Synthesis and characterization of Co(II) porphyrin complex supported on chitosan/graphene oxide nanocomposite for efficient green oxidation and removal of Acid Orange 7 dye

Catalytic degradation of Acid Orange 7 (AO7) by hydrogen peroxide in an aqueous solution has been investigated using cobalt(II) complex of 5, 10, 15, 20 Tetrakis [4-(hydroxy)phenyl] porphyrin [Co(II) TPHPP] covalently supported chitosan/Graphene Oxide nanocomposite [Co(II) TPHPP]-Cs/GO, as highly efficient and recoverable heterogeneous catalyst. The structures and properties of [Co(II) TPHPP]-Cs/GO nanocomposite were characterized by techniques such as UV–Vis, FT-IR, SEM, EDX, TEM, and XRD. The oxidation reaction was followed by recording the UV–Vis spectra of the reaction mixture with time at λmax = 485 nm. [Co(II) TPHPP]-Cs/GO nanocomposite demonstrated high catalytic activity and could decompose 94% of AO7 within 60 min. The factors that may influence the oxidation of Acid Orange 7, such as the effect of reaction temperature, pH, concentration of catalyst, Acid Orange 7, and hydrogen peroxide, have been studied. The results of total organic carbon analysis (TOC) showed 50% of dye mineralization under mild reaction conditions of AO7 (1.42 × 10−4M) with H2O2 (8 × 10−2M) in the presence of [Co(II) TPHPP]-Cs/GO nanocomposite (15 × 10−3 g/ml) and pH = 9 at 40 °C. The reuse and stability of the nanocomposite were examined and remarkably, even after six cycles of reuse, there was no significant degradation or deactivation of the recycled catalyst. Residual organic compounds in the reaction mixture were identified by using GC–MS analyses. The radical scavenging measurements and photoluminescence probing technology of disodium salt of terephthalic acid indicated the formation of the hydroxyl radical as the reactive oxygen species in the [Co(II) TPHPP]-Cs/GO nanocomposite/H2O2 system. A mechanism for the oxidation reaction has been discussed.


Instrumental measurements
On a Bruker Avance II spectrometer operating at 400 MHZ, 1HNMR spectra was measured in the presence of CDCl 3 and the chemical shifts were provided.The UV-visible spectra were measured by using a UV-1800 UV-visible scanning spectrophotometer (SHIMADZU, Kyoto, Japan) in the range of 200-600 nm.FTIR was performed using (JASCO FT-IR-4100, Japan) in the range 400-4000 cm −1 utilizing liquid samples of semi-solid polymers or KBr pellets for powdered polymers with chloroform as the solvent.Analyzing the crystallography structure was done using an X-ray powder diffractometer (APD 2000 pro-Italy).The experiment utilized Cu-K radiation, which has a wavelength of 1.5406 Å.The angle range for the scan was set from 5 to 90, with a scanning rate of 0.05/s at 45 kV and 0.8 mA.The pH of the medium was adjusted using a pH bench meter (AD1030, Adwa, Hungary).Scanning electron microscope (SEM) and Energy-dispersive X-ray spectroscopy (EDX) analysis at 10 kV to detect the existence of elements within the nanocomposite using (JSM-IT200 In Touch Scope™ Scanning Electron Microscope).A transmission electron microscope (TEM) JEM-1400Plus Electron Microscope was used to examine the sample.After undergoing ultrasonic treatment, a small amount of the sample was delicately placed onto a copper grid.The solution was allowed to air dry at room temperature, and any excess liquid was removed by gently blotting it with a delicate piece of cloth.Filter paper by Whatman.GC-MASS analysis was performed using (Thermo Scientific ISQ single quadrupole gas chromatography-mass spectrometry (GC-MS) instrument).

Grafting chloro-acetyl chloride on N-phthaloyl chitosan
Chloro-acetylation of N-phthaloyl chitosan was obtained according to the reported method 47 .IR (ν, cm

Preparation of N-phthaloyl chitosan supported of [Co(II) TPHPP]
Grafting of [Co(II) TPHPP] complex on N-phthaloyl chitosan was done as a reported method 48 , with some modification briefly (0.01 mmol) cobalt porphyrin complex in DMF (30 mL) was added to a solution of N-phthaloyl chitosan (1.0 g) and anhydrous potassium carbonate (0.5 g).

