The modification of an MB-2 bipolar ion exchange membrane with chromium (III) hydroxide was carried out by a chemical method, namely, by the sequential treatment of the membrane with a solution of chromium (III) salt and alkali. Data on the morphology, phase, and chemical composition of the modified membrane were obtained using scanning electron microscopy and energy-dispersive analysis. In particular, it was shown that the modifier was distributed in a layer 30–50 microns thick at the boundary of the cation- and anion-exchange layers of the bipolar membrane. The electrochemical behavior of the modified membrane in the process of sodium sulfate conversion was studied by measurements of the following characteristics: the current efficiency of the acid and base, the energy consumption of the process, and the degree of contamination of the target products with salt ions. It was shown that the resulting membrane has an alkali and acid yield of 61% and 57%, respectively. This is higher than the same yields for the industrial unmodified MB-2 membrane (38% and 30%). The results of this study demonstrated that the modified samples allowed obtaining a higher yield of acid and base, reducing the content of salt ions in the target products and also reducing the electricity consumption for obtaining a unit of the target product. The concentration dependences of the electrical conductivity of the MK-40 heterogeneous ion-exchange membrane, which is a cation-exchange layer of MB-2, in sodium sulfate solutions before and after its modification with chromium (III) oxide were obtained. A decrease in the specific electrical conductivity of the membrane with the introduction of a modifier was established. A quantitative assessment of the influence of the modifier on the current flow, volume fraction, and spatial orientation of the conductive phases of MK-40 was carried out using an extended three-wire model for the description of the model parameters of ion-exchange materials. When a modifying additive was introduced into MK-40, the fraction of the current passing through the inner solution and the intergel phase decreased. This was due to the substitution of part of the free solution in the pore volume by the modifier. A variant of the practical application of electrodialysis with the chromium-modified bipolar ion-exchange membranes is recommended. Over the past 10 years, research interest on the issue of chromium-containing wastewater treatment increased, which was due to the active use of chromium compounds in industrial processes and the improvement of existing methods as well as the appearance of new ones for the removal of the toxic metal . Industrial wastewater contains both trivalent Cr(III) ions and hexavalent Cr(VI) ions. Cr(VI) is considered more toxic than Cr(III) . The max2O72− to Cr3+:The most common treatment of chromium-containing wastewater in the electroplating industry is a method based on the reduction of chromium (VI) followed by the precipitation of trivalent chromium ,14. ThisThe reduction is carried out in an acidic medium at pH = 2–2.5. This requires the additional acidification of wastewater with a 10–15% sulfuric acid solution. For acidic solutions, instead of iron (II) salts, sodium sulfite or bisulfite is used:After the reduction of chromium, wastewater is neutralized with limewater to a pH of 8.5–9. Hydroxides are released as the result of neutralization:4 crystals in the pulp contributes to the improvement of the sedimentation and filtration processes. The use of alkali solutions for the precipitation of trivalent chromium in the form of hydroxide is also possible:During the precipitation of iron (III) hydroxide, the most complete transition of water impurities into the solid phase occurs due to the co-precipitation and adsorption on the surface of freshly precipitated iron hydroxide. The presence of CaSOSuch methods of processing of waste chromium-containing solutions result in sulfates remaining in the system after the separation of the sediments. Sulfates are secondary waste products which require disposal.2O72− 3).We propose to use electrodialysis with bipolar membranes for the conversion of these solutions. The acid and alkali obtained during the conversion can be used during earlier stages of the chemical treatment of wastewater containing CrBipolar ion-exchange membranes consist of a cation-exchange layer (CEL) tightly coupled with an anion-exchange layer (AEL). At the boundary of these layers, when an electric potential gradient is applied to the system, the irreversible dissociation of water molecules occurs, and the dissociation products leave the reaction