p -ANISIDINE

. The objective of this work is to study the release of p -anisidine by the hydrolysis of synthesized Schiff bases N-vinylbenzylidene-p -anisidine (Im) as a monomer and its copolymers (Cp 1 , Cp 2 ) with N,N-dimethylacrylamide (DMA). The hydrolysis behavior and kinetics are investigated in homogeneous media composed of ethanol/water (80% v/v) at 37 °C and at pH range of 4.0-10.0. The variation of the concentration over time is measured in thermostatically cells using UV-Vis spectroscopy. The results showed that the imine function hydrolysis obeyed to the first order for all compounds; the experimental kinetic constants are determined and the pH–rate diagram profile is established. It is noticed that the hydrolysis of imine function is extremely dependent on its carrier and the pH medium.


INTRODUCTION
Imine is sometimes called a Schiff base, its formation and hydrolysis have been developed by Patai [1].These compounds are highly interesting in biological chemistry [2][3][4][5] and in biomedical fields, the Schiff base-linked protein and polypeptide hydrogels are very interesting [6].Schiff bases are also known for their corrosion inhibition efficiency for steel in acidic solution [7] and a variety of active molecules have been grafted using Schiff base link [8][9][10][11].As well, the mechanism of Schiff base synthesis is tracked [12] and the effect of substitutes on Schiff bases' hydrolysis and kinetics has been largely studied [13,14].However, the present research is focused on the influence of structural parameters of aromatic polymeric backbone on the hydrolysis of Schiff bases which can provides supplementary data related to the chemical behavior of the imine function in aqueous media.
The first work carried out in homogeneous buffered medium is from Kamogawa et al. [15], their results showed that the hydrolysis of Schiff bases monomers derived from (m,p)vinilbenzaldehyde or (m,p)-aminostyrene is more rapid than their copolymers with N-vinyl-2pyrrolidone (VP).
Earlier works have introduced the catalytic effect of acids and bases on the hydrolysis of N-(m,p)-vinylbenzylidene-p-anisidine and its copolymer with N-vinyl-2-pyrrolidone at different incorporation rates in different pH of homogeneous buffered and hydro-ethanolic media [16].
It has been found that p-anisidine (4-methoxyaniline) is a new Δ6 desaturation inhibitor, for Mortierella alpina strains [17].In this work, as an active ingredient, p-anisidine is grafted on monomer and copolymers using imine link to get new controlled formulations which can release it progressively.So, in our previous study [18], N-(m,p)-vinylbenzylidene-p-anisidine Schiff base was synthesized and copolymerized with N,N-dimethyl-acrylamide (DMA) at two molecular ratios, where the hydrolysis of imine function was studied only in acidic homogeneous and heterogeneous media at pH = 4.7 and 4, respectively [18].Hence, the present paper is devoted to the study of the hydrolysis kinetics of these new p-anisidine based monomer and copolymers in homogeneous media at different pH; the work is intended to identify and discuss the hydrolysis mechanism in one hand and in the other hand to determine the effect of copolymerization on the hydrolysis constant.So, using these different carriers, i.e. monomer and copolymers, the panisidine release and hydrolysis can be modified and controlled over time.

