Investigation of ibuprofen drug using mass spectrometry,thermal analyses,and semi-empirical molecular orbital calculation

Ibuprofen (C₁₅H₁₈O₂) is an anti-inflammatory drug. It is important to investigate its structure to know the active groups and weak bond responsible for its medical activity. Consequently in the present study, ibuprofen was investigated by mass spectrometry (MS) and thermal analyses (TAs) (TG/DTG and DTA), and confirmed by semi-empirical molecular orbital (MO) calculation using PM3 procedure, on the neutral and positively charged forms of the drug. These calculations included bond order, bond length, and bond strain, and charge distribution, heat of formation, and ionization energy. The mass spectra and thermal analysis fragmentation pathways were proposed and compared to each other to select the most suitable scheme representing the correct fragmentation pathway of the drug in both techniques. From the electron ionization (EI) mass spectra, the primary cleavage site of the charged molecule is because of the rupture of COOH group (the lowest bond order) followed by propyl group loss. The TAs of the drug revealed high response of the drug to the temperature variation with very fast rate. It decomposed in several sequential steps in the temperature range 25–360 °C. The initial thermal decomposition is similar to that obtained by MS fragmentation of the first rupture (COOH), then subsequent one of propyl loss, and finally of ethylene loss. These mass losses appear as endothermic peaks required energy values of −214.83, −895.95, and −211.10 J g⁻¹, respectively. The order of these losses is also related to the values of the MO calculation parameters. Therefore, the comparison between MS and TA helps in the selection of the proper pathway representing the decomposition of this drug to give its metabolites in in vivo system. This comparison is also successfully confirmed by MO calculations.     

Structure characterization and spectroscopic investigation of ciprofloxacin drug

Ciprofloxacin (CPF, C₁₇H₁₈FN₃O₃) drug is used in the treatment of some bacterial infectious diseases. The drug was investigated using thermal analysis (TA) measurements (TG/DTG) and electron impact mass spectral (EI-MS) fragmentation at 70 eV techniques. Furthermore, the drug was characterized and investigated by other spectroscopic tools as IR, UV–Vis, ¹H-, and ¹³C-NMR. Semi-empirical MO calculation using PM3 procedure has been carried out on neutral molecule and positively charged species. The calculations included, bond length, bond order, bond strain, partial charge distribution, ionization energy, and heat of formation (ΔH _f). The PM3 procedure provides a basis for fine distinction among sites of initial bond cleavage, which is crucial to the rationalization of subsequent fragmentation of the molecule. The mass spectra and thermal analysis fragmentation pathways were proposed and compared to each other to select the most suitable scheme representing the correct fragmentation of this drug. From EI-MS, the main primary cleavage site of the charged molecule is that due to C–COOH bond cleavage with H-rearrangement to skeleton and CO₂ loss which can further decompose by piperazine loss. Thermal analysis of the neutral form of the drug reveals the high response of the drug to the temperature variation with very fast rate. Thermal decomposition has carried out in several sequential steps in the temperature range 40–650 °C. The initial thermal decomposition is similar to that obtained by mass spectrometric fragmentation (C–COOH fragment) but differ in that a rearrangement occurs by OH and CO loss. Therefore, comparison between MS and TA helps in selection the proper pathway representing the fragmentation of this drug. This comparison successfully confirmed by MO calculation. Finally, the effect of fluorine atom on the stability of the drug was discussed.     

Kinetic study of crystallization of PHB in presence of hydroxy acids

Poly(3-hydroxybutyrate), PHB, has been structurally modified through reaction with hydroxy acids (HA) such as tartaric acid (TA) and malic acid (MA). The crystallization kinetic of the samples was evaluated by isoconversional method through nonlinear fitting to obtain the estimation for activation energy (E _a ) and pre-exponential (A) values. The thermal behavior of the crystallization temperature, 44.8 and 58.9 °C at 5 °C/min, and results obtained to the average activation energy, 73 ± 9 kJ mol⁻¹ and 63 ± 1 kJ mol⁻¹, to PHB/MA and PHB, respectively, are suggesting that malic acid may be deriving plasticizer units from its own PHB chain. PHB/TA show increase in the medium value of E _a, 119 ± 2 kJ mol⁻¹ and T _c = 48.2 °C (at 5 °C/min), indicating that tartaric acid is probably interacts in different way to the of PHB chain (E _a=73 ± 9 kJ mol⁻¹, T _c = 44.8 °C at 5 °C/min).     

