Oxygen-Containing Quaternary Phosphonium Salts (oxy-QPSs): Synthesis, Properties, and Cellulose Dissolution

In the present study, the synthesis of oxygen-containing quaternary phosphonium salts (oxy-QPSs) was described. Within this work, structure–property relationships of oxy-QPSs were estimated by systematic analysis of physical–chemical properties. The influence of the oxygen-containing substituent was examined by comparing the properties of oxy-QPSs in homology series as well as with phosphonium analog-included alkyl side chains. The crystal structure analysis showed that the oxygen introduction influences the conformation of the side chain of the oxy-QPS. It was found that oxy-QPSs, using an aprotic co-solvent, dimethylsulfoxide (DMSO), can dissolve microcrystalline cellulose. The cellulose dissolution in oxy-QPSs appeared to be dependent on the functional group in the cation and anion nature. For the selected conditions, dissolution of up to 5 wt% of cellulose was observed. The antimicrobial activity of oxy-QPSs under study was expected to be low. The biocompatibility of oxy-QPSs with fermentative microbes was tested on non-pathogenic Saccharomyces cerevisiae, Lactobacillus plantarum, and Bacillus subtilis. This reliably allows one to safely address the combined biomass destruction and enzyme hydrolysis processes in one pot.


Introduction
Ionic liquids (ILs) are a well-known part of organic salts consisting of a cation (imidazolium, ammonium, phosphonium, etc.) and an inorganic anion with a melting point commonly below 100 • C [1].A unique set of properties, such as nonvolatility, nonflammability, thermal stability, and reusability, are more or less characterized by ionic liquids [2].The combination of the useful properties and a broad diversity of structures make ILs widely used as solvents for reactions, including those catalyzed by nanocatalysts [3], as active pharmaceutical drugs [4], as antibacterial agents [5,6], and in the design of accumulators [7] and smart materials [8].
Considering the tremendous potential for a wide structural diversity, a reasonable stage of development was the emergence of task-specific ionic liquids (TSILs) [9].This is a type of highly customized ILs that do not only serve as solvents; for example, through functionalization with oxygen-containing groups, their properties (viscosity, lyophilicity, hydrophilicity, hydrogen bonding ability, and electrical conductivity) can be easily tuned in a wide range, making them suitable for applications in organocatalysis, synthesis, gas absorption, supercapacitor development, and analytical chemistry [10][11][12][13].Interest in the functionalization of ILs is still maintained, and the effect of the ether group on the physical constants and crystal packing is investigated [14].
One of the most abundant oxygen-containing phosphonium salts, terakis(hydroxymethyl)phosphonium chloride, is produced in multi-ton quantities and is widely used as a biocide and flame retardant, in the oil industry, in the leather industry as a tanning agent, and in the production and stabilization of nanoparticles [15].Oxygen-containing phosphonium salts were quite effective solvents for the Grignard reaction [16].Phosphonium salts with multiple ester groups in the side chain have demonstrated applicable properties (viscosity and ionicity) suitable for use as electrolytes in lithium-ion batteries [17] and have been established as an excellent reaction medium for lipase-catalyzed reactions [18].Phosphonium salts with ester, ether, and hydroxyl groups as the fourth substituent are more suitable for biodegradation than tetraalkylphosphonium salts [19].In this regard, professor Ragogna's group demonstrated the antibacterial properties of the material containing functionalized phosphonium compounds [20,21].
The use of phosphonium salts as a cellulose dissolution agent has recently gained prominence.The ability of tetrabutylphosphonium 2-ethylhexanoate to dissolve cellulose from corn stover has been demonstrated.It is the best performing of the compounds used in this study, even over imidazolium compounds.At the same time, the authors refused to try to recycle the IL because of the challenges associated with the energy consumption of water evaporation [22].
The next generation of amino acid phosphonium ILs was introduced to dissolve cellulose for subsequent processing by enzymes [23].Using the tetrabutylphosphonium N,Ndimethylglycine system and DMSO co-solvent, 15 wt% cellulose dissolution was achieved at 30 • C, and enzymatic conversion of dissolved cellulose to sugars was completed.
The quantitative structure-activity relationship (QSAR) model was developed for predicting cellulose solubility using the example of oxygen-containing phosphonium salts [24].The designed model was successfully performed and showed an excellent R 2 factor, but it did not include a parameter characterizing the high viscosity of resulting IL/cellulose mixtures.Nevertheless, a triphenylphosphonium salt containing a methoxymethyl moiety and acetate anion exhibited high cellulose solubility as predicted by the modeling.It has also been previously reported that the size and structure of the phosphonium cation play a dramatic role in various processes [25].The search for the relationship between the ability of ILs to successfully participate in biomass preparation from various factors has long been underway.In general, comparisons are made from the structure and composition of cellulose or the composition of ionic liquids, their physical characteristics, and other parameters [26].
Earlier, we reported the synthesis and application in catalysis of sterically hindered ester containing phosphonium salt [27].In the current work, we systematically studied the influence of the oxygen-containing substituents in cation on the physical-chemical properties of oxy-QPSs.The obtained trends were compared with the previously reported alkyl-substituted analog [28,29].Some features of the crystal packing of oxy-QPSs were discussed.We utilized a series of compounds under the study for microcrystalline cellulose dissolution.The safety of oxy-QPSs for humans and the environment was also estimated.
Electro-spray ionization mass spectrometry (ESI-MS): ESI-MS measurements were performed using Maxis and MicroTOF II time-of-flight high-resolution mass spectrometers (Bruker Daltonic GmbH, Bremen, Germany) in positive mode in the mass range of m/z 50-3000.Direct syringe injection was used for all analyzed samples in acetonitrile solution at a flow rate of 5 µL/min.The capillary voltage was −4500 V, spray shield offset was −500 V, nitrogen nebulizer gas was −1 bar, nitrogen drying gas was 4 L•min −1 , and desol-vation temperature was 200 • C. The instruments were calibrated with a low-concentration tuning mixture (Agilent Technologies G2431A, Santa Clara, CA, USA).Data processing was performed by DataAnalysis 4.0 SP4 software (Bruker Daltonik GmbH, Bremen, Germany).
Thermogravimetry: A coupled system of a STA449-F3 TG/DSC synchronous thermal analysis instrument (Netzsch, Selb, Germany) with a Tensor 27 IR-Fourier spectrometer (Bruker, Ettlingen Germany) was used to study the decomposition temperature and thermal stability of the tested compounds.The samples (3.4-12.1 mg) were placed in aluminum crucibles with a perforated lid and heated in the range of 30-400 • C together with an empty crucible as a reference sample.TG/DSC measurements were carried out at a heating rate of 10 K/min in an argon flow of 50 mL/min.The resolution of Tensor 27 is 4 cm −1 .
Single crystal X-Ray analysis: The X-ray diffraction data for compounds were recollected at 100 K on a four-circle Synergy S diffractometer (Rigaku, Wrocław, Poland) equipped with a HyPix600HE area-detector (kappa geometry, shutterless ω-scan technique), using graphite monochromatized Cu K α -radiation; the intensity data were integrated and corrected for absorption and decay by the CrysAlisPro program [30].The structures were solved by direct methods using SHELXT [31] and refined on F 2 using SHELXL-2018 [32] or OLEX2 [33].
Optical microscopy: Optical microscope Micromed-1 (Saint-Petersburg, Russia) was used for the examination of cellulose dissolution.