Hydrolysis kinetics and UV-Vis analysis
The hydrolysis kinetics of Schiff bases monomer (Im) and copolymers (Cp1) and ( Cp2) are studied at a temperature of 37±0.1 °C using an UV-Vis spectrophotometer (Shimadzu UV-2401 PC; Tokyo, Japan) equipped with thermostatically cells.The kinetics are conducted in homogeneous media composed of buffer solutions which are prepared in 80:20 ethanol/water (v:v) where the ionic strength is maintained at 0.01 and the pH range varied from 4.0 to 10.0.The buffer solutions are prepared according to the methods described by Perrin [19], Michaelis-Mizutani [20] and Bates et al. [21] where water is substituted by a mixture of ethanol:water at 80:20 (v:v).A Tacussel digital pH meter is used for the pH measurements in cells.
For hydrolysis study procedure, the UV-Vis reference cell contained 2.70 mL of buffer solution (in 80:20 of ethanol:water), 0.06 mL of double distilled water and 0.24 mL of ethanol.However, the analysis cell contained 2.70 mL of buffer solution (in 80:20 of ethanol:water), 0.06 mL of double distilled water and 0.24 mL of 10 -3 M ethanolic solution of a monomer (Im) or a copolymer (Cp1 or Cp2).The total volume of liquid in cell is maintained at 3 mL.In these conditions, the initial concentration of all compounds in the analysis cell is kept at 8 x 10 -5 M. The molar masses of copolymers (MCp) are calculated on the basis of the repeating unit or pattern in copolymer according to the following equation: where and are molar masses of dimethylacrylamide (DMA) and vinylbenzylidene-panisidine (Im), respectively.

Hydrolysis study of monomer and copolymers Schiff bases
The UV-Vis spectra of Schiff bases have been described by many authors and particularly by El-Bayoumi et al. and Belletête et al. [22,23].This type of compounds have 4 bands (I to IV) which the first and the second one (I and II) correspond to n→π * transition and appear at wavelengths different from those of aldehydes.These four bands (I to IV) appear clearly in the UV-Vis spectrum of the monomer N-(m,p)-vinylbenzylidene-p-anisidine in ethanol and remain unchanged in the initial time (t1) of Im hydrolysis in the hydro-ethanolic study medium as shown in Figure 1a.Full UV spectra are registered for slow kinetics.An example is given in Figure 1a for Im hydrolysis at pH = 6.18; the UV spectra are registered at different times during the hydrolysis process.The comparison of UV spectra of Im (Figure 1a) and p-aniside (Figure 1c) and m,pvinylbenzaldehyde (Figure 1b) in same conditions revealed that the band I which corresponds to n→π * transition of imine function is the most appropriate to keep track the hydrolysis kinetic.It is observed that its intensity diminishes during the hydrolysis process indicating the imine function destruction.
Figure 2 showed the UV spectra of Cp2 hydrolysis at pH = 6.18, the first spectrum (n°1) which is registered at time ˂ 30 s displayed also four bands at 334, 266, 240 and 212 nm attributed to an aromatic Schiff base, that is the copolymer Cp2 before hydrolysis.In this case, also the band (I) is chosen for the imine constant hydrolysis determination.In the last spectrum (n°10), it is observed that the band (I) disappeared indicating completion of hydrolysis and obtaining the two spectra of amine and aldehyde (Scheme 3).
The presence of isobestic points "i1" and "i2" as mentioned in Figure 1a and 2 indicates that there was no accumulation of any intermediate, especially the α-amino alcohol, during the hydrolysis process both of imine (Im) and copolymers (Cp1) and (Cp2) (Scheme 3).For pH values less than 5, the hydrolysis process is more rapid; in this case, another method is used to track the hydrolysis kinetic.In fact, a quantitative method of the UV apparatus is used, the analytical wavelength is set at 350 nm (band I), and the optical density (D) is displayed as a function of time as shown in Figure 3 for Im hydrolysis in pH medium equal to 4.65.If the hydrolysis kinetic is considered as a first-order reaction, it obeyed to the equation 2: where and are corrected optical density at time t and initial optical density both are measured at the wavelength λ, is a hydrolysis constant.The corrected optical density is equal to actual optical density minus the measured optical density at infinite time.
On one hand, the experimental data are plotted and linear fits of log ( ) = f(t) for imine (Im) and copolymers (Cp1 and Cp2) at pH 4.16, 6.18, 7.71 and 9.91 are obtained.Examples of fits are given in Figure 4. Consequently, the hydrolysis kinetics of these Schiff bases in ethanol buffer mixtures are considered as first-order reactions.On the other hand, the initial optical densities read at λ = 350 nm from the linear plots of log ( ) = f (t) checked the Lambert-Beer law, as noticed in Table 2, at different concentrations of imine (Im), experimental hydrolysis constant (kobs), half reaction time (t1/2) and initial optical density (D 0 ) are measured and a linear plot of = f(C) is obtained as shown in Figure 5.The first order is also confirmed by the quasi-constancy of t1/2 whatever the initial concentrationC0 of the free imine (Im) or copolymer carriers (Cp).Then, experimental kinetic constants (kobs) of all compounds and at four values of pH are calculated from the slope of the log ( ) = f(t) curve and the initial optical density D 0 is determined by extrapolation at t = 0.The corresponding results are given in Table 3.It is noted that by varying the pH medium, D 0 is almost constant for each compound; in fact, the mean value of D 0 are respectively 1.615 for Im, 0.155 for Cp1 (β 0 = 35%) and 0.235 for Cp2 (β 0 = 48%).It is coherent that D 0 at λ = 350 nm of copolymers should be lower than D 0 of Im since the concentration of imine group in copolymers depends on the degree of incorporation (β 0 ).In fact, at initial time, the imine concentration in copolymer is significantly lower than pure imine.Also, by comparing Cp1 and Cp2, the value of D 0 is in agreement with copolymers' imine content, so higher is the incorporation rate (β 0 ), higher is the initial optical density (D 0 ).The results showed obviously that at same pH value, the hydrolysis constants of copolymers (Cp1, Cp2) are lower than of the monomer (Im).It is noticed that the carrier, i.e. copolymer backbone affects significantly the hydrolysis constant.As a result, copolymers and especially Cp2 exhibited delayed p-anisidine release since the hydrolysis constant is decreased.
During the hydrolysis process, the evolution of the UV spectrum of copolymers facilitates the clarification of the hydrolysis reaction that takes place according to the reaction in Scheme 4. Once the total reaction has been achieved, a mole of copolymer (Cp) gives an equal amount of aldehyde (VBA) based copolymer and β 0 mole of p-anisidine.Scheme 4. Complete hydrolysis reaction of copolymers.
During the hydrolysis process, at time t, the incorporation ratios α 0 of DMA remains constant and β 0 is equal to (β' + β") where β' is units of remaining Schiff basis and β" is the units obtained aldehyde as shown in Scheme 5: Scheme 5. Partial hydrolysis of copolymers.
The chemical modification of the copolymer during hydrolysis is related to the partial hydrolysis of the co-monomer carrying the imine function.

