Highly Efficacious Synthesis of Spiro-Fused Oxindole-Dioxolo[g][1]benzopyran Derivatives by Mechano-Chemical Approach using Ti3C2OH MXene 2D Nanomaterial

INTRODUCTION
The use of multi-component reactions (MCRs) as adaptable reactions to produce a wide variety of complex compounds with frequently biologically relevant scaffold structures using novel, efficient methods has become recognized as an intriguing and eco-friendly strategy [1]. According to published research, the mechano-chemistry method has created a chance to create novel processes for the quick synthesis of organic compounds since it allows for solvent-free reactions or the use of activators, which makes the process effective and affordable [2].
Among the various number of natural and unnatural bio-active spirocycles discovered heretofore, the spirooxindole moiety is a clinically significant drug used in the treatment and management of many diseases [3, 4]. Strong biological activities, such as antioxidant [5], antibacterial [6], antiviral [7], anticancer [8, 9], and antimalarial effects [10], are displayed by compounds with the spirooxindole structure. The importance of the spirooxindole framework in drug development is shown by the fact that it is present in many naturally occurring medicinally useful compounds. Typical examples with anti-inflammatory, anti-malarial, antimicrobial, and antitumor properties include mitraphiline [11], KAE609 [12], elacomine [13], and spirotryprostatin A [14] (Fig. 1). 
The MAX phases encompass a diverse category of stratified materials, consisting of hexagonal ternary early transition metal nitrides, carbides, and carbonitrides. This material demonstrated advantageous factors such as excellent thermal and electrical conductivity, ease of machinability, and resistance to oxidation and thermal shocks. They coined the term “Mn+1AXn” phase (where n = 1, 2, or 3), or “MAX” phase, to refer to these materials. In this material, M is a transition element, A is an element from the A group (group IIIA or IVA), and “X” represents carbon and/or nitrogen [15]. The MAX phases demonstrate chemical stability, although the A layers exhibit higher chemical reactivity due to the relatively weak M-A bonds between the Mn+1Xn layers. Hence, the Ti element can be reacted with Al and C to produce Ti3AlC2 or Ti2AlC MAX phases. Next, during the etching and delaminating procedures, the alumina is removed through centrifugation. The resulting colloidal suspension includes new materials denoted as MXene [16].
According to a review of the literature, one of the most important and desirable methods for chemists to promote environmentally friendly and sustainable chemical processes is the use of nanostructured materials as potential heterogeneous catalysts [17]. Since its discovery in 2011, MXenes, a form of two-dimensional material, have attracted a lot of attention due to their efficacy in certain situations [18]. These layered materials on a two-dimensional plane have sheet topologies similar to graphene, as indicated by the suffix “ene” in MXenes [18]. The usual formula for MXene is Mn+1XnTx, where n can be any number between 1 and 3. T stands for surface terminal functions, such as OH, O, F, and Cl, and M can be a transition metal atom, such as Zr, V, Ti, or Sc, positioned between n layers of either nitrogen or carbon (referred to as X) [19].
Here, we report for the first time on the catalytic use of Ti3C2OH MXene in the synthesis of a series of spiro-fused oxindole-dioxolo[g][1]benzopyran derivatives (4), considering our expertise in designing and developing new catalyzed synthetic methods for the preparation of significant heterocyclic frameworks [20-23]. The title compounds were produced by employing Ti3C2OH MXene as a novel, superior catalyst in a three-component reaction involving 3,4-methylenedioxyphenol (1), malononitrile (2), and 5-substituted isatins (3) utilizing a mechanochemical ethanol-assisted grinding technique (Fig. 2). Despite our previous publication on a practical approach, this work provides a recoverable catalyst/grinding-aid system and is strongly compatible with the concepts of green chemistry [24].

MATERIALS AND METHODS
Chemical reagents and solvents were purchased from Merck and Fluka Companies and utilized without additional purification. IR spectra were recorded on an ABB FT-IR (FTLA 2000) spectrometer. 1H NMR and 13C NMR spectra were recorded on a Bruker DRX-500 AVANCE at 500 and 125 MHz, respectively, in DMSO-d6 as the solvent and TMS as the internal standard. Elemental analyses were performed on a Foss-Heraeus CHN-O-rapid analyzer. X-ray diffraction (XRD) was recorded on a Philips PW1800/00 X-ray powder diffractometer employing CuKα X-ray radiation (0.154056 nm). Scanning electron microscopy (SEM) was captured with a TESCAN Vega TS 5136LM coupled to an energy-dispersive X-ray (EDS) detection system, typically at an acceleration voltage of 20 Kv.