Preparation of [Co(II) TPHPP] supported on to modified chitosan/graphene oxide nanocomposite
Graphene Oxide was synthesized via the modified Hummer's method 50 .Then, [Co(II) TPHPP]-Cs/GO nanocomposite [(4:1) ratio of [Co(II) TPHPP]-CS:GO] was synthesized via a facile two-step method according to the reported method 51 .IR(ν, cm www.nature.com/scientificreports/were taken out of the reaction flask and subjected to analysis.The UV of AO7 was calculated at wavelengths of 485 nm.The aliquots were then added back to the reaction flask.Pseudo-first order relation was applied to fit the results from destruction results: where A o is the dye's initial absorbance (at t = 0 min), A t is that absorbance at time = t, and K obs (min −1 ) is the observed rate constant calculated from the slope of the linear plot of ln (A o /A t ) vs time.

Recovery and recycling of catalyst
[Co(II) TPHPP]-Cs/GO nanocomposite were recycled.After the completeness of the reaction, the supported catalyst was removed from the reaction media by simple filtration, and they were reused for subsequent experiments after being washed with distilled water.Then directly used in the next degradation reaction 52 .Successive catalytic degradation experiments under optimum conditions were achieved to investigate the recycling ability of the catalyst for at least 6 cycles.For these tests, the same sample was utilized.The following parameters were set for the experiment: 10 mL solution volume; initial 1.42 × 10 −4 M dye concentration, 15 × 10 −3 g/mL of catalyst dosage, 8 × 10 −2 M H 2 O 2 concentration at 40 °C and reaction time: 60 min.The degradation efficiency was investigated according to: .Simultaneously the amino group of biopolymer chitosan was protected by using phthalic anhydride forming N-Phthaloyl chitosan 46 , which chloroacetylated using chloroacetyl chloride in the presence of triethyl amine forming Chloroacetylated N-phthaloyl chitosan 46 .[Co(II) TPHPP] covalently bonded to (chlorocetylated N-phthaloyl chitosan) by refluxing in the presence of potassium carbonate 48 .Then, the protected (phthaloyl group) was removed using Hydrazine monohydrate 49 .Finally, Chitosan/[Co(II) TPHPP] mixed, vigorously stirring with Graphene Oxide suspended solution to form [Co(II) TPHPP]-Cs/GO nanocomposite 51 as shown in (Supplementary Figs.S1-S7) Fig. 2.