Thermal behaviour of cephalexin in different mixtures

The thermoanalytical curves (TA), i.e. TG, DTG and DTA for pure cephalexin and its mixtures with talc, magnesium stearate, starch and microcrystalline cellulose, respectively, were drawn up in air and nitrogen at a heating rate of 10 °C min⁻¹. The thermal degradation was discussed on the basis of EGA data obtained for a heating rate of 20 °C min⁻¹. Until 250 °C, the TA curves are similar for all mixtures, up this some peculiarities depending on the additive appears. These certify that between the pure cephalosporin and the excipients do not exists any interaction until 250 °C. A kinetic analysis was performed using the TG/DTG data in air for the first step of cephalexin decomposition at four heating rates: 5, 7, 10 and 12 °C min⁻¹. The data processing strategy was based on a differential method (Friedman), an integral method (Flynn–Wall–Ozawa) and a nonparametric kinetic method (NPK). This last one allowed an intrinsic separation of the temperature, respective conversion dependence on the reaction rate and less speculative discussions on the kinetic model. All there methods had furnished very near values of the activation energy, this being an argument for a single thermooxidative degradation at the beginning (192–200 °C).     

Compatibility and decomposition kinetics studies of prednicarbate alone and associated with glyceryl stearate

In this work, TG/DTG and DSC techniques were used to the determination of thermal behavior of prednicarbate alone and associated with glyceryl stearate excipient (1:1 physical mixture). TG/DTG curves obtained for the binary mixture showed a reduction of approximately 37 °C to the thermal stability of drug ( T_{\textdm/\textdt = 0 \textDTG}^\textMax T_{{{\text{d}}m/{\text{d}}t = 0\,{\text{DTG}}}}^{\text{Max}} ). The disappearance of stretching band at 1280 cm⁻¹ (ν_as C–O, carbonate group) and the presence of streching band with less intensity at 1750 cm⁻¹ (ν_s C–O, ester group) in IR spectrum obtained to the binary mixture submitted at 220 °C, when compared with IR spectrum of drug submitted to the same temperature, confirmed the chemical interaction between these substances due to heating. Kinetics parameters of decomposition reaction of prednicarbate were obtained using isothermal (Arrhenius equation) and non-isothermal (Ozawa) methods. The reduction of approximately 45% of activation energy value (E _a) to the first step of thermal decomposition reaction of drug in the 1:1 (mass/mass) physical mixture was observed by both kinetics methods.     

Thermal characteristics of bitumen pyrolysis

The pyrolysis behavior of bitumen was investigated using a thermogravimetric analyzer–mass spectrometer system (TG–MS) and a differential scanning calorimeter (DSC) as well as a pyrolysis-gas chromatograph/mass spectrometer system (Py-GC/MS). TG results showed that there were three stages of weight loss during pyrolysis—less than 110, 110–380, and 380–600 °C. Using distributed activation energy model, the average activation energy of the thermal decomposition of bitumen was calculated at 79 kJ mol⁻¹. The evolved gas from the pyrolysis showed that organic species, such as alkane and alkene fragments had a peak maximum temperature of 130 and 480 °C, respectively. Benzene, toluene, and styrene released at 100 and 420 °C. Most of the inorganic compounds, such as H₂, H₂S, COS, and SO₂, released at about 380 °C while the CO₂ had the maximum temperature peaks at 400 and 540 °C, respectively. FTIR spectra were taken of the residues of the different stages, and the results showed that the C–H bond intensity decreased dramatically at 380 °C. Py-GC/MS confirmed the composition of the evolved gas. The DSC revealed the endothermic nature of the bitumen pyrolysis.     

Effect of particle size on pyrolysis characteristics of Elbistan lignite

In this study, the relationship between particle size and pyrolysis characteristics of Elbistan lignite was examined by using the thermogravimetric (TG/DTG) and differential thermal analysis (DTA) techniques. Lignite samples were separated into different size fractions. Experiments were conducted at non-isothermal conditions with a heating rate of 10°C min⁻¹ under nitrogen atmosphere up to 900°C. Pyrolysis regions, maximum pyrolysis rates and characteristic peak temperatures were determined from TG/DTG curves. Thermogravimetric data were analyzed by a reaction rate model assuming first-order kinetics. Apparent activation energy (E) and Arrhenius constant (A _r) of pyrolysis reaction of each particle size fraction were evaluated by applying Arrhenius kinetic model. The apparent activation energies in the essential pyrolysis region were calculated as 27.36 and 28.81 kJ mol⁻¹ for the largest (−2360+2000 μm) and finest (−38 μm) particle sizes, respectively.     