zone. Some studies ,16,17,18Both inorganic substances, including metals introduced into the ion exchanger in various forms  ,24,25,263 has the highest catalytic activity in the dissociation of water in the bipolar region [The MB-2 bipolar membrane is characterized by a very high operating voltage of water dissociation and a low current efficiency of acids and bases . It is kr region ,21. The The goals and objectives of this study were the modification of a commercially available bipolar membrane with chromium (III) hydroxide; an investigation of the features of the distribution of the modifier over the volume of the sample; an evaluation of the influence of the modifier on the characteristics of the salt (sodium sulfate) conversion process for the possible suggestion of a variant of the practical application of electrodialysis based on a chromium-modified bipolar membrane.3); washing the membrane with water; treatment of the membrane with a sodium hydroxide solution; washing the membrane with water again.The MB-2 membrane was modified chemically . This prAn analysis of the surface morphology and volume of the modified membrane was performed by means of electron microscopy using a JSM-6380LV  scanning electron microscope and a scanning electron microscope with a Quanta Nova field-emission cathode  in the mode of secondary electrons. The chemical composition of the membrane material was assessed by an energy-dispersive X-ray analyzer .m, was calculated according to the formula:2.The effect of the modifier, chromium (III) oxide, on the mechanism of current flow through the MB-2 cation-exchange layer was evaluated using an extended three-wire model for the description of the parameters of ion-exchange materials . For thi1, f2, α) and geometric  sample parameters based on the concentration dependence of the electrical conductivity of an ion-exchange material [1 and f2 are the volume fractions of the gel and intergel phases; α is a parameter that reflects the mutual arrangement of phases in the material with respect to the direction of the current and takes values from +1  to −1 (series). The value α should be taken into account for the description of the diffusion permeability of ion-exchange membranes. This method of parametrization of ion-exchange membranes is based on an approach that was developed for ion-exchange columns and resins. The current flows through the ion-exchange material through three parallel channels  hydroxide (designated as MB-2-Cr) and the commercially available MB-3 and MB-2 bipolar membranes were used . MK-40 a2, the intermembrane distance in the channels was 1 mm. The experiment was carried out in three variants. The working area of one membrane was 14 cm3, sulfuric acid (0.05 mol/dm3), and a sodium hydroxide solution (0.1 mol/dm3) were delivered to chambers 3 and 4. The rate of the saline flow was 20 cm3/min, and that of the acid and base solutions was –1 cm3/min. The number of elementary cells was n = 1. Such a saline flow rate in the desalination chambers allowed the process to be carried out in under-limit current modes. The formation of hydrogen and hydroxide ions occurred under the influence of the direct current in the bipolar membrane, and these ions migrated into the acidic (3) and basic (4) chambers. From the salt desalination sections (2 and 5), sulfate ions and sodium ions migrated through the anion- and cation-exchange membranes into the acidic and alkaline chambers, respectively. The process was carried out in galvanostatic mode: the current density was set (14.3–64.3 mA/cm2), and after a steady state was established in the system, the samples were analyzed. First variant. In the electrodialyzer : certain fixed volumes of salt, acid, and base solutions repeatedly passed through the electrodialyzer and returned to the storage tanks. The flows supplied to the chambers are shown in , a. The 42− ion flows through the bipolar membrane from chamber 3 to the main chamber 4 and Na+ ion flows from chamber 4 to chamber 3 , flux density of hydrogen and hydroxyl ions, formed inside the bipolar membrane ), specific energy consumption  (experiment according to variants 1 and 2) were calculated based on the results of experiments:The concentration of hydrogen and hydroxide ions in the acidic and basic chambers was determined using acid–base titration ; the conc is the change in the concentration of hydrogen H+ and hydroxyl OH− ions at the inlet and outlet of the concentration chamber, mol/m3; i is current density, A/m2; I is the operating value of the current strength, A; U is the operating voltage, V; τ is the time of the experiment, h; m is the mass of the target