.1. Interaction of DMAPIP-b with different analyte

DMAPIP-b has an absorption maximum at 345 nm in acetonitrile. With gradual addition of Fe³⁺, a new band maxima appears at 390 nm at the expense of 345 nm band (Figure 6.3.1.A). The red shift in the spectrum confirms the binding of metal ions at pyridyl and/or imidazo ring nitrogen.¹⁵⁹ It is due to the increase of charge flow from the

0 0.02 0.04 0.06 0.08 0.1

0 10 20 30

Absorbance

[Fe³⁺] (μM) (B)

0 0.03 0.06 0.09 0.12

250 300 350 400 450

Absorbance

Wavelength (nm) 20

[Fe⁺³] (µM) 0

(A)

Figure 6.3.1. (A) Absorption spectra of DMAPIP-b in acetonitrile with increasing Fe³⁺ concentration. (B) Increase in absorbance of DMAPIP-b at 390 nm with addition of Fe³⁺.

donor dimethylamino group to the acceptor moiety. The binding constant of the receptor is determined from the nonlinear curve fitting analysis of the titration curve using the equation 6.3.1 (Figure 6.3.1.B).

𝐴 = 𝐴₀+^{(𝐴𝑓−𝐴0)}

2𝐶ℎ [ 𝐶ℎ+𝐶𝑔+¹

𝐾𝑎−[(𝐶ℎ+𝐶𝑔+¹

𝐾𝑎)²−4𝐶ℎ𝐶𝑔] 1

2] (eq. 6.3.1.)

where A0, Af and A refer the absorbance of DMAPIP-b, Fe³⁺-DMAPIP-b complex and any intermediate Fe³⁺ concentration with DMAPIP-b. Ka refers the Fe³⁺ binding constant of DMAPIP-b. Ch and Cg indicate the concentration of DMAPIP-b and Fe³⁺, respectively.

The binding constant for the complex is 9.1 x 10⁶ M^-1. Along with the longer wavelength band a blue shifted band at 285 nm is also observed with a quasi isobestic point at 300 nm (Figure 6.3.1.A). Binding of metal ion with dimethylamino group decreases the charge flow. Accordingly, the hypsochromic band can be assigned to the complex formed due to the binding of metal ion at dimethyl amino group. The saturation limit is observed at around 20 μM of [Fe³⁺].

Upon excitation at 345 nm the molecular emission appears at 406 nm, but the intensity of the 406 nm emission band gradually decreases with the addition of Fe³⁺

(Figure 6.3.2.A). In presence of 20 μM of metal ion, the quantum yield of DMAPIP-b decreases from 0.76 to 0.02. However, upon excitation at 406 nm, two new emission bands of fluorophore-metal ion complexes appear at 432 nm and 560 nm (Figure 6.3.2.B).

The fluorescence decay measurements also suggest the emission from two different complexes when monitored at 432 nm (Table 6.3.1.). To find out the stoichiometric ratio of Fe³⁺ and DMAPIP-b in the complex, Job’s plot was constructed with the 432 nm emission of the complex. It confirms the 1:1 binding nature (Figure 6.3.3.).

0.0E+0 4.0E+5 8.0E+5 1.2E+6 1.6E+6

416 516 616 716

Intensity (a.u.)

Wavelength (nm) 0 20

Fe³⁺(µM) (B)

0.0E+0 1.0E+7 2.0E+7 3.0E+7 4.0E+7 5.0E+7

360 410 460

Intensity (a.u.)

Wavelength (nm) 0

20

Fe³⁺(µM) (A)

Figure 6.3.2. Emission spectra of DMAPIP-b with gradual increase of Fe³⁺ concentration,(A) 𝑒𝑥 = 345 nm and (B) _𝑒𝑥 = 406 nm.

Molecular Logic Gate

Table 6.3.1. Fluorescence lifetime (τ, ns) of DMAPIP-b in presence of different substrate.

Species τ χ2

DMAPIP-b^a 1.53 1.05

Fe³⁺-DMAPIP-b^b 0.33 (70.25 %)

1.86 (29.75 %)

1.09

F^--Fe³⁺-DMAPIP-b^a 1.53 1.01

_{𝑒𝑥= 375 nm,} ^[a]_{𝑒𝑚= 406 nm,} ^[b]_𝑒𝑚= 432 nm. The relative amplitudes of two different complexes have been shown in parenthesis.