Materials
All the work related to the preparation of the starting substrates, as well as the synthesis and the workup of products, was carried out in an inert N 2 atmosphere using the standard Schlenk apparatus.All solvents and purchased reagents were absoluted by the appropriate methods, mainly by distillation in an inert atmosphere.

General Procedure of the Synthesis of oxy-QPSs
Tri-tert-butylphosphine and alkylating agent were stirred at 50-100 • C for 4-8 h.The reaction mixture was cooled; white solid was washed with diethyl ether (see Supplementary Materials for details).For couple of syntheses, the solvent acetonitrile was used.After cooling, the reaction mixture solvent was evaporated and then washed with diethyl ether.

Screening Dissolution of Microcrystalline Cellulose in oxy-QPSs
1 mg of microcrystalline cellulose (50 µm, Avicell), 50 mg of oxy-QPS, and 47 µL of DMSO were added to the 5-mL glass tube equipped with magnetic stirring bar.The reaction vessel was sealed with a screw cap.The process was carried out at 80 • C. For a certain time interval, a drop of solution was collected and placed on glass slide.Monitoring the dissolution of cellulose was administrated by direct observation using optical microscope at 40-100× magnification.

Antimicrobial and Antifungal Activity of oxy-QPSs
The antimicrobial activity of the oxy-QPSs was determined by serial dilution technique in Mueller-Hinton broth 2 for pathogenic bacteria, MRS medium for the cultivation of lactobacilli, and Sabouraud broth for yeast.The cultures used for testing included the Gram-positive bacteria Staphylococcus aureus ATCC 6538P FDA 209P, Bacillus cereus ATCC 10702 NCTC 8035, Enterococcus faecalis ATCC 29212; Gram-negative pathogenic bacteria Escherichia coli ATCC 25922; Gram-positive non-pathogenic endospore-forming bacteria Bacillus subtilis ATCC 6633; and pathogenic yeast Candida albicans ATCC 10231 from the State Collection of Pathogenic Microorganisms and Cell Cultures "SCPM-Obolensk".Gram-positive non-pathogenic non-spore-forming bacteria Lactobacillus plantarum 8P-A3 and non-pathogenic yeast Saccharomyces cerevisiae 1986 were from the collection of Kazan Federal University.The bacterial load was 3.0 × 10 5 CFU/mL.The fungal load was 2.0 × 10 3 CFU/mL.Results were recorded every 24 h for 5-7 days.Bacterial cultures were incubated at 37 • C, and yeast cultures were incubated at 25 • C. The experiment was repeated three times.Compound dilutions were prepared immediately on nutrient media.The dilutions of the oxy-QPSs were prepared immediately in nutrient media; 5% aqueous solution of DMSO was used to improve the solubility of the oxy-QPSs, and the test strains were not inhibited at this concentration.The minimum inhibitory concentration (MIC) was defined as the minimum concentration of a compound that inhibits the growth of the respective test microorganism [34,35].The growth of bacteria, as well as the absence of growth due to the bacteriostatic action of a compound, was recorded.To determine the minimum bactericidal concentration (MBC), an aliquot of the culture of pathogenic bacteria was transferred onto Mueller-Hinton agar in a 10 cm Petri dish and incubated for 24 h at 37 • C. The MBC was the minimum concentration at which bacterial colonies were not detected, indicating that the bacteria were killed with >99.9% efficiency.

Hemolytic Activity
The hemolytic activity of oxy-QPSs was estimated by comparing the optical density of a solution containing the test compound with that of blood at 100% hemolysis.The experiments were carried out as described earlier [36].

