Diethyl Sulfate Research Articles

Facile N9-Alkylation of Xanthine Derivatives and Their Use as Precursors for N-Heterocyclic Carbene Complexes.

The xanthine-derivatives 1,3,7-trimethylxanthine, 1,3-dimethyl-7-benzylxanthine and 1,3-dimethyl-7-(4-chlorobenzyl)xanthine are readily ethylated at N9 using the cheap alkylating agents ethyl tosylate or diethyl sulfate. The resulting xanthinium tosylate or ethyl sulfate salts can be converted into the corresponding PF6− and chloride salts. The reaction of these xanthinium salts with silver(I) oxide results in the formation of different silver(I) carbene-complexes. In the presence of ammonia, ammine complexes [Ag(NHC)(NH3)]PF6 are formed, whilst with Et2NH, the bis(carbene) salts [Ag(NHC)2]PF6 were isolated. Using the xanthinium chloride salts neutral silver(I) carbenes [Ag(NHC)Cl] were prepared. These silver complexes were used in a variety of transmetallation reactions to give the corresponding gold(I), ruthenium(II) as well as rhodium(I) and rhodium(III) complexes. The compounds were characterized by various spectroscopic methods as well as X-ray diffraction.

Development and Validation of a Sensitive GC-MS/MS Method for the Determination of Five Potential Genotoxic Impurities in Abiraterone Acetate.

A sensitive and selective gas chromatography-tandem mass spectrometry method was developed for the identification and quantification of five potential genotoxic impurities (PGIs), i.e., chloromethane, 2-chloropropane, 2-bromopropane, 4-chloro-1-butanol and diethyl sulfate, in abiraterone acetate. The method was validated according to the International Council for Harmonisation (ICH) guidelines. The linearity was established for the concentration range of 30-480ng/mL (2-chloropropane, 2-bromopropane, 4-chloro-1-butanol and diethyl sulfate) and 90-1440ng/mL (chloromethane). The correlation coefficient of each PGIs was >0.995. The extraction recoveries ranged from 90.49 to 106.79% for the five PGIs. The quantitation limit, detection limit, accuracy, precision, repeatability and stability of the method demonstrated that the method was an adequate quality control tool for quantitation and identification of chloromethane, 2-chloropropane, 2-bromopropane, 4-chloro-1-butanol and diethyl sulfate at trace levels in drug substances and drug products.

Synthesis of 1-(4-Ethoxy-3-methoxybenzyl)-1,10-phenanthrolin-1-ium Bromide and Its Evaluation as Antiplasmodium through Heme Polymerization Inhibitory Activity (HPIA) Assay

This study describes the development of N-benzyl-1,10-phenantrolinium salt as an antiplasmodium agent. The salt, that is, 1-(4-ethoxy-3-methoxybenzyl)-1,10-phenanthrolin-1-ium bromide, was prepared using vanillin as the starting material in four simple synthetic steps. First, the alkylation of vanillin using diethyl sulfate produced 4-ethoxy-3-methoxybenzaldehyde in 79% yield. Second, the reduction of the protected vanillin by NaBH4 through the grinding method allowed us to obtain 4-ethoxy-3-methoxybenzyl alcohol in 96% yield. Next, the bromination of the benzyl alcohol under solvent-free condition led to the formation of the corresponding benzyl bromide, which in turn underwent bimolecular nucleophilic substitution with 1,10-phenanthroline to produce the desired product, that is, 1-(4-ethoxy-3-methoxybenzyl)-1,10-phenanthrolin-1-ium bromide, in 58% yield. The evaluation of N-benzyl-1,10-phenantrolinium salt as an antiplasmodium agent was conducted through heme polymerization inhibitory activity (HPIA) assay. The results showed that the phenantroline salt and chloroquine displayed the HPIA half maximal inhibitory concentrations of 3.63 and 4.37 mM, respectively. Therefore, 1-(4-ethoxy-3-methoxybenzyl)-1,10-phenanthrolin-1-ium bromide displays desirable HPIA and has a great potential to be further developed as an antiplasmodium.