Low-temperature behavior of Li3V2(PO4)3/C as cathode material for lithium ion batteries

The electrochemical performance of Li₃V₂(PO₄)₃/C was investigated at various low temperatures in the electrolyte 1.0 mol dm⁻³ LiPF₆/ethyl carbonate (EC)+diethyl carbonate (DEC)+dimethyl carbonate (DMC) (volume ratio 1:1:1). The stable specific discharge capacity is 125.4, 122.6, 119.3, 116.6, 111.4, and 105.7 mAh g⁻¹ at 26, 10, 0, −10, −20, and −30 °C, respectively, in the voltage range of 2.3–4.5 V at 0.2 C rate. When the temperature decreases from −30 to −40 °C, there is a rapid decline in the capacity from 105.7 to 69.5 mAh g⁻¹, implying that there is a nonlinear relationship between the performance and temperature. With temperature decreasing, R _ct (corresponding to charge transfer resistance) increases rapidly, D (the lithium ion diffusion coefficients) decreases sharply, and the performance of electrolyte degenerates obviously, illustrating that the low-temperature electrochemical performance of Li₃V₂(PO₄)₃/C is mainly limited by R _ct, D _Li, and electrolyte.     

Mechanistic studies on the intramolecular electron transfer in an adduct species of the oxo-centred trinuclear iron(III) cation and <Emphasis Type="SmallCaps">l</Emphasis>-ascorbic acid in aqueous solution

The kinetics of the intra-molecular electron transfer of an adduct of l-ascorbic acid and the [Fe₃^IIIO(CH₃COO)₆(H₂O)₃]⁺ cation in aqueous acetate buffer was studied spectrophotometrically, over the ranges 2.55 ≤ pH ≤ 3.74, 20.0 ≤ θ ≤ 35.0 °C, at an ionic strength of 0.50 and 1.0 mol dm⁻³ (NaClO₄). The reaction of l-ascorbic acid and the complex cation involves the rapid formation of an adduct species followed by a slower reduction in the iron centres through consecutive one-electron transfer processes. The final product of the reaction is aqueous iron(II) in acetate buffer. The proposed mechanism involves the triaqua and diaqua-hydroxo species of the complex cation, both of which form adducts with l-ascorbic acid. At 25 °C, the equilibrium constant for the adduct formation was found to be 86 ± 15 and 5.8 ± 0.2 dm³ mol⁻¹ for the triaqua and diaqua-hydroxo species, respectively. The kinetic parameters derived from the rate expression have been found to be: k ₀ = (1.12 ± 0.02) × 10⁻² s⁻¹ for the combined spontaneous decomposition and k ₁ = (4.47 ± 0.06) × 10⁻² s⁻¹ (ΔH ₁^‡ = 51.0 ± 2.3 kJ mol⁻¹, ΔS ₁^‡ = −100 ± 8 J K⁻¹ mol⁻¹), k ₂ = (4.79 ± 0.38) × 10⁻¹ s⁻¹ (ΔH ₂^‡ = 76.5 ± 0.8 kJ mol⁻¹, ΔS ₂^‡ = 6 ± 3 J K⁻¹ mol⁻¹) for the triaqua and diaqua-hydoxo species, respectively.     

Thermal characterization of starch-based polymers produced by <Emphasis Type="Italic">Ophiostoma</Emphasis> spp

The thermal behavior of modified starches (MS) produced by biosynthetic pathway is described based on a comparative analysis with native starches (NS). MS were produced by fermentation in presence of Ophiostoma spp. cultures. Thermogravimetric analysis (TG) with successive derivatives (DTG) and differential scanning calorimetry (DSC) were used for this study. NS results showed a single peak dominating both the TG (DTG) and DSC plots. A double thermal transition event was detected in samples of MS. The procedural decomposition temperature (T _i–T _f; lowest onset temperature of initial and final mass change) was carried out within a narrow interval of temperatures for NS (610–640 °C). This interval could not be reached within the 1,000 °C range in MS. Residues higher than 10% were recorded for MS at this temperature. The presence of the double thermal transition in MS is discussed.     