General procedure for the synthesis of Ti3AlC2 MAX phase
The milling process was performed in order to synthesize Ti3AlC2 MAX phase in Ti–Al–C system. For this purpose, a mixture of commercially available powders of Ti, Al, and C with a stoichiometric ratio corresponding to the preparation of Ti3AlC2, with Ti:Al:C = 3:1:2, was milled by a planetary ball mill for 10 h. A milling chamber made of tungsten carbide of 220 mL volume and balls made of zirconia of 2-8 mm diameter were applied for the milling operation.

General procedure for the synthesis of Ti3C2OH MXene Nanomaterial
The Ti3C2OH MXene was prepared using selective etching of Al from the prepared Ti3AlC2 MAX phase in the HF solution [25]. In a typical synthesis process, 2 g of the prepared MAX powders were slowly added to 50 mL of HF solution (40 wt%). The etching process was continued for 24 h, while stirring proceeded with the help of a magnetic Teflon-coated bar. Next, the mixture was washed several times with deionized water and finally centrifuged. After that, the powder samples were washed with ethanol and dried in a vacuum oven.

General procedure for the preparation of compounds 4a-g
A mixture of 3,4-methylenedioxyphenol (1, 1 mmol), malononitrile (2, 1 mmol), 5-substituted isatins 3 (1 mmol), and Ti3C2OH MXene (0.05 g) was grinded with a mortar and pestle at room temperature, in the presence of small amounts of ethanol, for about 20 minutes. TLC in a 3:2 ethyl acetate/n-hexane TLC solvent was used to monitor the reaction’s progression. After the reaction was finished, the reaction mixture was dissolved in 3 mL of hot ethanol and centrifuged for five minutes at 2000-3000 rpm to extract the catalyst for reuse. The mixture was then repeatedly rinsed with ethanol and dried. The ethanol solution was cooled to room temperature, diluted with 1 mL of water, and then allowed to crystallize to produce the pure product.

Spiro[3,8’]5-bromo-1,3-dihydro-2H-indol-2-one-6’-amino-8’H-[1’,3’]dioxolo[4’,5’-g][1]benzopyran-7’-yl cyanide (4b)
White powder, m.p. 355-357 ˚C, yield: 0.387 g (94%). IR (KBr): νmax 3324, 3289, 3228, 2862, 2119, 1660, 1541, 1325, 1218 cm-1. 1H NMR (500 MHz, DMSO-d6): δ 5.86 (1 H, d, J = 1.2 Hz, CH2), 5.96 (1 H, d, J = 1.2 Hz, CH2), 7.33 (1 H, brd, J = 7.9 Hz, HAr), 7.47 (1 H, s, HAr), 7.58 (1 H, s, HAr), 7.62 (2 H, brs, NH2), 7.81 (1 H, m, HAr), 7.90 (1 H, s, HAr), 10.93 (1 H, s, NH) ppm. 13C NMR (125 MHz, DMSO-d6): δ 59.6, 73.6, 97.6, 102.2, 104.5, 107.9, 116.1, 117.9, 123.0, 124.2, 128.4, 130.9, 144.9, 147.7, 146.4, 148.2, 159.4, 172.7 ppm. Anal. Calcd for C18H10BrN3O4 (412.19): C 52.45, H 2.45, N 10.19; Found: C 52.31, H 2.62, N 10.30.