Cell Toxicity Assay (MTT Assay)
The cytotoxic effect of the test compounds on normal human cells was determined using the colorimetric method of cell proliferation MTT (Thiazolyl Blue Tetrazolium Bromide, Sigma, Kawasaki, Japan).For this purpose, 10 µL of MTT reagent in Hank's balanced salt solution (HBSS) (final concentration 0.5 mg/mL) was added to each well.The plates were incubated at 37 • C for 2-3 h in an atmosphere humidified with 5% CO 2 .Absorbance was recorded at 540 nm using a microplate reader (Invitrologic, Novosibirsk, Russia).Experiments for all compounds were repeated three times.The Chang liver cell line (human liver cells) from N. F. Gamaleya Research Center of Epidemiology and Microbiology (Moscow, Russia) was used in the experiments.The cells were cultured on a standard nutrient medium, "Igla", produced by the Moscow Institute of Poliomyelitis and Viral Encephalitis.M.P. Chumakov (Moscow, Russia) with the addition of 10% fetal calf serum and 1% nonessential amino acids (NEAA) (PanEco, Moscow, Russia).
The cells were plated on a 96-well panel (Eppendorf SE, Hamburg, Germany) at a concentration of 5 × 10 3 cells per well in a volume of 100 µL of medium and cultured in a CO 2 incubator at 37 • C. In 48 h after planting the cells, the culture medium was taken into the wells, and 100 µL of solutions of the studied oxy-QPSs in the specified dilutions were added to the wells.Dilutions of the compounds were prepared directly in growth medium supplemented with 5% DMSO to improve solubility.The cytotoxic effect of the test compounds was determined at concentrations of 0.1-100 µM.The calculation of the IC 50 , the concentration of the drug causing inhibition of cell growth by 50%, was performed using the program MLA "Quest Graph™" IC 50 Calculator [37].

Ecotoxicity to D. magna
The test culture Daphnia magna was used to determine the LC 50 of the studied oxy-QPSs.The crustaceans were kept under standard conditions according to the recommendations published in ISO 6341 [38] and NBR 12713 [39].The sensitivity to K 2 Cr 2 O 7 was determined one week before the test; LC 50 for potassium dichromate was 1.443 mg/L.A total of 10 crustaceans were planted in each falcon with the studied solutions; the glasses were kept in a luminostat for 48 h at a temperature of 22 • C, with a 12 h illumination regime at an intensity of 600 lux.The pH of the solutions was 7.2-7.4,and the O 2 content was 6.8-7.2 mg/L.The crustaceans were not fed the day before the study and during the test.
After 48 h, a visual count of surviving daphnia was performed [40].The results were recorded in the protocol and calculated using the program "R" version 2.13.0 (13 April 2011).All compounds under the study at 1% stock concentration were readily soluble in distilled water.Further dilutions were made with biogenerated water.At the same time, a control group of daphnia was placed in biogenerated water without any of the studied substances.All the daphnia in the control group survived.
The test substances were classified as Acute 1, Acute 2, or Acute 3 to aquatic organisms based on LC 50 definitions [41].

Synthesis of oxy-QPSs
The synthesis of oxygen-containing quaternary phosphonium salts (oxy-QPSs) involves the quaternization reaction of tertiary phosphine and halogenated reagent, as in the case of alkyl-substituted QPSs.Ethers, carboxylic acids, and esters were applied as the functional groups in the cation (Scheme 1, Table 1).The coupling reaction requires the usage of an inert atmosphere due to the high sensitivity of initial tri-tert-butylphosphine to atmospheric oxygen and humidity.In turn, the reactivity of the alkylating agent reduces as chain length increases.In this respect, the synthesis conditions were harsher when a longer halogenated reagent came into the reaction (see Supplementary Materials for experimental details).According to the atom economic principle, the quaternization reactions were carried out without a solvent in most cases except for the synthesis of oxy-QPSs 7 and 8 containing carboxylic groups in the side chain.Acetonitrile was involved in these runs to exclude the formation of side products caused by the interaction of acidic proton with phosphorus.
A total of 10 crustaceans were planted in each falcon with the studied solutions; the glasses were kept in a luminostat for 48 h at a temperature of 22 °C, with a 12 h illumination regime at an intensity of 600 lux.The pH of the solutions was 7.2-7.4,and the O2 content was 6.8-7.2 mg/L.The crustaceans were not fed the day before the study and during the test.
After 48 h, a visual count of surviving daphnia was performed [40].The results were recorded in the protocol and calculated using the program "R" version 2.13.0 (13 April 2011).All compounds under the study at 1% stock concentration were readily soluble in distilled water.Further dilutions were made with biogenerated water.At the same time, a control group of daphnia was placed in biogenerated water without any of the studied substances.All the daphnia in the control group survived.
The test substances were classified as Acute 1, Acute 2, or Acute 3 to aquatic organisms based on LC50 definitions [41].

Synthesis of oxy-QPSs
The synthesis of oxygen-containing quaternary phosphonium salts (oxy-QPSs) involves the quaternization reaction of tertiary phosphine and halogenated reagent, as in the case of alkyl-substituted QPSs.Ethers, carboxylic acids, and esters were applied as the functional groups in the cation (Scheme 1, Table 1).The coupling reaction requires the usage of an inert atmosphere due to the high sensitivity of initial tri-tert-butylphosphine to atmospheric oxygen and humidity.In turn, the reactivity of the alkylating agent reduces as chain length increases.In this respect, the synthesis conditions were harsher when a longer halogenated reagent came into the reaction (see Supplementary Materials for experimental details).According to the atom economic principle, the quaternization reactions were carried out without a solvent in most cases except for the synthesis of oxy-QPSs 7 and 8 containing carboxylic groups in the side chain.Acetonitrile was involved in these runs to exclude the formation of side products caused by the interaction of acidic proton with phosphorus.

oxy-
The general reaction of oxy-QPS synthesis.
The oxy-QPSs 1 and 4 were introduced into the metathesis reaction to give 2 and 5-6 included tetrafluoroborate-or hexafluorophosphate-anions.The procedure took place in an aqueous solution to ensure complete conversion due to the insolubility of products in water.
The oxy-QPSs with halogen anions were found to be soluble with most polar organic solvents (chloroform, DMSO, acetone, ethanol, etc.) and water.The compounds 1 and 3-4 are also soluble in diethyl ether and other non-polar solvents as well.The phosphonium salts 2 and 5-6 are poorly dissolved in diethyl ether.