Stability in solution and chemoprotection by octadecavanadates(IV/V) in E. coli cultures

Two mixed-valence octadecavanadates, (NH4)2(Me4N)5[VIV12VV6O42I]·Me4NI·5H2O (V18I) and [{K6(OH2)12VIV11VV7O41(PO4)·4H2O}n] (V18P), were synthesized and characterized by single-crystal X-ray diffraction analysis and FTIR, Raman, 51V NMR, EPR and UV/Vis/NIR spectroscopies. The chemoprotective activity of V18I and V18P towards the alkylating agent diethyl sulfate was assessed in E. coli cultures. The complex V18I was nontoxic in concentrations up to 5.0 mmol L−1, while V18P presented moderate toxicity in the concentration range 0.10 - 10 mmol L−1. Conversely, a ca. 35% enhancement in culture growth as compared to cells treated only with diethyl sulfate was observed upon addition of V18I (0.10 to 2.5 mmol L−1), while the combination of diethyl sulfate with V18P increased the cytotoxicity presented by diethyl sulfate alone. 51V NMR and EPR speciation studies showed that V18I is stable in solution, while V18P suffers partial breakage to give low nuclearity oxidometalates of vanadium(V) and (IV). According to the results, the chemoprotective effect depends strongly on the direct reactivity of the polyoxidovanadates (POV) towards the alkylating agent. The reaction of diethyl sulfate with V18I apparently produces a new, rearranged POV instead of poorly-reactive breakage products, while V18P shows the formation and subsequent consumption of low-nuclearity species. The correlation of this chemistry with that of other mixed-valence polyoxidovanadates, [H6VIV2VV12O38PO4]5- (V14) and [VIV8VV7O36Cl]6- (V15), suggests a relationship between stability in solution and chemoprotective performance.

Quantitative evaluation of DNA damage caused by atmospheric and room-temperature plasma (ARTP) and other mutagenesis methods using a rapid umu-microplate test protocol for microbial mutation breeding

• Mutation breeding is an important technique used in the improvement of strain traits in fermentation industry. • However, the evaluation of the effects after different mutagenic treatment lacks quantification strategies. • Here we developed the umu -microplate test providing a feasible method for a quick quantitative evaluation of DNA damage caused by different mutagenic treatments in the study of the microbial mutation breeding. Mutagenesis is an important technique for microbial mutation breeding. As the source of mutations, DNA damage extent is a key indicator for the effectiveness of mutagenesis. Therefore, a rapid and easy DNA damage quantification method is required for the comparison of mutagenesis effects and development of mutagenesis tools. Here, we used the umu -microplate test system to quantitatively compare the DNA damage strength caused by atmospheric and room-temperature plasma (ARTP) and other traditional mutagenesis methods including: ultraviolet radiation (UV), diethyl sulfate (DES) and 4-nitroquinoline-1-oxide (4-NQO). The test strain of Salmonella typhimurium TA1535/pSK1002 was used to monitor the time-course profile of β-galactosidase activity induced by DNA damage caused by different mutagenesis methods using a microplate reader. The umu -microplate test results showed that ARTP caused higher extent of DNA damage than UV and chemical mutagens, which agrees well with the result obtained by SOS-FACS-based quantification method as reported previously. This umu -microplate test is accessible for broad researchers who are lack of the expensive FACS instruments and allows the quick quantitative evaluation of DNA damage among living cells for different mutagenesis methods in the study of the microbial mutation breeding.

A Novel Synthesis Technology of Ethyl-β-Cyclodextrin

A novel green synthesis process about ethyl-β-Cyclodextrin have been investigated through the reaction between β-cyclodextrin and diethyl carbonate by used anhydrous potassium carbonate as catalyst in DMF. The influence of experimental factors including reaction time, the molar ratio of β-cyclodextrin to diethyl carbonate, reaction temperature and contents of anhydrous potassium carbonate on the average degree of substitution of ethyl-β-cyclodextrin were carried out by a design method of orthogonal experiments. The results shown that the average degree of substitution of ethyl-β-cyclodextrin can be depend on the reaction temperature and the molar ratio of raw material primarily. The optimal maximum average degree of substitution of ethyl-β-cyclodextrin is 6.0 when the molar ratio of β-cyclodextrin to diethyl carbonate, reaction temperature, reaction time and dosage of catalyst are 1:28, 120°C, 24h and 2.0 g, respectively. The structures of ethyl-β-cyclodextrin were characterized by TLC, IR, MS, ¹H-NMR and ¹³C-NMR, and these results are concordant with former ones which synthesized ethyl-β-cyclodextrin by diethyl sulfate or ethyl iodide.