A comparative study of kinetic hydrolysis of monomer (Im) and copolymers (Cp1) and (Cp2)
The formation or hydrolysis reaction of imine is quite sensitive to pH.The value of the apparent kinetic constants of Schiff bases of imine and copolymers at each pH are regrouped in Table 3.For a better comparability, the ratios k(Im)/k(Cp1) and k(Im)/k(Cp2) are calculated and given in Table 3 and the pH-rate diagram profile, i.e. log kobs = f (pH) are drawn, as shown in Figure 6.Firstly, the results revealed that the mono-imine (Im) is rapidly hydrolyzed comparing to copolymers.In fact, the ratios k(Im)/k(Cp) are always greater than unity as remarked in Table 3 and depends on the pH medium.In fact, the ratio kIm/kCp1 varied from 1.02 to 1.74 and is a higher at pH = 4.68 and however the ratio kIm/kCp2 is more valuable and varied from 2.02 to 2.77 and is a higher in basic medium at pH = 9.91.
It is noted that the observed hydrolysis constant decreased when the pH increased until pH = 7.71 for the three compounds (Table 3).It is reported that the hydrolysis curve of Schiff bases derived from aromatic and aliphatic amines can be divided into different zones [13].In Figure 6, two zones of hydrolysis called A and P appeared that are meaning acidic zone catalysis and basic plateau, respectively.Between 4.65 and 7.71 of pH value corresponding to a zone A, the hydrolysis reaction is catalyzed by acidity since the observed hydrolysis constant decreases noticeably when pH increases.In these conditions, it is considered that the protonated imine is attacked by water molecules but the values of the slops of log kobs = f (pH) is lower than unity indicating that the base catalysis cannot be neglected [13].A same shape of curves log kobs = f (pH) (Figure 6) is obtained both for the free imine and copolymers that confirmed that the hydrolysis mechanism remains unchanged whatever the imine carrier.Similar results have been noticed for other Schiff bases [24][25].
Also, as reported in previous research [8], the hydrolysis curves of Schiff bases derived from pyrrolic aldehydes gave similar zones from pH = 4 to 12.However, as reported by Barmatov and Hugues [7], for the Schiff base N-(4-(dimethylamino)benzylidene) benzothiazol-2-amine 3; a specific acid catalysis mechanism is observed from pH = 2.76 to 7.51.For Im, Cp1 and Cp2 and from pH 7.71 and 9.91 appeared a zone P of basic plateau of the pHrate curve (Figure 6), in this zone, it is demonstrated that the rate-limiting step corresponded to the nucleophile attack of hydroxide anion on the protonated imine and the kinetic constant is independent from pH [13,25].
In the two zones of pH, the results showed that the hydrolysis of free imine (Im) is more rapid than copolymers Cp1 and Cp2.So, the delayed effect due the copolymerization is distinguished.Besides, if we compare the two copolymers, it is seen that the hydrolysis constant kobs of Cp1 is higher than kobs of Cp2 whatever the pH value.From the copolymers' characterization, Cp2 contained more Schiff bases or imine groups (β 0 = 48%) and had a low molecular mass (Mn).So, it seems that there is no correlation between the molecular weight Mn and the kinetic constants but rather a specific effect of the close statistical pelota of polymer is occurred.In fact, in Cp2, the amide:imine ratio is close to 1:1, intra-molecular and intermolecular interactions can be arise and so the hydrolysis reaction is inhibited and the constant kobs decreased.
The similar hydrolysis curves are obtained for both the free imine and its copolymers (amideimine), so the hydrolysis mechanism of the imine function remained virtually unchanged regardless of its backbone carrier.As well, same hydrolysis curve profiles of aromatic Schiff base are obtained by other researchers [13,24,[26][27].Finally, it is noticed that the hydrolysis constants of these aromatic Schiff bases (Im and Cp) are higher than those of p-anisidine Schiff bases derived from N-pyrrolic and N-methyl pyrrolic aldehydes [13].