31 P NMR Spectral Data
The 31 P{ 1 H} NMR spectra of oxy-QPSs exhibited a singlet in a range of 44.0-52.0ppm (Table 2), except for 6, where the signal was accompanied by a septet of a hexafluorophosphate group.The chemical shift of the compounds depends on the proximity of the oxygen atom to the phosphorus one.The chemical shift of 1-2 and 4-6 appears in higher magnetic fields in narrow regions (44.0-44.7 ppm) due to the compensation of electronic density withdrawn by the oxygen of the ether group.The 31 P{ 1 H} NMR spectra of 7 and 8 are around 50 ppm and close to those for alkyl-substituted sterically hindered phosphonium salts.The anion weakly influenced the chemical shift in 31 P{ 1 H} NMR spectra; the difference is 0.4 and 0.5 ppm for 1-2 and 4-6, consequently.

1 H NMR Spectral Data
Both sterically hindered QPSs with the linear alkyl substituent and oxy-QPS α-protons are the most sensitive to the structure modification (Figure 1).The signal of α-protons in 1 H NMR spectra exhibited a multiplet and varies depending on the functional group neighboring to (Table 2).The chemical shift to the high field is characteristic of the substituents where α-protons are separated by three single bonds or more from the oxygen atom (for 3, 7-8).

1 H NMR Spectral Data
Both sterically hindered QPSs with the linear alkyl substitu tons are the most sensitive to the structure modification (Figure 1) in 1 H NMR spectra exhibited a multiplet and varies depending neighboring to (Table 2).The chemical shift to the high field is cha uents where α-protons are separated by three single bonds or mor (for 3, 7-8).Anion exchange influences the chemical shift of α-protons as phonium tetrafluoroborate or hexafluorophosphate appears in (4.55-4.62 ppm) than in halides (5.11-5.17ppm).The chemical sh QPSs with carboxylic group 9-12 is observed in a range of 4.07-4 3.2.3. 13

C NMR Spectral Data
The most influenced carbon atom in oxy-QPSs is the α-carb phorus one in the functional substituted group (Figure 1, Table 2 phosphorus and oxygen atoms in 1-2 and 4-6 is the most desh 13 C{ 1 H} NMR spectra as a doublet in a range of 59.8-61.9ppm.Th on the chemical shift of α-carbon is about 0.6 ppm.

IR
The IR spectral data of oxy-QPSs showed bands in the regi was associated with the stretching vibrations of the P-C groups served at around 2910-2980 cm −1 , which were assigned with the the dangling CH2/CH3 groups (Figure 2, Figures S37-S48 in Supp Anion exchange influences the chemical shift of α-protons as well.The signal in phosphonium tetrafluoroborate or hexafluorophosphate appears in a higher magnetic field (4.55-4.62 ppm) than in halides (5.11-5.17ppm).The chemical shift of α-protons in oxy-QPSs with carboxylic group 9-12 is observed in a range of 4.07-4.27ppm.

13 C NMR Spectral Data
The most influenced carbon atom in oxy-QPSs is the α-carbon nearest to the phosphorus one in the functional substituted group (Figure 1, Table 2).The α-carbon between phosphorus and oxygen atoms in 1-2 and 4-6 is the most deshielded and presented in 13 C{ 1 H} NMR spectra as a doublet in a range of 59.8-61.9ppm.The influence of the anion on the chemical shift of α-carbon is about 0.6 ppm.

IR
The IR spectral data of oxy-QPSs showed bands in the region 810-800 cm −1 , which was associated with the stretching vibrations of the P-C groups.Other bands were observed at around 2910-2980 cm −1 , which were assigned with the stretching vibrations of the dangling CH 2 /CH 3 groups (Figure 2, Figures S37-S48 in Supplementary Materials).
For compounds 1-6 containing ether functional groups, bands of significance were around 1200-1050 cm −1 , which were due to stretching antisymmetric vibrations of the C-O-C groups.The spectra of 1 and 3 have absorbance at 1182 and 1097 cm −1 and 1175 and 1094 cm −1 , respectively.Four bands were assigned for 4-5 in the same region: 1178, 1135, 1116, and 1086 cm −1 .
Strong absorbance at 1734 and 1741 cm −1 was associated with the stretching vibrations of the C=O groups in 7 and 8.The stretching vibrations of the C=O ester groups in 9-12 have absorbance at 1740, 1735, 1732, and 1727 cm −1 , accordingly.

IR
The IR spectral data of oxy-QPSs showed bands in the region 810-800 cm −1 , which was associated with the stretching vibrations of the P-C groups.Other bands were observed at around 2910-2980 cm −1 , which were assigned with the stretching vibrations of the dangling CH2/CH3 groups (Figure 2, Figures S37-S48 in Supplementary Materials).Weak absorbance at around 3440 cm −1 was associated with the stretching vibrations of the COO-H groups of 7-8.Broadening of these bands indicates the hydrogen bond formation, which is correlated well with the analysis of crystal packing of oxy-QPSs.
The FTIR spectrum of the 9-12 has strong absorbance at 1320 and 1160 cm −1 , which was assigned to the stretching symmetric and antisymmetric vibrations of C-O groups.