Thermal behavior and thermal safety on 3,3-dinitroazetidinium salt of perchloric acid

3,3-Dinitroazetidinium (DNAZ) salt of perchloric acid (DNAZ·HClO₄) was prepared, it was characterized by the elemental analysis, IR, NMR, and a X-ray diffractometer. The thermal behavior and decomposition reaction kinetics of DNAZ·HClO₄ were investigated under a non-isothermal condition by DSC and TG/DTG techniques. The results show that the thermal decomposition process of DNAZ·HClO₄ has two mass loss stages. The kinetic model function in differential form, the value of apparent activation energy (E _a) and pre-exponential factor (A) of the exothermic decomposition reaction of DNAZ·HClO₄ are f(α) = (1 − α)⁻¹/2, 156.47 kJ mol⁻¹, and 10^15.12 s⁻¹, respectively. The critical temperature of thermal explosion is 188.5 °C. The values of ΔS ^≠, ΔH ^≠, and ΔG ^≠of this reaction are 42.26 J mol⁻¹ K⁻¹, 154.44 kJ mol⁻¹, and 135.42 kJ mol⁻¹, respectively. The specific heat capacity of DNAZ·HClO₄ was determined with a continuous C _p mode of microcalorimeter. Using the relationship between C _p and T and the thermal decomposition parameters, the time of the thermal decomposition from initiation to thermal explosion (adiabatic time-to-explosion) was evaluated as 14.2 s.     

Synthesis and electrochemical characterization of aPEO-based polymer electrolytes

In this paper, the preparation and purification of an amorphous polymer network, poly[oxymethylene-oligo(oxyethylene)], designated as aPEO, are described. The flexible CH₂CH₂O segments in this host polymer combine appropriate mechanical properties, over a critical temperature range from −20 to 60 °C, with labile salt-host interactions. The intensity of these interactions is sufficient to permit solubilisation of the guest salt in the host polymer while permitting adequate mobility of ionic guest species. We also report the preparation and characterisation of a novel polymer electrolyte based on this host polymer with lithium tetrafluoroborate, LiBF₄, as guest salt. Electrolyte samples are thermally stable up to approximately 250 °C and completely amorphous above room temperature. The electrolyte composition determines the glass transition temperature of electrolytes and was found to vary between −50.8 and −62.4 °C. The electrolyte composition that supports the maximum room temperature conductivity of this electrolyte system is n = 5 (2.10 × 10⁻⁵ S cm⁻¹ at 25 °C). The electrochemical stability domain of the sample with n = 5 spans about 5 V measured against a Li/Li⁺ reference. This new electrolyte system represents a promising alternative to LiCF₃SO₃ and LiClO₄-doped PEO analogues.     

A kinetic investigation of the oxidation of <Emphasis Type="SmallCaps">dl</Emphasis>-pipecolinate by bis(hydrogenperiodato)argentate(III) complex anion

Kinetics of oxidation of dl-pipecolinate by bis(hydrogenperiodato)argentate(III) complex anion, [Ag(HIO₆)₂]⁵⁻, has been studied in aqueous alkaline medium in the temperature range of 25–40 °C. The oxidation kinetics is first order in the silver(III) and pipecolinate concentrations. The observed second-order rate constant, decreasing with increasing [periodate] is virtually independent of [OH⁻]. α-Aminoadipate as the major oxidation product of pipecolinate has been identified by chromatographic analysis. A reaction mechanism is proposed that involves a pre-equilibrium between [Ag(HIO₆)₂]⁵⁻ and [Ag(HIO₆)(H₂O)(OH)]²⁻, a mono-periodate coordinated silver(III) complex. Both Ag(III) complexes are reduced in parallel by pipecolinate in rate-determining steps (described by k ₁ for the former Ag(III) species and k ₂ for the latter). The determined rate constants and their associated activation parameters are k ₁ (25 °C) = 0.40 ± 0.02 M⁻¹ s⁻¹, ∆H ₁^≠ = 53 ± 2 kJ mol⁻¹, ∆S ₁^≠ = −74 ± 5 J K⁻¹ mol⁻¹ and k ₂ (25 °C) = 0.64 ± 0.02 M⁻¹ s⁻¹, ∆H ₂^≠ = 41 ± 2 kJ mol⁻¹, ∆S ₂^≠ = −110 ± 5 J K⁻¹ mol⁻¹. The time-resolved spectra, a positive dependence of the rate constants on ionic strength of the reaction medium, and the consistency of pre-equilibrium constants derived from different reaction systems support the proposed reaction mechanism.     