The prototropic studies of DMAPIP-b and analogues molecules established that the protonation might occur at the imidazole and pyridyl nitrogens to form two different kind of monocations.^{109, 159} The emission spectrum of the monocation formed by the

0.0E+0 5.0E+3 1.0E+4 1.5E+4 2.0E+4 2.5E+4

0.4 0.6 0.8 1

Intensity at 432 nm (a.u)

1/(1+[DMAPIP-b]/[Fe³⁺]

0 0.2 0.4 0.6 0.8 1 1.2

250 350 450

Normalized Intensity

Wavelength (nm)

b a c

(B)

0 0.3 0.6 0.9 1.2

300 500 700

Normalized Intensity

Wavelength (nm) a

b c

(A)

Figure 6.3.3. Job’s plot for Fe³⁺-DMAPIP-b complex. Due to the interference of the DMAPIP-b emission at low mole fraction of [Fe³⁺], those points were not included in the plot).

Figure 6.3.4. (A) Normalized emission spectra of (a) DMAPIP-b, 𝑒𝑥= 280 nm, (b) DMAPIP-b with 20 μM of Fe³⁺, _𝑒𝑥= 280 nm, (c) DMAPIP-b with 20 μM of Fe³⁺, _𝑒𝑥= 406 nm. (B) Normalized excitation spectra of (a) DMAPIP-b, _𝑒𝑚 = 406 nm, (b) DMAPIP-b with 20 μM of Fe³⁺, _𝑒𝑚 = 350 nm, (c) DMAPIP-b with 20 μM of Fe³⁺, _𝑒𝑚 = 560 nm.

protonation of the pyridyl nitrogen is more red-shifted than that of the monocation formed by the protonation of the imidazole nitrogen.¹⁴⁹ Accordingly, the band at 432 nm and 560 nm can be assigned to the complexes formed by the binding of ferric ion at imidazole nitrogen and pyridyl nitrogen, respectively. The larger bathochromic shift in the spectra of pyridyl complex are due to the higher conjugation in this complex (as in this complex metal binds at the terminal (pyridyl) nitrogen which increases the charge flow from the dimethylamino group to the pyridyl ring). However, excitation at 280 nm shows the emergence of a new band in addition to the main band at ~350 nm. This is due to the emission from the complex formed by the binding of Fe³⁺ at dimethylamino nitrogen of DMAPIP-b (Figure 6.3.4.A). This assignment is based on the fact that the binding of the metal ion at dimethylamino nitrogen decreases the charge flow from the dimethylamino group to the acceptor, which causes a blue shift compared to the uncomplexed molecule. The excitation spectra corresponding to all three new bands corroborate the existence of three different Fe³⁺-complexes in the ground state (Figure 6.3.4.B).

The molecular fluorescence at 406 nm can be recovered by the addition of anion to the complex. The process occurs more efficiently with increasing the anionic strength (basicity). The efficiency of different anions follows the order F^- > OAc^- > Cl^- > I^- > SCN^- ~ H²PO^4-> NO^3- >> HSO^4- (Figure 6.3.5.). The fluorescence of the molecule is completely regained in presence of equivalent amount of fluoride anion (20 µM). The fluorescence lifetime obtained at 406 nm (Table 6.3.1) exactly matches with the earlier reported

Figure 6.3.5. Regaining of fluorescence of DMAPIP-b by different anions in presence of 20 µM of Fe³⁺, _𝑒𝑥

= 345 nm.

Molecular Logic Gate

fluorescence lifetime of DMAPIP-b.¹⁵⁹ This confirms that the addition of anions to the complex solution liberates the free fluorophore in solution.

The absorption spectra of DMAPIP-b with increasing fluoride concentration is shown in Figure 6.3.6.A. Unlike Fe³⁺, little amount of fluoride ([F^-] < 20 µM) does not affect the absorption spectrum of DMAPIP-b. Only at higher concentrations changes are observed in the absorption spectrum. Same as absorption spectrum, up to 20 µM of fluoride, the emission spectrum of DMAPIP-b exhibits negligible change but upon further increase in anion concentration, a new blue shifted band appears at the cost of the 406 nm emission band with an isoemissive point at 389 nm (Figure 6.3.6.B). As reported earlier, the absorption spectrum and the emission spectrum of DMAPIP-b undergo a blue shift when the hydrogen of the imidazole >NH group is deprotonated to form anion.⁸² The formation of negative charge on the imidazole ring increases the electron density on the acceptor moiety, which decreases the charge transfer from donor to acceptor moiety.¹⁰⁹ Therefore the new blue shifted emission band is attributed to the deprotonated DMAPIP-b anion which generates in the excited state.