product  obtained during the experiment, kg; ω is the volumetric flow rate of the solution, m3/s; n is the number of elementary cells  in the electrodialyzer; A is the active membrane area, m2; F is the Faraday constant, 96485 C/mol. The degree of acid and base contamination with salt ions that crossed the membrane  was determined using the formula (12):Na+) is the amount of Na+ cations (in moles) that crossed into the acidic chamber for a certain time, ν(H+) is the amount of H+ ions (in moles) generated during the same time, ν(SO42−) is the amount of SO42− anions (in moles) that crossed into the alkali chamber for a certain time, and ν(OH−) is the amount of OH− ions (in moles) generated during the same time [Here, Δame time .When the MB-2 and MK-40 membranes were treated with a chromium salt solution, metal ions entered the cation exchanger and were localized near ionogenic centers as counterions . An ion-3 was formed near the ionogenic groups in the polymer:After treatment of the membrane with a sodium hydroxide solution, a sparingly soluble Cr(OH)In this case, the cation exchanger (the cation-exchange layer of a bipolar membrane or monopolar cation-exchange membrane) acquired the gray-green color characteristic of chromium (III) hydroxide. Such treatment resulted in the following reactions in the MB-2 anion-exchange layer:3 is present in the membrane in a nanoscale form, with a gradient distribution over the ionite particles [3 decreased exponentially from the surface deep into the ionite granules.It was shown in  that, unarticles : the conMicrographs of the cross section of MB-2-Cr are shown in The distribution of elements in the considered MB-2-Cr section is shown in 1, f2, and α determined from the concentration dependence of the specific electrical conductivity of the membrane are shown in For the assessment of the effect of the modifier on the cation-exchange layer of the bipolar membrane, the electrical conductivity of only the MB-2 cation-exchange layer of the MK-40 membrane was studied before and after its modification with chromium (III) hydroxide. The obtained concentration dependences of the electrical conductivity of the MK-40 and MK-40-Cr membranes are shown in 2) decreased. The decrease in c and f2 was due to the substitution of part of the free solution in the pore volume by the modifier. The decrease in the α parameter indicated the disordering effect of the modifier on the relative position of the conductive phases of the membrane. The symbatic change of c and f2 was noted in [The analysis of the model parameters showed that when a modifying additive was introduced into MK-40, the fraction of the current passing through the channel c and the intergel phase  hydroxide allowed reducing the contamination of the target products with salt ions and obtaining higher concentrations of acid and base with lower power consumption.Based on the data obtained, we can recommend the use of chromium-modified bipolar membranes for the electrodialysis treatment of secondary waste solutions formed during the processing of chromium-containing wastewater. In this case, the possible degradation of the modifier is not critical. In addition, the modification of industrial bipolar membranes with chromium can be carried out directly in industries where there is chromium-containing wastewater; obviously, this approach is more cost-effective.The scheme of the improved process for the treatment of chromium-containing wastewater is shown in 2O72- enters the chromium recovery stage (block 1). Next, is the neutralization stage (block 2), then the precipitation of sparingly soluble hydroxides (block 3) and the processing of the salt solution in an electrodialysis synthesizer with bipolar membranes (block 4). The resulting diluate and acid and alkali solutions can be used during the earlier stages of the chromium wastewater treatment process. Wastewater containing CrThe treatment of sodium sulfate by electrodialysis with bipolar chromium-modified membranes proposed in this study can also be useful to improve the technology considered in , where t2CrO4 after the oxidation of Cr(III) to Cr(VI) with H2O2 in an alkaline medium in an electrodialyzer with bipolar membranes was proposed, or in relation to another study [The use of a chromium-modified bipolar membrane can be recommended in a number of applications, for example, as a continuation of a previous study , where ter study , where tA chromium-modified bipolar ion-exchange membrane was obtained by the sequential chemical treatment of an industrial membrane with a chromium (III) salt and alkali. The structure of the membrane and its influence on the