Effects of strontium silicate on structure and magnetic properties of electrospun strontium ferrite nanofibers

The composite nanofibers of xSrSiO₃/(100 − x)SrFe₁₂O₁₉ (x = 0–13 wt%) with diameters around 110 nm have been prepared by calcination of the electrospun SrSiO₃/SrFe₁₂O₁₉/poly (vinyl pyrrolidone) (PVP) composite fibers at 800–900 °C. The composite nanofibers were characterized by Fourier transform infrared spectroscopy, X-ray diffraction, scanning electron microscopy and vibrating sample magnetometer. After calcined at 800° the M-type strontium ferrite is formed and the strontium silicate exists as an amorphous state when the calcination temperature below about 950 °C. The addition of SrSiO₃ has an obvious suppression effect on the strontium ferrite grain growth and the ferrite grain size decreases from 66.9 to 33.5 nm corresponding SrSiO₃ content from 0 to 9 wt% in the composite. The specific saturation magnetization (Ms) of the xSrSiO₃/(100 − x)SrFe₁₂O₁₉ composite nanofibers exhibits a continuous reduction from 58.0 to 45.6 A m² kg⁻¹ with the increase of SrSiO₃ content from 0 to 13 wt%. With addition of SrSiO₃ from 0 to 13 wt%, the coercivity of the composite nanofibers obtained at 900 °C initially increases, reaching a maximum value 501.1 kA m⁻¹ at the silicate content 7 wt%, and then shows a reduction tendency with the strontium silicate content increase further up to 13 wt%. This influence on the coercivity by strontium silicate can be attributed mainly to the ferrite grain growth suppression and the non-magnetic phase barrier for the domains misalignment.     

Kinetic determination of silver at trace level based on its catalytic effect on a ligand substitution reaction

It is observed that Ag(I) catalyzes the rate of substitution of phenylhydrazine (PhNHNH₂) into hexacyanoferrate(II), producing a cherry red colored complex, [Fe(CN)₅PhNHNH₂]³⁻. The reaction was monitored at 488 nm leading to the formation of the complex under the conditions: [Fe(CN)₆]⁴⁻ (5.0 × 10⁻³ mol dm⁻³), PhNHNH₂ (2.0 × 10⁻³ mol dm⁻³), temperature (25 ± 0.1 °C), pH (2.8 ± 0.02), and ionic strength, I (0.02 mol dm⁻³), (KNO₃). Under optimum conditions, absorbance at fixed times (A _t ) is linearly related to Ag(I) in the concentration range 10.79–97.08 ng cm⁻³, in the presence of several diverse ions. The highest percentage error and relative standard deviations in the entire range of Ag(I) determination are found to be 2.5% and 0.16, with a detection limit of 8.75 ng cm⁻³ of silver(I). The experimental accuracies expressed in terms of percentage recoveries are in the range of 97.87–102.50. The method was successfully applied for the determination of Ag(I) in a few synthetic samples and found to be in good agreement with those obtained from atomic absorption spectrophotometry (AAS). The validity of the proposed method has also been tested for Ag(I) determination in spiked drinking water samples. The present catalytic kinetic method (CKM) is highly sensitive, selective, reproducible, and inexpensive. A review of recently published catalytic spectrophotometric methods for determination of Ag(I) has also been presented for comparison.     

Kinetics and mechanism of interaction of hexaaquachromium(III) with <Emphasis Type="SmallCaps">l</Emphasis>-Dopa in aqueous medium: comparative antiparkinsonian studies

The kinetics and mechanism of the substitution reaction between [Cr(H₂O)₆]³⁺ and l-Dopa in aqueous medium has been studied over the range 1.8 ≤ pH ≤ 2.6, 1.68 × 10⁻² mol dm⁻³ ≤ [Dopa] ≤ 5.04 × 10⁻² mol dm⁻³, I = 0.1 mol dm⁻³ (KNO₃) at 50 °C. The reaction takes place via an outer sphere association between Cr³⁺ and l-Dopa followed by chelation. The product was characterized by physicochemical and infrared spectroscopic methods. The antiparkinsonian activity of the product was found to be higher than that of l-Dopa.     