CONCLUSION
The present research discusses the p-anisidine controlled release by hydrolysis process of Schiff bases derived from aromatic backbone.The hydrolysis of free imine or N-(m,p)-vinylbenzylidenep-anisidine (Im) and its copolymers with N,N-dimethylacrylamide (Cp1) and (Cp2) is studied in pH range of 4 to 10 and using UV-Vis analysis.The experimental results showed that the mechanism of hydrolysis reaction is a pseudo first order.The experimental kinetic constants are calculated and the morphology of the hydrolysis curves log kobs = f(pH) confirmed that the hydrolysis mechanism did not undergo any modification when moving from the free mono-imine molecule (Im) to the polymer carrier; in this range of pH, two zones namely A and P are obtained.Whatever the pH, the hydrolysis kinetic constant (kobs) of mono-imine is higher than those of copolymers.However, for copolymers, it is observed that the kobs decreased when the degree of imine incorporation increased.Finally, the copolymer Cp2 displayed a slow hydrolysis and a delayed p-anisidine release.

Scheme 1 .
Scheme 1. Global reaction of Schiff base synthesis.

Figure 2 .
Figure 2. UV spectra of Cp2 at different time of hydrolysis at pH = 6.18.

Figure 4 .
Figure 4. Plots of log ( ) as a function of time for monomer and copolymer Cp2 hydrolysis at different pH values.

Figure 5 .
Figure 5.A linear plot of log (D 0 350) = f (C) of Im for Lambert-Beer law verification.

Table 1 .
Characterization results of copolymers.

Table 2 .
Confirmation of the first order by checking Lambert-Beer law for imine Im.