characteristics of the electrodialysis conversion of sodium sulfate were investigated.Chromium (III) hydroxide was found to be distributed in the layer of 30–50 μm, at the boundary of the cation and anion exchanger. When the modifier was introduced into the bipolar region, the acid and alkali performance during the conversion of sodium sulfate increased, and the specific energy consumption decreased almost twice as compared to the unmodified sample.It was shown that the chemical modification of the MB-2 bipolar membrane with chromium (III) oxide allowed not only an increase in the yield of acids and bases and a reduction in the energy consumption of the process, but also a reduction in the degree of contamination of the target products with salt ions by three to four times. The latter was associated with the limitation of the diffusion transfer of the salt, limited by the transfer of coions, and was a consequence of a change in the current transfer paths in the modified sample.3, solutions of sulfuric acid and sodium hydroxide with concentrations of 0.47 and 1.5 mol/dm3, respectively, were obtained. This was achieved with a volume ratio of salt to acid  solutions equal to 10:1.When implementing the process in the recycle mode, with the conversion of sodium sulfate at an initial concentration of 0.5 mol/dmThe use of a chromium-modified bipolar membrane in the electrodialysis treatment of both chromium-containing wastewater and secondary stock solutions obtained during the processing of chromium-containing wastewater is recommended.


In this study, a modified version of electrospun nylon 66 nanofibers by silica particles were blended into ordinary Portland cement to investigate the microstructure and some mechanical properties of cementitious material. The addition of silica into the nanofibers improved the tensile and compressive properties of the hardened cement pastes. The observations from the mechanical strength tests showed an increase of 41%, 33% and 65% in tensile strength, compressive strength, and toughness, respectively, when modifying the cement pastes with the proposed nanofibers. The observations from scanning electron microscopy and transmission electron microscopy showed the morphology and microstructure of the fibers as well as their behaviors inside the cement matrix. Additionally, X-ray diffraction and thermal gravimetric analysis clarified the occurrence of the extra pozzolanic reaction, as well as the calcium hydroxide consumption by the attached silica inside the cement matrix. Finally, the observations from this study showed the successful fabrication of the modified nanofibers and the feasibility of improving the tensile and compressive behaviors of cement pastes using the proposed electrospun nanofibers. Today, cement is a basic material that plays an important role in the construction sector. However, there is an increased demand for higher cement strength, due to the feasibility of reducing structural size and cement consumption, which can lead to significant ecological improvement and financial efficiency. Therefore, it has posed a considerable challenge for researchers and manufacturers. Cement matrix strength can be affected by different factors, such as the component ratio, the cement hydration process, the change in the cement matrix microstructure, the curing condition, the addition of additives, and so on ,2. TradiFurthermore, recent findings showed significant performance of different nanosized fibers in improving the cement matrix. The findings by Liu et al.  showed tType I ordinary Portland cement was purchased from Ssangyong Co., Korea . The proThe polymer solutions were prepared in two steps: (1) The solvent was prepared by stirring formic acid and chloroform in a volume ratio of 4:1. (2) Precursors, silica particles, and nylon 66 were added to the prepared solvent to obtain the polymer solutions at a precursor-to-solvent ratio of 1:4 by weight of the polymer solution. This proportion was determined after conducting a trial-and-error process to determine the optimal and spinnable viscosity of the polymer solution using a size 20 Taylor cone. To be more specific, 13 wt% of silica was added to two bottles containing 80 wt% of solvent. Then, the solutions were ultrasonicated for 2 h under laboratory conditions to break the van der Waals forces between the nanomaterial molecules and obtain homogeneous dissolution in the final solutions ,22. FinaElectrospun nanofiber fabrication and the blending process to obtain the composite cementitious materials containing electrospun nanofibers have been reported in previous works ,16. AfteOwing to the changes