DSC Measurements and TG Analysis
The melting points (T m ) and decomposition temperatures (T d ) were measured by differential scanning calorimetry and thermogravimetric analysis (Table 3, Figures S49-S60 in Supplementary Materials).One can define studied compounds into three main homologous series.The limited range of available series does not allow us to demonstrate all the trends in melting points and decomposition temperatures; however, some tendencies could be noticed.Commonly, the melting temperature depends on various factors.Both the higher symmetry of the structure and the presence of donor-acceptor groups in the molecule may result in an increase in the lattice energy and, as a consequence, in the rise of the melting and decomposition temperature [42].
Typically, an elongation of the substituent in cation leads to a reduction in the melting point.Our study agrees with this claim (for example, 1 and 3).On the contrary, an additional oxygen atom in the side chain increases the melting temperature in spite of an overall chain length growth ( 1 and 4).This effect is related to the rising interactions between the chains through weak hydrogen bonds driven by an additional oxygen atom [43,44].As expected, replacing the halide anion typically strongly bonding to the cation with a weakly coordinating tetrafluoroborate anion led to a decrease in the melting point.Here, we observe that the melting point of 5 and 6 drops by half compared to 4. Notably, the decomposition temperature increases slightly from 187 to 210 and 221 • C, respectively.The oxy-QPSs with the ester functional group (9-11) melt with decomposition at around 170 • C with a loss of about half their mass.Probably, the same mechanism of thermal decomposition takes place involving the detachment of the acetoether fragment.Compound 12, with a total substituent length of seven atoms, has a phase transition at 125 • C.This is in agreement with the previously observed lengthening effect of the alkyl substituent for sterically hindered phosphonium salts and with the literature data for other organic salts [45], including the calculated data [46].
It is worth mentioning that the DSC curve of 3 has two more phase transitions at temperatures 41 • C and 71 • C, accordingly, prior to complete melting at 109 • C (Figure 3).These phase transformations occur without the loss of mass.Here, we observe that the melting point of 5 and 6 drops by half compared to 4. Notably, the decomposition temperature increases slightly from 187 to 210 and 221 °C, respectively.The oxy-QPSs with the ester functional group (9-11) melt with decomposition at around 170 °С with a loss of about half their mass.Probably, the same mechanism of thermal decomposition takes place involving the detachment of the acetoether fragment.Compound 12, with a total substituent length of seven atoms, has a phase transition at 125 °С.This is in agreement with the previously observed lengthening effect of the alkyl substituent for sterically hindered phosphonium salts and with the literature data for other organic salts [45], including the calculated data [46].
It is worth mentioning that the DSC curve of 3 has two more phase transitions at temperatures 41 °С and 71 °С, accordingly, prior to complete melting at 109 °С (Figure 3).These phase transformations occur without the loss of mass.

Single Crystal X-ray Analysis
Crystals suitable for analysis by single crystal X-ray diffraction were obtained only for some compounds from the presented series.The main crystallographic information for oxy-QPSs is provided in Supplementary Materials.The analysis and comparison of

Single Crystal X-ray Analysis
Crystals suitable for analysis by single crystal X-ray diffraction were obtained only for some compounds from the presented series.The main crystallographic information for oxy-QPSs is provided in Supplementary Materials.The analysis and comparison of the features of molecular and crystal structures were carried out within the three main homologous series, and some general patterns can be noted.
All compounds form monoclinic or orthorhombic crystals without solvent molecules in the crystal lattice and with one cation-anion pair in the independent part of the unit cell.Compound 6 is the only oxy-QPS in the investigated series that exhibits a partial disorder in the linear substituent the cation, as well as possesses the entire anion.Such behavior may be attributed to the nature of the substituent in cation.The introduction of oxygen atoms leads to the flexibility of the side chain of oxy-QPSs and causes a slight change in the conformation in the crystal, even in the case of the same type of cation, for example, in crystals 4 and 6 (Figure 4a).Meanwhile, in the crystals of compounds 7 and 8, the alkyl part of the substituent is in a characteristic zigzag conformation (see Figures S64  and S65 in Supplementary Materials).In the homologous series of compounds 9-11, the conformations of the side chain are fairly very close despite the differences in their length (Figure 4b).S65 in Supplementary Materials).In the homologous series of compounds 9-11, the c formations of the side chain are fairly very close despite the differences in their len (Figure 4b).In the crystals of compounds 7 and 8, an uncharacteristic conformation of the c boxyl group can be noted.The hydrogen atom of the -COOH fragment is in the tr position with respect to the C=O group (see Figures S64-S65 in Supplementary Materia This orientation prevents the formation of a classical centrosymmetric dimer, which characteristic of molecules with a carboxyl fragment.As a result, the classical hydrog bonds of the O-H⋯Br type are realized in both cases. In the example of COOH-functionalized imidazolium ionic liquids, it was shown t in the crystals of hydrophilic ionic liquids, the formation of classic O-H⋯X (X = Hali bonds between the cation and anion of the system is preferred.Classical dimers betw two cations can be realized only for hydrophobic ionic liquids.Nevertheless, in b groups of ionic liquids, the classical cis-conformation of the carboxyl group is realized [4 In our case, another type of conformation of the carboxyl system leads to some shorten of the donor-acceptor distance.The O⋯Br − distances of the O-H⋯Br − hydrogen bonds 3.1124(12) Å and 3.177(2) Å in crystals 7 and 8, respectively.
The crystal structure of oxy-QPSs essentially depends on the type of side chain s stituent and on the type of anion.Specifically, the compounds form non-isostructural cr tals, so no one can observe any trend in the features of crystal packing.For example, co pounds 9-11 with homologically close cations and identical anions are characterized b different three-dimensional lattice and localization of anion (Figure 5).In the crystals of compounds 7 and 8, an uncharacteristic conformation of the carboxyl group can be noted.The hydrogen atom of the -COOH fragment is in the trans position with respect to the C=O group (see Figures S64 and S65 in Supplementary Materials).This orientation prevents the formation of a classical centrosymmetric dimer, which is characteristic of molecules with a carboxyl fragment.As a result, the classical hydrogen bonds of the O-H• • • Br type are realized in both cases.
In the example of COOH-functionalized imidazolium ionic liquids, it was shown that in the crystals of hydrophilic ionic liquids, the formation of classic O-H• • • X (X = Halide) bonds between the cation and anion of the system is preferred.Classical dimers between two cations can be realized only for hydrophobic ionic liquids.Nevertheless, in both groups of ionic liquids, the classical cis-conformation of the carboxyl group is realized [47].In our case, another type of conformation of the carboxyl system leads to some shortening of the donor-acceptor distance.The crystal structure of oxy-QPSs essentially depends on the type of side chain substituent and on the type of anion.Specifically, the compounds form non-isostructural crystals, so no one can observe any trend in the features of crystal packing.For example, compounds 9-11 with homologically close cations and identical anions are characterized by a different three-dimensional lattice and localization of anion (Figure 5).The crystal structure of oxy-QPSs essentially depends on the type of side chain substituent and on the type of anion.Specifically, the compounds form non-isostructural crystals, so no one can observe any trend in the features of crystal packing.For example, compounds 9-11 with homologically close cations and identical anions are characterized by a different three-dimensional lattice and localization of anion (Figure 5).