Prediction of viscosity and surface tension properties of aqueous solution of ionic liquids EMIES

1-Ethyl-3-methylimidazolium ethyl sulfate (EMIES) was synthesized by using N-methylimidazole and diethyl sulfate as raw materials, the density, viscosity and surface tension of aqueous EMIES with various concentrations were measured in the temperature range from 288.15 to 328.15 K. According to the Jones-Dole empirical equation, the viscosity B-coefficients were calculated. Using the empirical equation of the relationship between Δμ2≠0 and T, the activation entropy (ΔS2≠0) and activation enthalpy (ΔH2≠0) were further obtained, the viscosity values of aqueous EMIES were predicted by using the Eyring semiempirical equation, which were highly consistent with experimental values of the viscosity. Furthermore, on the basis of the traditional Eӧtvӧs empirical equation, the new Eӧtvӧs equation was further derived by using the linear relationship between the molar surface Gibbs energy (gs) and temperature, each parameter has the definite physical meaning and a new unit of energy in new Eӧtvӧs equation. The predicted values of the surface tension of aqueous EMIES were obtained by the definition of the molar surface Gibbs energy, which were highly consistent with the experimental values of the surface tension.

Chrysomutanin and related meroterpenoids from a DES mutant of the marine-derived fungus Penicillium chrysogenum S-3-25

A new meroterpene, chrysomutanin (1), two new meroterpenoids (4 and 5) together with nine known ones were isolated from the diethyl sulphate (DES) mutant 3d10-01 of the marine-derived fungus Penicillium chrysogenum S-3-25. The structures of the isolated compounds were determined by their spectroscopic data, and the absolute configuration of 1 was determined by Rh2-induced electrical circular dichroism (ECD) analysis or by comparison of the measured ECD with that of the known compounds. The cytotoxic activity was preliminarily evaluated against five human cancer cell lines. HPLC-UV analysis showed that compounds 1–12 were all newly produced by the mutant, and were not detected from the initial strain S-3-25. Chrysomutanin (1) is a new member with a chain sesquiterpene unit to the family of meroterpenes. Present results confirm that DES mutagenesis strategy is an effective method to exploit the dormant metabolites of fungi.

AFLP-Based Analysis of Variation and Population Structure in Mutagenesis Induced Faba Bean

Genetic diversity enrichment is urgently necessary to develop climate-resilient faba bean cultivars. The present study aimed to measure the enrichment of genetic diversity and changes in the population structure of faba bean, following induced mutagenesis. 120 samples, including 116 M2 mutant plants, generated by exposing the ILB4347 accession to four mutagen treatments (25 and 50 Gray gamma radiation and 0.01%, and 0.05% diethyl sulfate) and four reference genotypes were characterized using 11 amplified fragment length polymorphism (AFLP) primer combinations. The AFLP markers generated 1687 polymorphic alleles, including 756 alleles (45%) that were detected infrequently (f ≤ 0.1). The total allele count of the mutant plants ranged from 117 to 545. We observed a wide range of banding patterns and counts among the mutant plants, showing the high genetic diversity induced by mutation. Mutations also changed the population structure, by altering 31.78% of the total membership coefficient (Q). Although mutations changed the population structure, Nei’s genetic distance showed that the mutant population remained closely related to its control parent. This is the first report examining genetic diversity and population changes in faba bean mutant populations and, thus, could facilitate the application of induced mutagenesis during faba bean breeding.