in the microstructure of the matrix and the difficulty in controlling the spontaneous cracking that occurs during the curing process, it is difficult to conduct a study on the mechanical properties of hardened cement pastes. Therefore, the cement paste samples were designed with small dimensions according to the specifications in the ASTM C307  and ASTMIn this study, the tensile strength and compressive strength tests were evaluated using a 5 kN mortar tensile strength test device in compliance with ASTM C307 and a 1000 kN hydraulic universal testing machine in compliance with ASTM C109/109M, respectively.The FE–SEM analysis was conducted using a Zeiss Merlin Compact system. Input parameters, a working distance of 9 mm, and an acceleration voltage of 5 kV were used. In addition, to increase the observation quality, a 5 Å platinum cover was coated on the sample surfaces.The FE–TEM analysis was conducted using an FEI Tecnai F30 Twin system under an acceleration voltage of 300 kV.XRD analysis was conducted using the D8 Advance system with a scanning speed of 0.4 s/step, a step size of 0.02° (2θ)/step from 5° to 70°, and CuKα radiation .TGA was conducted using a TA instrument SDT-Q600 under a nitrogen atmosphere with a flow rate of 100 mL/min, a heating velocity of 10 °C/min, and a heating range from 25 °C to 1000 °C.The results of the tensile and compressive strength tests indicate that the electrospun nylon 66 nanofibers modified with silica particles are effective in improving some select mechanical properties of the hardened cement paste see . OverallIn the case of compressive strength, an increase of 33% was observed with the addition of NS-N66 NFs, as compared to that of the unmodified paste a. In the4.2H2O), hillebrandite (Ca2SiO3(OH)2) and some dehydrated clinker grains, belite (C2S), and alite (C3S). The hydration products of all the samples were consistent and suitable with the results obtained from Jiang et al. [The XRD patterns of the plain cement and the electrospun nanofiber-blended cement are shown in g et al.  (Figure g et al. ,32.The decrease–increase tendency of the CH and CSH contents of the three samples was consistent with the order of the observations from the mechanical strength tests. The XRD analysis showed that the CH phase in the matrices was consumed. The increase in the CSH content was observed thanks to the pozzolanic reaction between the attached NS and the generated CH phase from the hydration processes.2 from the ambient environment. As can be observed from the TGA results, the CSH content from the nanofiber-blended cement pastes was higher than that of the plain paste, whereas that of CH was lower. It is also worth noting that despite the decomposition range of the CH phase being similar to the decomposition range of the electrospun nanofibers from 400 °C to 500 °C, as presented above, the CH content of the hardened modified cement pastes still decreased compared to that of the hardened plain paste, as indicated by TGA analysis. Consistent with the XRD results, these observations showed a decreasing tendency of the CH and CSH contents in the presence of NS-N66 NFs in the cement. Therefore, the attached silica particles on the nylon 66 nanofibers influenced the final product of the cement paste. Above all, these observations confirmed the initial hypothesis regarding the role of the pozzolanic agent in the attached silica particles.This study determined the influence of electrospun nylon 66 nanofibers containing silica particles on the microstructure and selected mechanical properties of the hardened cement pastes. The following conclusions were drawn:An increase of 41%, 33%, and 65% in tensile strength, compressive strength, and toughness, respectively, was observed when modifying the cement pastes with the proposed nanofibers containing silica particles.The nanosilica particles attached along the axis of the nylon 66 nanofibers were effective in increasing the generated CSH content from the pozzolanic reaction. In addition, the bridging and filling effects of the nanofibers inside the matrix were observed, which increased the mechanical strength of the hardened cement pastes.The XRD and TGA results demonstrated the decrease–increase tendency of the CH and CSH contents when blending the cement with NS-N66 NFs. Therefore, the role of the attached silica particles is clarified.Finally, electrospun nylon 66 nanofibers containing silica particles is a promising candidate for strengthening cementitious materials. It can be seen that an improvement in the mechanical properties of cement can lead to extending the durability and service life of concrete. However, for the practical application of this product, further research needs to be conducted to improve the performance of the composite cement fabrication process, as well as investigate the effectiveness of these nanofibers on other properties of cement paste and concrete. Additionally, the analysis of the life cycle and the life cycle cost of the concrete structure utilizing the proposed cement is necessary, which can evaluate the competitiveness of the proposed cement solution and help authorities in the decision-making process . In addi