Spiro[3,8’]1,3-dihydro-5-hydroxy-2H-indol-2-one-6’-amino-8’H-[1’,3’]dioxolo[4’,5’-g][1]benzopyran-7’-yl cyanide (4d)
Pale yellow powder, m.p. 319-321 ˚C, yield: 0.325 g (93%). IR (KBr): νmax 3368, 3276, 3213, 2944, 2112, 1664, 1533, 1334, 1210 cm-1. 1H NMR (500 MHz, DMSO-d6): δ 5.79 (1 H, d, J = 1.2 Hz, CH2), 5.86 (1 H, d, J = 1.2 Hz, CH2), 6.88 (1 H, brd, J = 7.5 Hz, HAr), 7.04 (1 H, s, HAr), 7.53 (1 H, s, HAr), 7.67 (2 H, brs, NH2), 7.75 (1 H, brd, J = 7.5 Hz, HAr), 7.89 (1 H, s, HAr), 9.31 (1 H, s, OH), 11.02 (1 H, s, NH) ppm. 13C NMR (125 MHz, DMSO-d6): δ 60.0, 74.5, 99.3, 101.8, 103.8, 108.6, 114.3, 118.4, 122.7, 125.3, 129.6, 144.6, 146.1, 147.4, 148.5, 158.6, 160.1, 173.4 ppm. Anal. Calcd for C18H11N3O5 (349.30): C 61.89, H 3.17, N 12.03; Found: C 62.07, H 3.27, N 11.91.

RESULTS AND DISCUSSION
The main aim of this research, as shown in Fig. 2, is to develop the catalytic applications of the Ti3C2OH MXene 2D nanomaterial for the environmentally friendly synthesis of a series of spiro-fused oxindole-dioxolo[g][1]benzopyrans. Ti₃C₂OH MXene was obtained through HF etching of Ti₃AlC₂ MAX phase, while the MAX phase precursor was synthesized using a ball milling-assisted method that reduces energy consumption and improves the sustainability of the overall preparation process [25]. The EDS analysis confirms the presence of oxygen on the Ti3C2TX surface, signifying the successful formation of oxygen-containing functional groups as a hallmark of etched MXene materials. The structural correctness of the synthesized nanomaterial was then assessed using several techniques, including X-ray diffraction (XRD) analysis, scanning electron microscopy (SEM), and energy dispersive X-ray spectrometry (EDS). The XRD patterns of the separated pure objects, Ti3AlC2 MAX and Ti3C2OH nanomaterials, are illustrated in Fig. 3.
According to the XRD analysis, the peaks corresponding to Ti3AlC2 MAX and Ti3C2OH were observed, while Ti3AlC2 was the main phase. By transforming the Ti3AlC2 MAX phase into Ti3C2OH MXene, a remarkable shift of the (002) plane to the lower angles and vanishing of the sharpest peak of Ti3AlC2 at 2θ = 39˚ in the XRD patterns can be observed. These changes reveal the successful conversion of Ti3AlC2 into Ti3C2OH [26]. The (002) diffraction peak at an angle of 6.6˚ can be utilized for the assessment of the d-spacing between the MXene nanosheets and the monolayer thickness. Compared to the MAX phase precursor, the etching procedure yielded MXene with expanded interlayer spacing, evidenced by an increase in the c-lattice parameter of their crystallites.
Fig. 4 shows the SEM micrograph of the MAX product after 10 h milling and its EDX analysis. The layer structure of Ti3AlC2 together with granular grains can be seen. The formation of the observed agglomerates suggested that a mechanically induced self-propagating reaction occurred. This procedure releases a large amount of heat, which not only makes reactions occur but also raises the temperature in the microzone at a high level, resulting in the sintering of small parts of powders to coarse granules. However, this agglomerate can easily be de-agglomerated [27]. The existence of Ti, Al, and C elements and their elemental percentage can be confirmed via the EDX curve and the presented elemental maps.
According to Fig. 4g, a more developed layered structure of MXene can be observed, indicating the efficient removal of Al from the MAX phase. The EDS analysis shows that Al was drastically reduced during the etching process, confirming the near-complete extraction of Al from the parent MAX structure. The EDS spectrum confirms the presence of oxygen on the MXene surface, suggesting the existence of oxygen-containing surface terminations.
To further elucidate the chemical nature and origin of these oxygen species, FT-IR spectroscopy was employed (Fig. 5). The spectrum exhibits characteristic absorption bands at approximately 3400 and 1083 cm−1, which are attributed to OH and C-O stretching vibrations, respectively. These features suggest the integration of hydroxyl and epoxy terminal groups within the MXene structure. Furthermore, the prominent peak at 563 cm−1 is assigned to the Ti–O lattice vibrations [28]. The sharp band at 1637 cm−1 corresponds to the H–O–H bending mode of structural water molecules intercalated within the MXene interlayer galleries. Notably, the absence of distinct peaks in the regions of 1700-1725 and 1210-1320 cm−1 confirms the lack of carboxylic groups (COOH), indicating that the surface remains free from significant oxidative degradation into organic acid forms.