FT-IR analysis
The FTIR spectrum of THPP is consistent with its chemical structure as shown in (Fig. 3a).The stretching vibrations of O−H and N−H groups showed a broad band at ν 3423 cm −1 .Additionally, the peaks at ν 1228 and 1169 cm −1 were attributed to C−N stretching vibrations of amine groups, while peaks at ν 1599 and 1469 cm −1 were assigned to N-H bending and C=N vibrations, respectively.Finally, the peak at ν 808 cm −1 was referred to as the macrocycle ring's N-H out-of-plane bending vibration.In the [Co(II) THPP] spectrum (Fig. 3b), the peak at ν 3423 cm −1 became broad and slightly shifted to 3420 cm −1 .The new peak at ν 1001 cm −1 has appeared, which is characteristic absorption of the Co-N(equatorial) bond formed in [Co(II) THPP] 24,53 .The spectrum of Cs (Fig. 3c) shows a broad band at ν 3424 cm −1 , which is the region where the stretching vibration of O−H and N−H groups are situated due to intermolecular H-bond.The absorption bands at ν 1634 cm −1 and 1389 cm −1 correspond to the C=O stretching vibration (amide band) and C−N stretching vibration in amide.And band at ν 1069 cm −1 corresponds to C−O stretching vibration.Unlike chitosan, the FT IR spectrum of N-phthaloyl chitosan (Fig. 3d) exhibits characteristic peaks at ν 1715, 1759 corresponding to C=O of the phthalimide group.And another peak at ν 1555 and 724cm −1 for C=C stretching vibration and C−H bending vibration of an aromatic ring 54 .
For the chloroacetylated of N-phthaloyl chitosan (Fig. 3e), a new signal around ν 2958 cm −1 (methylene of the chloroacetyl group) was found to correlate with the decrease of the signal around ν 3424 cm −1 (hydroxyl groups).Also, a new peak at ν 717 cm −1 confirmed the presence of C-Cl in the chloroacetyl group [55][56][57] .
For [Co(II) TPHPP] supported on N-Phthaloyl chitosan spectrum (Fig. 3f), the broad peak around at ν 3486 cm −1 was slightly shifted to ν 3432 cm −1 , and the intensity was diminished which related to vibration of O−H and N−H groups.Also, the band at ν 1759 and 1715 cm −1 was shifted at ν 1720 and 1661 cm −1 , and the significant peak at ν 717 cm −1 which is related to C-Cl in the chloroacetyl group disappeared due to attaching [Co(II) THPP] 1 to N-phthaloyl chitosan.This proves that the [Co(II) THPP] 1 molecules were covalently bonded to N-phthaloyl chitosan.As seen, signature bands for [Co(II) THPP] did not appear anywhere in the spectrum of [Co(II) THPP]-N-phthaloyl chitosan due to the overlap of the spectra of N-phthaloyl chitosan and [Co(II) THPP] in the entire infrared region 58 .
FT-IR spectrum for the protected group (phthaloyl group) removed (Fig. 3g), chitosan/Co-porphyrin show disappearing characteristic peaks at ν 1720,1661,717 cm −1 correspond to C=O of phthalimide group and C−H bending vibration of aromatic ring.Another peak appeared at ν 1656 cm −1 this evidence removed the protected group (phthalic anhydride) 59 .
The spectrum of GO shown in (Fig. 3h), exhibited a broad band at 3442 cm −1 for O-H groups, and two characteristic bands for C=O and C=C at 1720, and 1627 cm −1 , respectively.Furthermore, the other peaks corresponding to C-OH and C-O stretching vibration appeared at 1387 and 1074 cm −1 , respectively.These characteristic peaks indicated that the GO nanosheets were successfully synthesized 60 .

XRD structural characterization
The X-ray powder diffraction pattern was illustrated in Fig. 4 where (Fig. 4a) the original chitosan shows a weak peak at 2θ = 9.60° and a more intense peak at 2θ = 20.22° that is caused by diffraction from the (020) and (110) planes of the crystalline lattice with interplane distances of 0.92 nm and 0.438 nm, respectively.It is worth noting that the corresponding degree of acetylation was 81% and its crystalline index was 55 [62][63][64][65] .
N-phthaloyl chitosan (Fig. 4b) showed the disappearance of the peak at 2θ = 9.6° with the appearance of another peak at 6.42°, and the peak at 2θ = 20.22° was shifted to 2θ = 18.62°.This explains why the amount of hydrogen bonds decreased by blocking amino groups, leading to a typical diffraction pattern with lower crystallinity.It was reported that N-phthaloyl chitosan prepared in DMF/water showed a certain crystallinity, despite the introduction of the phthaloyl group; this also supports the uniform structure of the product.It is noteworthy that N-phthaloyl-chitosan exhibited certain crystallinity even though such a bulky substituent had been introduced 64 .
Figure 4d shows that the deprotection of the amino group of chitosan (phthaloyl group) shows a characteristic peak at 2θ = 20.81°,which indicates a higher crystallinity of this compound.Figure 4e show a sharp peak at 2θ = 10.32°, which confirms that graphite powder was oxidized well, utilizing concentrated acids and KMnO 4 to obtain GO.After modifying GO with modified [Co(II) TPHPP]-Cs, the significant peak of GO at 2θ = 10.32°shifted to a smaller angle and appeared at 2θ = 9.60° as shown in (Fig. 4f).This shift is due to the intercalation of GO by modified metalloporphyrin-chitosan chains.The two peaks of Cs are not observed in the XRD patterns of GO-Cs nanocomposite, indicating that [Co(II) TPHPP]-Cs chains are well intercalated amongst the GO sheets.This confirms the good attachment of [Co(II) TPHPP]-Cs to GO layers 67,68 .SEM and EDX technique.SEM micrograph of the [Co(II) TPHPP] supported on chitosan before and after GO incorporation, where the uniform distribution of [Co(II) TPHPP]-chitosan network is displayed in (Fig. 5a).
The typical morphology of GO is shown in (Fig. 5b), which has a smooth, flat, soft surface and many layers stacked on top of one another in a range of sizes and forms.Furthermore, as seen in (Fig. 5c,d), the irregularly shaped GO layers and their surface developed wrinkles as a result of being occupied by [Co(II) TPHPP]-Cs beads, where the [Co(II) TPHPP]-Cs beads bind the bunches of flake GO sheets, these bound transpire on the surface and from edges confirm the tightly bound with each other and illustrate the successful interaction between [Co(II) TPHPP]-Cs and the GO.Additionally, similar results were supported using the EDX analysis, which shows the presence of elements such as C, O, and N, Co, which indicate the combination between them forming [Co(II) TPHPP]-Cs/G'O nanocomposite in (Fig. 5e).
TEM analysis.The TEM imaged in (Fig. 6a,b), shows the nanoparticles morphologies of [Co(II) TPHPP]modified Chitosan are mostly spherical.Some of these beads spread and others aggregated within Graphene Oxide layers.The TEM image shows that the changes to the inner layers of GO are dramatic and clearly visible as some layers intercalated and others exfoliated.Where the bulk of GO sheets contain beads with wrinkles due to the dispersion of [Co(II) TPHPP]-chitosan networks on the GO sheets.Figure 6c shows the histogram of the particle size distribution curve of [Co(II) TPHPP]-Cs/GO nanocomposite where the irregular spherical shape of these beads and its average particle size (< 30 nm).