Microcrystalline Cellulose Dissolution
Recently, lignocellulosic biomass treatment has gained a reputation as a key process of near-future chemical technology [49,50].Biomass recycling provides access to renewable feedstock for energy, chemicals, and advanced materials [51][52][53].Polysaccharides are the major structural component of plant cell walls.The challenging step of the recycling process is biomass pretreatment due to the poor solubility of natural polysaccharides in water and organic solvents [54,55].The most common carbohydrate polymers are cellulose, chitin, chitosan, and starch.The ILs (organic salts in a broad sense) have found an application in the field of cellulose and other polysaccharides' dissolution, and a number of advances have been made [56,57].
Among effective ILs in biomass pretreatment are nitrogen-containing heterocycles and ammonium salts with oxygen-containing functional groups [58][59][60].However, the high efficiency in cellulose dissolution has been demonstrated by the phosphonium salts as well [24,61].Recent research has highlighted a number of structure-capacity patterns for effective cellulose dissolution in organic salts, which have been emphasized [62,63].Regardless, the strictly defined structural motives for IL design in this area do not exist.
In the first series of experiments, we used phosphonium salts under the study to dissolve microcrystalline cellulose.As oxy-QPSs are solids at room temperature and melt above 100 • C, a co-solvent was used to reduce the viscosity.Mixtures of ILs and organic solvents are extensively used for biomass pretreatment [64].The co-solvent also acts to decrease the cost of the dissolution process.The most popular of them is dimethyl sulfoxide (DMSO).It is an aprotic, inexpensive, and non-volatile organic solvent that is obtained as a by-product of the pulping industry in lignin production [65].DMSO is readily miscible (1:1 by mass) with most of the oxy-QPSs under study.DMSO has no significant influence on the specific interactions between organic salts and dissolved cellulose [66].Stirring of 1 wt% microcrystalline cellulose (50 µm) in pure DMSO at 80 • C for 17 h did not lead to the dissolution of the biopolymer (Figure 6).
Microcrystalline cellulose of 50 µm (1 wt%), the mixture of corresponding oxy-QPSs and DMSO (1:1 by mass), was placed in a 5 mL Duran test tube with a magnetic stirrer and tightly screwed cap.The mixture was stirred at 800 rpm at 80 • C. At certain time intervals, a drop of the mixture was picked and placed on a glass slide, and the solubility process was analyzed under an optical microscope at 40× magnification (Table 4).
decrease the cost of the dissolution process.The most popular of them is dimethyl sulfoxide (DMSO).It is an aprotic, inexpensive, and non-volatile organic solvent that is obtained as a by-product of the pulping industry in lignin production [65].DMSO is readily miscible (1:1 by mass) with most of the oxy-QPSs under study.DMSO has no significant influence on the specific interactions between organic salts and dissolved cellulose [66].Stirring of 1 wt% microcrystalline cellulose (50 µm) in pure DMSO at 80 °C for 17 h did not lead to the dissolution of the biopolymer (Figure 6).Microcrystalline cellulose of 50 µm (1 wt%), the mixture of corresponding oxy-QPSs and DMSO (1:1 by mass), was placed in a 5 mL Duran test tube with a magnetic stirrer and tightly screwed cap.The mixture was stirred at 800 rpm at 80 °С.At certain time intervals, a drop of the mixture was picked and placed on a glass slide, and the solubility process was analyzed under an optical microscope at 40× magnification (Table 4).During the experiment, the cellulose fibers split into smaller ones, then visible particles disappeared, and a homogeneous solution was observed (Figure 7).During the experiment, the cellulose fibers split into smaller ones, then visible particles disappeared, and a homogeneous solution was observed (Figure 7).The mixtures of 1 and 4 with DMSO partly dissolve cellulose fibers during heating till 80 °С.After stirring for 5 min at 80 °С, the solution became clear with no visible particles of cellulose (Figure 8).The mixture of 3 and DMSO dissolves cellulose slower than two other oxy-QPSs with the ether functional group and halide anion.Polymer fibers disappear after 90 min of heating (Figure 7).However, at this time, oxy-QPS 3 is partly destructed, confirmed by the 1 H NMR spectrum.The destruction occurs with the loss of one of four substituents, and tertiary phosphonium salt is formed.We suppose that compounds 1 and 4, with their similar structure, possess chloride anions that probably make the molecule more stable under process conditions due to the small size of the anion and stronger ionic binding.At 80 • C, the phosphonium compounds 9 and 11 are not fully miscible with DMSO, preventing cellulose dissolution.The heating was terminated after 4 h.The destruction of 10 and 12 is detected by the NMR method after several hours of heating, although some splitting of the cellulose fibers is observed.The mixture of 5 and DMSO dissolves cellulose slowly; probably, the appearance of tetrafluoroborate-anion in oxy-QPS structure prevents the formation of hydrogen bonds with cellulose molecules.
In the next step of the study, oxy-QPSs with the ether group and halide anion were introduced into the dissolution process at a reduced temperature (Table 5).The temperature decrease to 60 • C became the breaking point when the cellulose fibers showed no visible dissolution for up to 8 h.In the example of 3, one can see gradual moderation in the time of cellulose splitting and disappearing.Oxy-QPS 4 appeared not to be fully miscible with DMSO at 60 • C, which influenced its solubilizing properties on cellulose.The distinctive feature made 4 the solvent of choice for carrying out a number of experiments with the aim of optimizing the process of cellulose dissolution.The first enhancement was made to reduce the ratio of the oxy-QPSs to DMSO.When the proportion of oxy-QPSs to DMSO was 1:4, the time of cellulose dissolution increased from 5 min to 4 h at 70 • C.
The same effect of elongation of dissolution time was observed when DMSO was substituted with water.An amount of 1 wt% of cellulose and the mixture of 4 and water (1:1) was heated to 70 • C, and the complete dissolution of polymer fibers was observed after 4 h of stirring.
Due to the high solubility of 4 in water, a three-component system was tested for cellulose dissolution.The mixture of 4, DMSO, and water (2:1:1) with 1 wt% of cellulose were stirred at 60 • C. In contrast to the two-component mixture of 4 and DMSO, the oxy-QPS was fully miscible with co-solvents when half of DMSO was replaced by water.However, after 8 h of stirring, cellulose fibers had not been dissolved in the mixture.
Taking into account the ready dissolution of cellulose in the mixtures of 1 and 4 with DMSO at 80 • C, the enlarged loading of cellulose was tested.Each of the two mixtures had soluted up to 5 wt% of cellulose for around 30 min.It is noteworthy that the resulting solution of cellulose in 4 + DMSO is less viscous than in 1 + DMSO.
The best solvent for cellulose among studied oxy-QPSs is the mixtures 4 + DMSO and 1 + DMSO.Both 1 and 4 possess ether functional groups in cation and Cl-anion.The chloride anion, due to its small size, acts as a hydrogen acceptor, facilitating the dissolution of cellulose.The oxygen atom in 1 and 4 is structurally tethered in the vicinity of the charged phosphorus closer than in other oxy-QPSs.
Hydrogen bonds play a very important role in processes where ionic liquids are used as solvents.To date, numerous studies have indicated that hydrogen bonding formation between anions and the -OH group of cellulose is the main driving force for cellulose dissolution in ILs [58][59][60]64].The geometrical neighborhood of the charged head group could have an electronic impact on H-bonding [67].Indirect evidence of the strong influence of functional groups on the possibility of changing the strength of hydrogen interactions can be changes in chemical shifts in 1 H NMR spectra.This is exactly what we see in the case of compounds 1 and 4, where the protons of the CH 2 fragments "tear" between the positive charged phosphonium cation and the electronegative oxygen atom, which leads to a shift of signals of corresponding protons to the region of 5 ppm.Of course, one cannot discount the small chloride anion, which makes a significant contribution to the interaction with the bare nuclei of protons [68].Thus, in phosphonium salts 1 and 4, we obtained the necessary combination of critical structural components for the formation of strong hydrogen bonds, which allows us to successfully dissolve cellulose.