As shown in Fig. 6, a typical reaction of 3,4-methylenedioxyphenol (1), malononitrile (2), and 5-bromoisatin (3b) was employed for the synthesis of compound 4b under various reaction conditions, with the results presented in Table 1. Conducting the reaction without any catalyst at reflux temperature in ethanol was examined first. After 6 hours, only 52% of the product was obtained (Table 1, entry 1). The reaction was also performed without any catalyst under ethanol-drop grinding at room temperature; a low conversion was observed on TLC after 60 minutes (Table 1, entry 2). However, this typically requires a catalyst to drive the reaction. Then, the reaction was evaluated using Ti3C2OH MXene as a new heterogeneous catalyst in both conditions: ethanol reflux or ethanol-drop grinding. As can be observed, grinding is more efficient than reflux and requires less reaction time and temperature (Table 1, entries 3 & 4). To identify the optimum amounts of the catalyst, the reaction was carried out with 0.04, 0.05, and 0.06 g of Ti3C2OH MXene; conversion yields of 80%, 94%, and 92% were obtained, respectively, after 20 minutes grinding (Table 1, entries 4-6). Next, the replacement of the Ti3C2OH MXene with some Brønsted acids such as HCl and p-TSA (p-toluenesulfonic acid) in the model reaction led to moderate conversions after 20 minutes grinding (Table 1, entries 4 & 6-7). Additionally, the impact of grinding duration was examined, and it was found that only 20 minutes of grinding could successfully convert the particular reactants (Table 1, entries 4 & 9-10).
We investigated the substrate scope by selecting several of 5-substituted isatins after establishing the optimal conditions for the aforementioned reaction. This was tested in the reaction with 3,4-methylenedioxyphenol, malononitrile, resulting in the production of the target compounds whitin 20 minutes in high yields (Table 2).
For the synthesis of 4b under optimal reaction conditions, the next step was to examine the catalyst’s recyclability. The recovered catalyst can be recycled five times, although there is a slight decrease in activity (Table 3).
Fig. 7 illustrates a feasible pathway for producing product 4, based on the explanatory processes reported in earlier research. The Lewis acidic sites on the nanostructure are believed to enhance the initial Knoevenagel reaction between isatin 3 and malononitrile 2, resulting in the formation of alkene 6 in the presence of Ti₃C₂OH MXene. When 3,4-methylenedioxyphenol 2 is added to alkene 6, it creates the Michael adduct 7, which then cyclizes to form intermediate 8. Compound 4 is generated when intermediate 8 undergoes further tautomerization.
In order to evaluate the benefits of the current method over the previously published procedure, we also evaluated the yields, conditions, scope, and generality of this approach in synthesizing spiro[3,8’]1,3-dihydro-2H-indol-2-one-6’-amino-8’H-[1’,3’]dioxolo[4’,5’-g][1]benzopyran-7’-yl cyanides (Table 4).

CONCLUSION
In this study, we developed a mechanochemical method for synthesizing Ti₃C₂OH MXene and employed it for the first time as an effective heterogeneous catalyst to produce spiro-fused oxindole-dioxolo[g][1]benzopyran derivatives. The successful fabrication of the nanocatalyst, which exhibits strong structural integrity, thermal stability, and oxygen-containing surface functionalities, possibly including OH groups, was confirmed through comprehensive characterization tests using FT-IR, XRD, SEM, and EDS. We assessed and validated the target products using melting points, FT-IR spectroscopy, ¹H NMR, ¹³C NMR, and elemental analysis. This approach offers a faster, more efficient, and environmentally friendly alternative compared to traditional methods. From an industrial perspective, this strategy is likely to reduce costs, minimize waste, and enhance energy efficiency.

ACKNOWLEDGMENT
Shahrzad Abdolmohammadi expresses her gratitude to the Research Council of the South Tehran Branch, Islamic Azad University, for their technical help.

CONFLICT OF INTEREST
The authors declare that there is no conflict of interests regarding the publication of this manuscript.