Gene expression response of mutagenic breeding of Acidithiobacillus sp. FJ2 to different concentrations of uranium low grade ore

Acidithiobacillus ferrooxidans ( At. f ) is a bacterium involved in the bioleaching process . In the present study, we investigated the effects of Acidithiobacillus sp. FJ2 induced with diethyl sulfate (DES) as mutagen on the bioleaching of low grade uranium ore. The comparison was carried out within 5, 10, 15, 25 and 50% uranium ore pulp densities in the bioleaching system. The impact of the induction of Acidithiobacillus sp. FJ2 was determined by Eh and pH values, concentrations of Fe 2+ and uranium extraction rates at 24 h intervals. The results showed that DES with 0.8% may lead to an obvious change on bacteria leading to improvement of bioleaching capability in 5, 10, 15% pulp densities. However, the bioleaching activity of the original bacteria was more efficient than DES-induced bacteria at 25 and 50% pulp densities. The gene expression results in 0.8% DES treated bacteria indicated that the bacteria attempt to adjust in the bioleaching systems (with different pulp densities) through decreased cyc2 and increased rus , cyc1 and coxB levels. These results suggest that uranium may induce oxidative stress in the wild and treated strains in the high pulp density, while the bacteria tried to survive and gain more energy from the iron oxidation. However, when the amount of uranium increased, the mutants couldn’t cope up woth the enhanced stress in 25 and 50% pulp densities. It may be due to inhibitory effect of uranium toxicity on adaptive processes which may change the trends.

Morphological diversity of faba bean (Vicia faba L.) M2 mutant populations induced by gamma radiation and diethyl sulfate

Induced mutagenesis is one of the most widely used breeding methods for rapidly increasing genetic diversity in many crops. This study used gamma radiation and diethyl sulfate (DES) to increase the genetic diversity in two faba bean genotypes. The genetic diversity assessment was based on 12 qualitative traits. The purpose of this study was to assess the genetic diversity in the M2 faba bean mutant populations and to select promising mutant. A total of 5271 M2 seeds generated from the M1 generation of two genotypes—Hassawi 2 and ILB4347—were used in this study. The seeds of M1 were obtained by gamma-ray exposure (25 and 50 Gy) and treatment of 0.01, 0.02, and 0.05% DES. A total of 4333 (82.2%) seeds germinated, of which 4102 plants survived and were screened for morphological diversity. A total of 36 mutant types are presented in this study. Determinate plant type, white flower color, white hilum, and black seed coat color were some of desirable traits that were considered for further generations. Multivariate-based principal component analyses showed that high genetic diversity was induced by gamma radiation and DES in faba bean mutant populations. This study revealed the potency of gamma radiation and DES on increasing genetic diversity and demonstrated the successful program of induced mutagenesis in the faba bean.

Improvement of vincamine production of endophytic fungus through inactivated protoplast fusion.

Improvement of the production of vincamine in endophytic fungus VINI-7 was performed by using the inactivated protoplast fusion method. The preparation conditions of protoplasts were optimized by systematic trials with various parameters, and inactivated protoplast fusion was subsequently performed. The mycelium in logarithmic growth phase was treated with 1500U/mL lywallzyme, 1500U/mL lysozyme, 2000U/mL cellulase, and 1000U/mL snailase solution for 3h at 30°C and had the best conditions, in which the concentration of the protoplast was 3.17 × 107 cells/mL. Protoplasts were inactivated by heat, ultraviolet, microwave, sodium nitrite, and diethyl sulfate, respectively. Subsequently, protoplasts inactivated by different methods were subjected to respective protoplast fusion. The results showed that the yield of vincamine in fusants inactivated by mutagens was generally higher than that of fusants inactivated by heat. The highest yield of vincamine in two fusants (U-U1 and N-N1) was 31.6 and 38.7mg, which increased to 162.24 and 221.16%, respectively, as compared to the parent strain (12.05mg). LC-MS/MS analysis showed that U-U1 and N-N1 fusants could produce vincamine. Furthermore, the results of genetic stability experiments indicated that U-U1 and N-N1 were genetically stable.