Antimicrobial Activity of oxy-QPSs
Despite spreading opinion, not all ILs are toxic in the same way [43,57], although it is generally known that phosphonium salts have quite high antibacterial activity [69], which is reflected in their use to create an antibacterial coating for plastic surfaces [70] or even create polymers with antibacterial properties [71].At the first approximation, the biological activity of oxy-QPSs was estimated on several pathogenic bacteria and fungi as the model systems.The phosphonium salts under study were tested for antimicrobial activity against a number of Gram-positive Staphylococcus aureus (Sa), Bacillus cereus (Bc), and Enterococcus faecalis (Ef ) and Gram-negative Escherichia coli (Ec) bacteria.Antifungal activity was studied on a culture of a yeast-like fungus, Candida albicans (Ca).The results on antimicrobial activity of oxy-QPSs are presented in Table 6.
Based on the data obtained, one can conclude that the tested compounds have a selective effect on the Gram-positive bacteria S. aureus.The most effective compounds were 7 and 10, which showed antimicrobial activity at the level of the reference drug chloramphenicol.Chloramphenicol inhibits bacterial protein synthesis by interfering with the 'transfer' of the elongated peptide chain to the newly attached aminoacyl t-RNA at the ribosome-RNA complex.It binds specifically to the 50 S ribosome and may prevent the aminoacyl t-RNA from accessing the acceptor site for amino acid incorporation.Chloramphenicol probably acts as a peptide analog, preventing the formation of peptide bonds.At high doses, it can also inhibit mammalian mitochondrial protein synthesis.Marrow cells are particularly susceptible [72][73][74][75].It was also shown that 7 and 10 have weak bactericidal activity (the difference between MIC and MBC did not exceed 4) against S. aureus 209p, in contrast to chloramphenicol, which is a bacteriostatic drug.The test compounds were inactive against Bacillus cereus, Enterococcus faecalis, Escherichia coli, and Candida albicans.
The fermentation process could be included in a one-pot biomass treatment or follow the dissolution procedure in a stage-by-stage process.Both of them require the usage of non-toxic solvents due to the microorganisms that are utilized during the fermentation.The biocompatibility of ILs is a serious problem for the choice of an effective biomass solvent [76].The tolerance study of potential solvents against fermented microbes should be an essential procedure [77].
The biocompatibility of the oxy-QPSs with enzymatic microorganisms was tested against non-pathogenic microbes that could be used in the fermentation process (Table S1 in Supplementary Materials): Saccharomyces cerevisiae 1986 (Sc), Lactobacillus plantarum 8P-A3 (Lp), and Bacillus subtilis ATCC 6633 (Bs).The obtained results demonstrated low toxicity of oxy-QPSs against all non-pathogenic microorganisms.All tested compounds were absolutely inactive (MIC > 500 µg/mL) against Saccharomyces cerevisiae and Lactobacillus plantarum.Compounds 1, 7, and 9 showed negligible activity against Bacillus subtilis bacteria compared to the antibiotic amoxicillin.Their MIC manifested in the range of 250-500 µg/mL.
The oxy-QPSs were shown to be a suitable environment for bacteria and fungi and could be safely used in a biomass pretreatment process followed by fermentation.It would be safe to assume that IL utilization prevents the formation of inhibitors for the growth of microorganisms that are capable of enzymatic biomass degradation.