Catalytic oxidation of AO7
The catalytic oxidation of (AO7) has been investigated using hydrogen peroxide as an oxidant and [Co(II) TPHPP]-Cs/GO nanocomposite as a catalyst in aqueous solution.The oxidation reaction was followed by recording the UV-Vis spectra of the reaction mixture with time at λ max = 485 nm.(Fig. 7a) represents the collapse of the main absorbance band of AO7 at λ max = 485 nm almost vanished, and the degradation percent of AO7 reached 94% within 60 min, this is due to the lost conjugation in the dye leading to colorless oxidation products.This reverberates an essential role of using H 2 O 2 and [Co(II) TPHPP]-Cs/GO nanocomposite in the degradation of AO7. Figure 7b also shows the kinetic curve for the destruction of AO7, the plot of (lnA 0 /A t ) against time presents a straight line, this indicates that the degradation of AO7 is considered a first-order rate kinetic 69,70 and it could be simply described as ln A 0 /A t = k obs t.Before studying the behavior of this supported catalyst, it is important to evaluate the AO7 decolorization process, i.e., whether AO7 removal occurs through adsorption, through a catalytic reaction, or both processes.For that reason, several runs were then performed.The first one was a blank, carried out to evaluate the ability of H 2 O 2 to eliminate AO7 in aqueous solutions without the addition of the heterogeneous catalyst which shows that AO7 degradation due to hydrogen peroxide is almost negligible (Supplementary Fig. S8) which might be attributed to its low oxidation potential as compared to hydroxyl radicals.
To determine the influence of the adsorption processes experiment without, H 2 O 2 and in the presence of [Co(II) TPHPP]-Cs/GO nanocomposite was carried out as shown in (Supplementary Fig. S9) and also carried out at different pH used (7,9 and 11) (Supplementary Fig. S10), which indicated no significant decolorization of dye, this evidence for the degradation pathway occurred through a catalytic reaction.

Influence of the experimental conditions on oxidation of AO7
The factors that may influence the oxidation of AO7, such as the Influence of reaction pH, AO7, concentration of catalyst, hydrogen peroxide, and temperature have been investigated.

Influence of initial PH
The influence of pH on the oxidation reaction was investigated at a constant concentration of the dye (1.42 × 10 -4 M), H 2 O 2 (8 × 10 −2 M) as well as a fixed amount of [Co(II) TPHPP]-Cs/GO nanocomposite (15 × 10 -3 g/mL) at 40 °C.The pH varied in the range of 7-11.Data illustrated in (Fig. 8) show that the decolorization of AO7 increased with an increase of pH and reached optimum at pH = 9.0 71,72 .The factors likely responsible for the decrease in the observed rate of constant k obs at higher pH are the formation of hydroperoxide anion HO − 2 in an alkaline medium, which reacts with the non-dissociated molecule of H 2 O 2 according to reaction Eq. ( 3).