Phenotypic and molecular cytogenetic variability in calendula (Calendula officinalis L.) cultivars and mutant lines obtained via chemical mutagenesis

The morphological, meiotic and chromosomal variability were studied in two cultivars of Calendula officinalis L. and their mutant lines obtained though chemical mutagenesis using diethyl sulphate (DES) (0.04%, 0.08%) and dimethyl sulphate (DMS) (0.025%, 0.05%). The studied cultivars displayed different sensitivity to DMS and DES mutagens. More M1 plants with morphological changes were observed in C. officinalis cv. ‘Zolotoe more’ than in cv. ‘Rajskij sad’. DMS and DES at low concentrations had positive effects on main agro-metrical traits in both cultivars including plant height, inflorescence diameter and number of inflorescences per plant. Dose-dependent increase in number of various meiotic abnormalities was revealed in both mutant lines. Comparative karyotype analysis and FISH-based visualization of 45S and 5S rDNA indicated a high level of karyotype stability in M1 and M2 plants. Seed treatments with DMS and DES at certain concentrations resulted in higher yields of inflorescences in M1 plants compared to the control. In M2 generation, dose-dependent reduction in the yields of inflorescences was observed. Our findings demonstrate that DMS and DES at low concentrations have great potential in calendula mutation breeding.

Engineering Corynebacterium glutamicum Mutants for 3-Methyl-1-butanol Production.

3-Methyl-1-butanol (3MB) is a promising biofuel that can be produced from 2-ketoisocaproate via the common L-leucine biosynthesis pathway. Corynebacterium glutamicum was chosen as a host bacterium because of its strong resistance to isobutanol. In the current study, several strategies were designed to overproduce 3MB in C. glutamicum through a non-fermentation pathway. The engineered C. glutamicum mutant was obtained by silencing the pyruvate dehydrogenase gene complex (aceE) and deleting the lactic dehydrogenase gene (ldh), followed by mutagenesis with diethyl sulfate (DES) and selection with Fmoc-3-4-thiazolyl-L-alanine (FTA). The mutant could produce 659mg/L of 3MB after 12h of incubation. To facilitate carbon flux to 3MB biosynthesis, the engineered recombinant was also constructed without branched-chain acid aminotransferase (ilvE) activity by deleting the ilvE gene. This recombinant could produce 697mg/L of 3MB after 12h of incubation.

Continuous synthesis of tetraalkylammonium‐Based ethyl sulphate ionic liquid and its kinetic study in microreactors

A tetraalkylammonium‐based ionic liquid (IL) was synthesized with diethyl sulphate and triethylamine in a capillary microreactor. Solvent‐free conditions coupled with microreactor technology greatly simplified the IL construction process. Optimization of process parameters (e.g., temperature, residence time, and inner diameter of capillary) was performed, and the maximum yield was 94% with the residence time of 300 s at 90°C. Interestingly, the obtained product could exist as a molten salt with low viscosity at a low melting point of 24.1°C. Key dimensionless groups such as Pe, Bo, and DaII numbers were calculated to account for the mass transport phenomena and reaction performance in this IL synthesis or to validate the reliability of the corresponding reaction kinetics study. Based on fast heat and mass transfer rates provided by the microreactor, intrinsic reaction kinetics could be obtained. Moreover, gold (III) extraction was remarkably improved by the application of the synthesized IL. © 2018 American Institute of Chemical Engineers AIChE J, 65: 1245–1255, 2019

Nitrous Acid and Diethyl Sulfate, as Chemical Mutagenic Agents, to Improve the Biomass, Metabolites, Enzyme Activity, and Antioxidant Activity of Wild <i>Phellinus igniarius</i>

Nitrous Acid and Diethyl Sulfate, as Chemical Mutagenic Agents, to Improve the Biomass, Metabolites, Enzyme Activity, and Antioxidant Activity of Wild <i>Phellinus igniarius</i>

Two new polyketides isolated from a diethyl sulphate mutant of marine-derived Penicillium purpurogenum G59

Two new polyketides, purpurofuranone (1) and purpuropyranone (2), were isolated along with the known polyketides, cillifuranone (3) and taiwapyrone (4), from a mutant BD-3n-1 derived from the diethyl sulfate (DES) mutagenesis of a marine-derived Penicillium purpurogenum G59. The structures of 1 and 2 were elucidated by spectroscopic methods especially on the basis of X-ray diffraction and calculated optical rotations data. The plausible biosynthesis of 1 − 4 was also proposed and discussed. In preliminary MTT assay, 1 − 4 showed no notable inhibitory effects on the tested four human cancer cell lines.