Cytotoxicity of oxy-QPSs
The impact of ILs on living cells is an important challenge to explore in the development of the strategy for their industrial application [78].As a part of the study of the safety of compounds for humans, the hemolytic and cytotoxic activities of oxy-QPSs were evaluated.The ability of the tested compounds to destroy human erythrocytes illustrates the toxic effect on the internal environment of the body.The hemolysis assay is a simple screening test that can represent the cytotoxicity in more complex models [79].Such ex-perimental models can be cell lines derived from different human organs and tissues and provide an adequate assessment of the effect of the compounds on cell metabolism.
The cytotoxicity of oxy-QPSs was studied using human erythrocytes, normal liver cell line Chang liver, and human embryo lung WI38.Within the study, the values of HC 50 , the concentration causing hemolysis of 50% of erythrocytes, and IC 50 , the concentration causing the death of 50% of human liver cells, were determined (Table 7).According to the test results, all tested oxy-QPSs showed no hemolytic activity and low toxicity to normal human cell lines, which characterizes their safety toward mammalian cells.

Ecotoxicity of oxy-QPSs
The industrial use of a new solvent requires the checking of its safety.Modern solvents for natural polysaccharides should be safe, non-toxic, and nonhazardous to humans and the environment [76].
The planktonic crustacean Daphnia magna is a widespread water animal that possesses known sensitivity to the slightest environmental change and, therefore, is a convenient organism in ecotoxicological research.One of the factors that profoundly affects the survival rate of D. magna is environmental pollution.
The acute aquatic toxicity of oxy-QPSs was determined using a D. magna population.A 48 h test was performed with selected water solutions of oxy-QPSs at different concentrations (Table S2 in Supplementary Materials).Each test involved 30 mature animals.The compounds with shorter substituents (1 and 7) could be classified as nonhazardous according to the Globally Harmonized System of Classification and Labelling of Chemicals.The oxy-QPSs 4-5 and 11-12 are referred to in the category Acute 3.

Conclusions
We synthesized new quaternary phosphonium salts with oxygen-functionalized groups in sterically hindered cations.The NMR, FTIR, and TG/DSK study and crystal structure showed that trends in properties of oxy-QPSs are, to some extent, similar to those for alkyl-QPSs.However, the introduction of the oxygen atom in the side chain is responsible for the hydrogen bond formation, which causes some novel tendencies in properties, such as melting point or chemical shift in NMR spectra.
The structural features were found to determine the ability of oxy-QPSs to dissolve microcrystalline cellulose.The ether group in the cation and chloride anion was found to be necessary for effective cellulose dissolution.Notably, the oxygen atom in the side chain is preferred to be no more than one carbon atom across from phosphorus.This is attributed to the acidity of the α-protons and the strong hydrogen bond formation.The conditions of the dissolution process, such as temperature, time, co-solvent, cellulose loading, and solvent ratio, were selected.Pretreatment with ether-containing phosphonium salts and DMSO co-solvent (1:1) results in the dissolution of up to 5 wt% of microcrystalline cellulose at 80 • C for 30 min.The use of 4 allows DMSO to be replaced by water for cellulose dissolution but with a significant increase in process time.
The antimicrobial activity of the oxy-QPSs under investigation was expected to be low due to the short length of substituents in the cation.The studied compounds are sufficiently safe for the non-pathogenic microbes that can stand as fermenting ones in a one-pot biomass treatment process.The current study has also shown the low toxicity of oxy-QPSs to the human cell lines and to the crustacean D. magna.The present work can be used to guide the design of new task-specific phosphonium salts.

Figure 5 .
Figure 5.The fragments of crystal packing of oxy-QPSs from left to right: 9 (view along the 0b axis), 10 (view along the 0a axis), and 11 (view along the 0a axis).The Mercury program suite [48] was used for molecular graphics.

in 13 C NMR Spectra (ppm)
* The chemical shift in non-deuterated acetone is given in brackets.

Table 3 .
Summary of melting points T m (by DSC) and decomposition temperatures T d (by dynamic TGA) for oxy-QPSs.

Table 4 .
The time of cellulose dissolution in oxy-QPSs.

Table 4 .
The time of cellulose dissolution in oxy-QPSs.

Table 5 .
The time of cellulose dissolution at varied temperatures.
* The experiment was repeated three times.
* The experiment was repeated three times.