Influence of AO7 concentration
The variation of the decolorization efficiency of the AO7 with H 2 O 2 and [Co(II) TPHPP]-Cs / GO nanocomposite with varying the concentration of the dye from 6.65 × 10 −5 M to 2.85 × 10 −4 M. (Fig. 9) shows that the decolorization efficiency reduces with a further increase in dye concentration.This phenomenon may be attributed to the high concentration of AO7 molecules that may aggregate on the catalyst surface and inhibit contact between H 2 O 2 and the catalyst, which reduces the number of hydroxyl radicals involved in the decolorizing process 72 .The molar ratio oxidant/dye is low (because the amount of hydrogen peroxide molecules initially present in the reaction is the same). (3)

Influence of catalyst concentration
Data illustrated in (Fig. 10) show that the rate of oxidation of AO7 increases with an increase of the catalyst concentration from 4 × 10 −3 g/mL to 15 × 10 −3 g/mL with all other reaction parameters fixed, at 40 °C.This may be attributed to the increase in the number of available active sites on the catalyst surface for H 2 O 2 activation and the increased rate of formation of hydroxyl radicals 73 .After 15 × 10 −3 g/mL concentration, adding more catalyst may not have a significant effect on the rate of reaction.This point is called steady-state concentration where no further increase in the rate of reaction, due to the limited number of attacking active species involved in the degradation process, k obs remained almost constant.Thus, the optimum catalyst dosage 74 .4) and ( 5) compete with the destruction of the dye chromophore 72 .These reactions reduced the probability of attacking the AO7 molecules by hydroxyl radicals, which decreased the decolorization rate of the dye at a high concentration of H 2 O 2 .

Influence of reaction temperature
The effect of temperature on the decolorization of AO7 with [Co(II)-TPHPP]-Cs/GO nanocomposite /H 2 O 2 has been studied in the range of 25-55 °C.(Fig. 12) shows that the decolorization rate increased with an increase in reaction temperature.Increasing temperature led to a shorter time for decolorization of AO7.Thermodynamic parameters for degradation AO7 such as Gibbs free energy (ΔG°), enthalpy (ΔH°), and entropy (ΔS°) describe the nature and type, and disorder of the system at the liquid-solid interface 75 .
Data illustrated in Table 1 where: a positive ∆G° value validates a non-spontaneous process under standard conditions, which means that the reaction still prefers reactants over products under the given conditions.Overall, a positive ΔG with a catalyst doesn't necessarily mean an ineffective reaction.It just means the reaction leans toward the reactants, and the catalyst helps achieve equilibrium faster 76,77 .A positive ∆H° value is indicative of an endothermic, irreversible process, and the negative value of ΔS° revealed that the degree of the disorder decreased at the solid-liquid interface during the reaction and the positive E a is an indication the system requires energy (increased temperature) to drive the process, therefore is termed endothermic 78 .

Mineralization and proposed degradation pathway of AO7
Total organic carbon analysis (TOC) is an important method for evaluating the mineralization of the oxidative degradation reactions of dyes.Oxidation of AO7 under standard reaction conditions at 40 °C showed after 60 min TOC removal of 50% indicating incomplete mineralization of the dye to CO 2 and H 2 O.However, raising the reaction temperature to 55 °C mineralization of the dye increased and the mineralization of AO7 also enhanced with a higher concentration of catalyst up to 15 × 10 -3 g/mL [79][80][81][82] .

Comparison with other systems
Table 3 compared the results obtained in terms of catalytic degradation efficiency, stability, reaction time, temperature, pH, and dye mineralization of AO7 in the presence of the [Co(II) TPHPP]-Cs/GO nanocomposite presented in this work, to other previously published results.It can be seen from Table 3.A heterogeneous catalyst used [Co(II) TPHPP]-Cs/GO nanocomposite gave higher degradation yield and mineralization of AO7 with a shorter reaction time required and also showed high catalytic stability and no significant changes up to the sixrun compared to the other heterogeneous catalysts used in the comparison.

