Over the past few decades, supramolecular chemistry has advanced intensively establishing itself as a central discipline in modern chemistry . This tremendous growth allowed the field to transcend the traditional chemistry boundaries and move toward real-life applications . In this context, supramolecular materials with pre-organized structures have been designed and exploited for diverse environmental, industrial and biological applications .

Among these macrocycles, calix[4]pyrroles (CPs) represent a modern generation of supramolecular materials with interesting chelation properties . These nonplanar, non-aromatic, tetrapyrrolic macrocycles consist of four pyrrolic units linked together at their 2,5-positions through sp3-hybridized meso-carbon atoms . The ability of CPs to selectively bind anions , cations, ion pairs and neutral guest species, combined with the versatility to introduce diverse functional groups, has enabled the fabrication of calix[4]pyrrole-based supramolecular matrices for a broad range of applications . Notable examples include catalysis, chromogenic sensors and fluorescent sensors, as well as heavy metals extraction .

One CP molecule that has attracted the attention of several research groups is the meso-tetra-methyltetrakis(4-hydroxyphenyl)calix[4]pyrrole, commonly referred to as phenoxycalix[4]pyrrole (PCP). It was first reported in 1999 by two independent research groups, Floriani et al. and Sessler et al., via the acid-catalyzed condensation of pyrrole with p-hydroxyacetophenone . PCP and its derivatives were first known for their anion chelation capability achieved through the formation of hydrogen bonds with the pyrrolic units . Subsequent studies have demonstrated that the introduction of additional chelation sites via extension of the phenolic groups of PCP can yield anionic receptors, ditopic receptors bearing both anion- and cation-binding sites, as well as systems capable of promoting ion-pair formation .

As effective chelating agents for a broad range of transition metals, hydroxamic acids constitute an important class of organic compounds that have attracted considerable attention and have found diverse applications across both biomedical and industrial fields . Hydroxamic acids are also well known for their strong chelating ability toward uranyl, forming stable complexes through the synergistic coordination of the carbonyl and hydroxylamine groups . Uranium is a naturally occurring radionuclide and is thus an integral constituent of the environment . However, beyond its extensive use as fuel for nuclear power generation, uranium is released into the environment through various industrial processes, such as phosphate fertilizer production and metal refining, contributing to the increased accumulation of uranium in ecosystems . Owing to its radioactivity, chemical toxicity, and long half-life, environmental uranium contamination poses serious ecological and public health risks . In this context, hydroxamic acid functionalities have been successfully incorporated on supramolecular architectures to explore their potential for the removal of uranium from aqueous media. For instance, it has been long known that hydroxamic acid derivatives of calix[4]arenes and calix[6]arenes act as excellent uranophiles as demonstrated in early studies from 1991 . A calix[4]resorcinarene hydroxamic acid has also shown a pronounced binding tendency and selectivity for uranyl and proved to be applicable for the determination of uranium in standard and environmental samples . Importantly, more recent studies have demonstrated that pre-organized or cyclic hydroxamate ligands can display significantly enhanced binding toward uranyl relative to their linear hydroxamic acid counterparts, emphasizing the beneficial role of structural organization in uranyl chelation . This beneficial role of pre-organization of chelating groups within one molecule was also highlighted for supramolecular platforms like calixarenes for actinides and lanthanides extraction .

In light of this background, we report here the incorporation of hydroxamic acid into phenoxycalix[4]pyrrole and examine its effectiveness in extracting uranyl from aqueous solution using gamma spectroscopy. The extraction performance was systematically evaluated as a function of pH and ligand-to-metal molar ratio to study the effect of these two key parameters on the extraction process. In doing so, this work aligns with current research dedicated to the design of functional supramolecular materials capable of remediating uranium from aqueous environments .

Various strategies have been developed for the synthesis of hydroxamic acid derivatives over the years . Classical and most popular routes involve the direct reaction of carboxylic acid derivatives such as esters, acyl chlorides and anhydrides with hydroxylamine salts . A variety of coupling or activating agents were also employed in case of simple addition of hydroxylamine to carboxylic acid compounds . In addition, alternative methods starting from aldehydes , alcohols , and amides have also been reported. However, the literature states that there is no particular reagent or reaction condition that can serve as a general rule for the synthesis of hydroxamic acids .

In this study, the direct reaction of hydroxylamine with the PCP ester was first examined, since the ester functions as the synthetic precursor to the corresponding acid used in the preparation of PCP derivatives from PCP . For that, PCP was initially synthesized through the molecular cyclisation of pyrrole with 4-hydroxyacetophenone in methanol in the presence of methanesulfonic acid as first reported by Floriani et al. and Sessler et al. . This condensation reaction produces several PCP isomers, including the targeted pre-organized α,α,α,α-isomer, which is isolated by crystallization from glacial acetic acid, followed by removal of the acetic acid using a mixture of acetonitrile and acetone. Using this protocol, the PCP α,α,α,α-isomer was recovered in 47% yield, as previously reported by Namor and Shehab . Subsequently, the synthesis of the PCP ester derivative (PCP E) was carried out via O-alkylation of PCP with ethyl bromoacetate in dry acetone/K2CO3, following the method reported by Camiolo and Gale , affording a yield of 85%.

Figure 1: Synthetic route of PCP HA.