Optimization on fabrication and performance of A-site-deficient La0.58Sr0.4Co0.2Fe0.8O3-δ cathode for SOFC

A-site-deficient perovskite cathode material La_0.58Sr_0.4Co_0.2Fe_0.8O_3 − δ (L58SCF) is coated on the yttria-stabilized zirconia electrolyte by screen-printing technique. Several key fabrication parameters including selection of additives (binder and pore former), effect of coating thickness, sintering temperature and time on the microstructure, and electrochemical performance of cathode are investigated by scanning electron microscopy and electrochemical impedance spectroscopy. We study the microstructure and the electrochemical property of the cathode with different kinds of additives. Results show that the cathode possesses fine microstructure, enough porosity, and ideal electrochemical property when polyvinyl butyral serves as both binder and pore former in the cathode. The cathode with three screen-printing coats (thickness 28 ± 7 μm, weight 6.07 ± 0.72 mg cm⁻²) sintering at 1,000 °C for 2 h shows lower polarization resistance of 0.183 Ω cm² at 800 °C. Based on the optimized parameters, the polarization resistances of the L58SCF–Ce_0.8Gd_0.2O_{1.9 – δ} composite cathode display the R _p values of 0.067 Ω cm² at 800 °C, 0.106 Ω cm² at 750 °C, 0.225 Ω cm² at 700 °C, and 0.550 Ω cm² at 650 °C.     

On Water Structure in Concentrated Salt Solutions

In concentrated salt solutions the average distances between the ions, d ^av=1.1844⋅(∑ν _i c _i )^−1/3 nm, are commensurate with the sizes of the solvated ions, so that no ‘bulk solvent’ remains. This is illustrated with two saturated aqueous solutions, where 16.67 mol⋅dm⁻³ CsF at 75 °C has d ^av(Cs–F)=0.368 nm and 14.54 mol⋅dm⁻³ LiI at 80 °C has d ^av(Li–I)=0.385 nm. The minimal distance required for the bare ions (sum of their radii) are 0.303 nm for CsF and 0.289 nm for LiI. Hence no water molecule, diameter 0.276 nm, can be fitted between the ions to form linear or slightly bent hydrogen bonds. Some recent work ignoring such constraints, even in 3–6 mol⋅dm⁻³ solutions, is criticized on this account.     

Preparation and electrochemical performance of Li3V2(PO4)3/C cathode material by spray-drying and carbothermal method

Composite Li₃V₂(PO₄)₃/C cathode material can be synthesized by spray-drying and carbothermal method. The monoclinic-phase Li₃V₂(PO₄)₃/C was prepared with the process of double spray drying at 260 °C and subsequent heat treatment at 750 °C for 12 h. The results indicate that the Li₃V₂(PO₄)₃/C presents large reversible discharge capacity of 121.9 mA h g⁻¹ and charge capacity of 131.8 mA h g⁻¹ at the current density of C/5, good rate capability with 61.1 mA h g⁻¹ at 20C, and excellent capacity retention rate close to 100% at various current densities in the region of 3.0–4.3 V.     

Isolation,Purification and Characterisation of Low Molecular Weight Xylanase from <Emphasis Type="Italic">Bacillus pumilus</Emphasis> SSP-34

Low molecular weight endo-xylanase from Bacillus pumilus SSP-34 was purified to homogeneity using ion exchange and size exclusion chromatographies. Xylanases were isolated by novel purification protocol which includes the use of anion exchange matrix such as DEAE Sepharose CL 6B with less affinity towards enzyme protein. The purified B. pumilus SSP-34 have a molecular weight of 20 kDa, with optimum pH and temperature at 6.0 and 50 °C, respectively. The enzyme was stable at 50 °C for 30 min. It showed remarkable stability at pH values ranging from 4.5 to 9 when the reaction was carried out at 50 °C. K _m and V _max values, determined with oats spelts xylan were 6.5 mg ml⁻¹ and 1,233 μmol min⁻¹ mg⁻¹ protein, respectively, and the specific activity was 1,723 U mg⁻¹