Identification of reactive oxygen species (ROS)
Hydroxyl radical analysis using isopropyl alcohol Hydroxyl radicals were able to be the reactive species in the oxidation of the dye AO7 by our catalytic system [Co(II) TPHPP]-Cs/GO nanocomposite/H 2 O 2 , hence, the inhibiting effect of isopropyl alcohol as hydroxyl radicals scavenging agent has been investigated on the oxidation reaction of AO7 by H 2 O 2 /catalyst.As shown in (Fig. 13), the rate of degradation of AO7 decreased with the addition of isopropyl alcohol to the reaction mixture and was inhibited by increasing the concentration of isopropyl alcohol in the reaction solution.This indicates that decolorization of AO7 by [Co(II) TPHPP]-Cs/GO nanocomposite/H 2 O 2 involved the formation and participation of • OH radicals as the active species.

Hydroxyl radical determination
To identify the reactive oxygen species formed in the [Co(II) TPHPP]-Cs/GO nanocomposite/H 2 O 2 system, disodium salt of terephthalic acid (NaTA) photo-luminescence probing technology measurements were carried out.NaTA could react with • OH to give 2-hydroxy terephthalic acid (HTA), which exhibits a bright stable fluorescence 91 .This reaction is unaffected by the presence of other reactive species such as H 2 O 2 , HO  It's observable that the fluorescence intensity increases sharply to 220 within 75 min, implying that • OH radicals were indeed generated in the system.Moreover, when AO7 and NaTA were simultaneously added into the solution, the generated fluorescence significantly decreased, with the intensity only increasing to 80 simultaneously.

Suggested mechanism
The suggested mechanism for the oxidation reaction involves an activation of the H 2 O 2 molecule, leading to the formation of hydroxyl radicals( • OH). as shown in the following equations.where S is the supported nanocomposite, [TPHPP] is the ligand, and Co is the metal ions.Cobalt can be used as a catalyst, preferably in a heterogeneous system.Cobalt is known to undergo Fenton-type reaction and has been used as an activator in the decomposition of H 2 O 2 and the degradation of various dyes 72 .The suggested mechanism claims that the catalyst activates the H 2 O 2 molecules, leading to the formation of hydroxyl radicals, • OH 93 .The latter attacks the dye forming an active intermediate, which decomposes in the rate-determining step giving the final oxidation product.

Recovery and recycling of catalyst
There is a wide diversity of benefits resulting from using heterogeneous catalysts in the catalytic oxidation process as reducing reaction costs, diminishing waste generation, and bringing about more environmentally and economically saving methods for separation and recycling 24,94 .[Co(II) TPHPP]-Cs/GO nanocomposite was  www.nature.com/scientificreports/readily recovered from the solution by simple filtration and recycled for successive reactions after being rained several times with distilled water.Degradation percentages of AO7 were illustrated (Fig. 15).The proportion of AO7 degradation after six consecutive cycles was shown in Fig. 15.No significant changes were detected for the first five successive cycles of [Co(II) TPHPP]-Cs/GO nanocomposite, then the foregoing catalyst losses (15%) of its activity in the sixth cycle.The results clearly showed that the mentioned supported catalyst was active in oxidative degradation and could be stable and reused without a significant decrease in activity and selectivity as shown also in FT-IR (Supplementary Fig. S11).The breakdown rate and mineralization of the dye have been found to increase with an increase in reaction temperature and catalyst concentration up to 15 × 10 −3 g/mL subsequently no significant increase.The rate of dye decolorization decreased with increasing the concentration of dye, H 2 O 2, and at higher pH than 9.0.GC-MS analyses examined all the degradation products of AO7 with different retention times.Remarkably, even after six cycles of reuse, there was no significant degradation in the catalytic activity of the recycled catalyst.This breakthrough highlights the potential of the catalyst in addressing water pollution challenges efficiently and sustainably.

Figure 8 .
Figure 8.Effect of pH on the decolorization of AO7.The pH was adjusted to 8 and 9 using borax and HCl buffer mixture and the pH was adjusted to 10.0 using NaHCO 3 and NaOH mixture.Phosphates were used to adjust the pH to 7 and 11.

Figure 11 .
Figure 11. .Effect of concentration of H 2 O 2 on the decolorization of AO7.

Figure 13 .
Figure 13.Effect of increased isopropyl alcohol on the decolorization of AO7.

Table 1 .
Effect of temperature on the decolorization of AO7.Rate constant and activation parameters of catalytic degradation of AO7 by [Co(II) TPHPP] -Cs / GO nanocomposite with H 2 O 2 .

Table 2 .
Identified reaction intermediates degradation of AO7 by GC-MS.