Screening and Mutation of Saccharomyces cerevisiae UV-20 with a High Yield of Second Generation Bioethanol and High Tolerance of Temperature, Glucose and Ethanol.

A wild-type strain was isolated from slightly rotted pears after three rounds of enrichment culture, identified as Saccharomyces cerevisiae 3308, and evaluated for its fermentation capability of second generation bioethanol and tolerance of temperature, glucose and ethanol. S. cerevisiae 3308 was mutated by using the physical and chemical mutagenesis methods, ultraviolet (UV) and diethyl sulfate (DES), respectively. Positive mutated strains were mainly generated by the treatment of UV, but numerous negative mutations emerged under the treatment of DES. A positive mutated strain, UV-20, produced ethanol from 62.33 ± 1.34 to 122.22 ± 2.80g/L at 30-45°C, and had a maximum yield of ethanol at 37°C. Furthermore, UV-20 produced 121.18 ± 2.51g/L of second generation bioethanol at 37°C. Simultaneously, UV-20 exhibited superior tolerance to 50% of glucose and 21% of ethanol. In a conclusion, all of these results indicated that UV-20 has a potential industrial application value.

Optimization of the method for the determination of diethyl sulfate at workplaces

Diethyl sulfate (DES) is a substance classified to the group of carcinogens. The value of maximum admissible concentration for this substance in workplace air is not specified in Poland. Due to the use of DES in domestic companies there is a need to develop a sensitive method for the determination of diethyl sulfate in the work environment. Studies were performed using gas chromatography (GC) technique. An Agilent Technologies chromatograph, series 7890A, with a mass selective detector (5975C, Agilent Technologies, USA) was used in the experiment. Separation was performed on a capillary column with Rtx-5MS (30 m × 0.25 mm × 0.25 μm) (Restek, USA). The possibility of using sorbent tubes filled with activated carbon (100 mg/50 mg), silica gel (100 mg/50 mg) and Porapak Q (150 mg/75 mg) for absorption of diethyl sulphate was investigated. The method of sampling air containing diethyl sulfate was developed. Among the sorbents to absorb DES Porapak Q was chosen. Determination of the adsorbed vapor includes desorption of DES, using dichloromethane/methanol mixture (95:5, v/v) and chromatographic analysis of so obtained solution. Method is linear (r = 0.999) within the investigated working range of 0.27- -5.42 μg/ml, which is an equivalent to air concentrations 0.0075-0.15 mg/m3 for a 36 l air sample. The analytical method described in this paper allows for selective determination of diethyl sulfate in the workplace air in the presence of dimethyl sulfate, ethanol, dichloromethane, triethylamine, 2-(diethylamino)ethanol, and triethylenetetramine. The method meets the criteria for performing procedures aimed at measuring chemical agents, listed in EN 482. Med Pr 2018;69(3):291-300.

A new polyketide purpurogenic acid: the activated production of polyketides by the diethyl sulphate mutagenesis of marine-derived Penicillium purpurogenum G59

A new polyketide, purpurogenic acid (1), and two known polyketides, (-)-mitorubrin (2) and (-)-mitorubrinol (3), were isolated from a fungal mutant derived from the diethyl sulphate (DES) mutagenesis of marine-derived Penicillium purpurogenum G59. The planar structure of new 1 was elucidated by spectroscopic methods and the absolute configuration was assigned on the basis of [α]D and CD data. In our preliminary MTT assay, 1 inhibited human cancer K562, HL-60, HeLa and BGC-823 cells with the inhibition rates of 52.7, 78.8, 38.4 and 35.3% at the 100 μg/mL, respectively.