In this study, two parameters that can influence the solid-liquid extraction efficiency of PCP-HA toward uranyl, namely pH and ligand-to-metal ratio, were studied. To ensure comparison between extraction experiments, parameters like temperature, shaking time and volume of uranyl solution used were kept constant. In a typical experiment, 20 mL of a 1 mM aqueous uranyl acetate solution was adjusted to the target pH and then added to solid PCP HA. The solution was shaken at a fixed temperature of 25 °C for 4 h. This time was estimated to be sufficient to reach equilibrium. The heterogeneous solution was centrifuged and the aqueous layer was filtered prior to gamma spectroscopy analysis. Uranium activity concentrations before and after extraction were determined, and extraction efficiencies were calculated from the decrease in activity concentration. It is important to note that the extraction efficiency of PCP HA was evaluated based solely on the quantification of 238U. Uranium is a naturally occurring radionuclide that exists predominantly as a mixture of three isotopes: 238U, 235U and 234U. Among these, 238U is by far the most abundant, accounting for 99.28 % by mass, followed by 235U (0.72 %) and 234U (0.0058 %). Although these isotopes possess identical chemical properties, they differ in their radioactive characteristics . Since the extraction experiments depend on the chemical behavior of uranium rather than its radiological properties, any of these isotopes could, in principle, be used. Two main factors justify the choice of 238U: its natural abundance and its reliable detectability by gamma spectroscopy, in contrast to 235U, which exhibits weak gamma emission intensity .

Table 1: Effect of pH on uranyl extraction by PCP HA.a

aA₀ and A₁ represent the activity concentrations of 238U in the uranyl acetate working solution before and after extraction, respectively. σ(A₀) and σ(A₁) denote the uncertainties associated with the measured activity concentrations determined by gamma spectroscopy. σ(% E) represents the uncertainties of the individual extraction efficiencies. Extractions were carried out at 25 °C using aqueous uranyl acetate solutions of 1 mM, with the pH being adjusted with 0.1 M HCl. PCP HA was added at a ligand-to-metal molar ratio of 1:1. Samples were equilibrated for 4 h under constant agitation before phase separation and activity measurements.

Figure 2: Effect of pH on the mean uranyl extraction efficiency (% Emean) of PCP HA.

This behavior is consistent with previous studies in which ligands displayed high uranyl uptake within an optimal pH window. For instance, calix[4]arene-based 8-hydroxyquinoline ligands showed nearly quantitative extraction of uranyl between pH 4 and 9, with efficiency decreasing under more acidic conditions . Likewise, phosphoramide-functionalized magnetic nanoparticles showed high uranyl adsorption (80–95%) between pH 4 and 8, with reduced uptake outside this range .

Although hydroxamic acids typically have pKa values of 7–9 , supramolecular systems bearing these groups do not exhibit uniform uranyl extraction behavior under identical pH conditions. Consequently, some systems achieve maximum extraction under weakly acidic conditions, while others perform better at higher pH values. For instance, an octa-functionalized calix[4]resorcinarenehydroxamic acid exhibits quantitative extraction of uranyl into ethyl acetate solution at pH value of 8 . On the other hand, calix[4]arenes and calix[6]arenes functionalized with hydroxamic acid groups show a marked increase in uranyl extractability from aqueous solution into chloroform at pH 3–4, reaching nearly 100% extractability at pH 5. The enhanced extraction efficiency observed in the acidic pH region has been attributed to the metal-ion-assisted deprotonation. The coordination of the hydroxamic acid groups to the uranyl cation stabilizes the deprotonated hydroxamate form, effectively lowering the apparent pKa and enabling strong binding even at low pH . It is important to mention that the formation of the stable 5-membered chelate between the uranyl ion and the Z-isomer of hydroxamic acid effectively shifts the Z/E equilibrium towards the Z-conformation through a metal-induced fit mechanism . Consistent with known hydroxamate coordination chemistry, coordination-induced deprotonation of the hydroxamic acid at low pH provides the conditions required for effective uranyl complexation in our system.

To further understand the complexation behavior of PCP HA toward uranyl, the effect of the ligand-to-metal molar ratio on the extraction efficiency was investigated. Although the previous experiment revealed maximum extraction at pH values of 2 and 3, the investigation of the ligand-to-metal molar ratio was conducted at pH 4. This adjustment was made to allow a clearer assessment of the molar ratio effect. At pH 3, a 1:1 ratio of PCP HA to uranyl already yielded about 95% extraction, leaving little sensitivity to detect variations at higher ligand mass. By performing the experiment at pH 4, where the extraction efficiency was approximately 86%, the system remained sufficiently responsive to changes in ligand mass.

Table 2: Effect of ligand–metal ratio on uranyl extraction by PCP HA.

Figure 3: Effect of ligand-to-metal molar ratio on the mean uranyl extraction efficiency (% Emean) of PCP HA.

The observed efficiency depends not only on ligand mass but also on the intrinsic coordination preferences with uranyl. In aqueous solution, hexavalent uranium, the most dominant oxidation state, exists predominantly as the linear uranyl ion UO22+. In this ion, two oxo ligands occupy axial positions while the equatorial plane accommodates four to six donor atoms . Complexes of UO22+ often adopt either a pseudoplanar pentacoordinate or hexacoordinate structure, as shown by X-ray crystallographic studies. Consequently, to bind UO22+ effectively, a ligand must present donor atoms positioned to match the uranyl equatorial coordination sites .

Hydroxamic acids act as bidentate ligands, with each functional unit offering two donor atoms, a carbonyl oxygen and a hydroxy oxygen, that can simultaneously coordinate to a uranyl ion . In supramolecular systems, not all hydroxamic acid groups necessarily bind uranyl ions, as steric hindrance and site accessibility can limit coordination to a subset of available sites . A relevant example is a calix[6]arene functionalized with three hydroxamic acid groups, where a theoretical study using density functional theory (DFT) calculations showed that the complex with the uranyl is most stabilized when only two of the three hydroxamic acid units participate in binding . This concept of partial participation and cooperative interactions between hydroxamic acid units may also be relevant in the PCP HA system, where not all functional groups are necessarily involved in uranyl binding.