The experimental design was in randomized blocks with four repetitions and six treatments . The following analyses were performed: K and Cl content in the leaves and the soil, stocks of Cl in soil, yield, removal of K and Cl with the beans, cup quality of the beverage, polyphenol oxidase activity (PPO), electric conductivity (EC), potassium leaching (KL), the content of phenolic compounds, the content of total sugars (TS), and total titratable acidity (TTA). The stocks of Cl in the soil decreased as the proportion of KCl in the fertilizer was reduced. The fertilization with KCl reduces the cup quality and the activity of the polyphenol oxidase, probably due to the ion Cl. The increase in the application of Cl directly relates to the increase in potassium leaching, electric conductivity, and titratable acidity. Indirectly, these variables indicate damages to the cells by the use of Cl in the fertilizer. The activity of the polyphenol oxidase enzyme and the cup quality indicate that the ion Cl- reduces the quality of the coffee beverage. K content in the leaves was not influenced by the application of blends of K fertilizer while Cl content increased linearly with KCl applied. The application of KCl and K2SO4 blends influenced coffee yield and the optimum proportion was 25% of KCl and 75% of K2SO4. The highest score in the cup quality test was observed with 100% K2SO4.The present study had the objective to evaluate the effect of blends of KCl and K Coffea arabica accounting for 77% the total revenue (R$ 47.48 billion). The state of Minas Gerais is the largest producer, responding for R$ 33.28 billion or 54% of national revenues  × 100, where SI is the saline index, ECa is the electric conductivity of the sample, and ECb is the electric conductivity of the sodium nitrate solution. The SI found were 142, 130, 121, 112, and 104%, for T1, T2, T3, T4, and T5, respectively.Before the experiment, the saline index of each blend of K fertilizer was determined by comparing a 10 g L2O applied were 150, 200, and 300 kg ha−1 for the respective agricultural years of 2017/2018, 2018/2019, and 2019/2020. All K fertilizations were divided into three applications.The maintenance fertilization was done according to Guimarães et al.  using thK and Cl content in the leaves−1) was determined with a selective electrode  coupled to a Hanna® device, model HI2221. The determination curve was built using the concentrations of 2, 20, 200, and 1000 mg L−1 of Cl.The third and fourth pair of leaves on both sides of the plants were collected from the useful area 20 days after the second application of the cover fertilization. The leaves were washed in deionized water, dried at 65 °C, and grounded in a Willey mill. The plant material was digested in a solution of nitric-perchloric acid (4 parts of nitric acid to 1 part of perchloric acid), and K was determined with inductively coupled plasma (ICP). To determine Cl, 1 g of grounded material was added to 50 mL of ultrapure water under agitation for 15 min . After fYield®, model DPM-02) and submerged for 24 h to remove the mucilage. After removing the peels and the rotten beans, samples were air-dried until a 10.8% to 11.2% moisture level.The harvests were done when more than 70% of the fruits were mature. For the chemical analyses, 4 L of beans at the cherry stage were collected two days before each harvest. Fruits were peeled with an electric peeler .K and Cl content and removal in the beansK and Cl contents in the beans were determined at the cherry stage after air-drying  and grounding the beans in a Willey mill. K content was determined after nitric-perchloric digestion with measures done by ICP. Cl content followed the same procedures to quantify Cl in the leaves. The amounts of these elements removed from the soil were obtained by multiplying their content in the beans by the yield on each treatment.Stocks of Cl in the soilStocks of Cl in the 0–20 and 20–80 cm layers of soil were checked during the experiment. Six soil samples were taken from the soil underneath the projection of the tree canopies (three from each side of the parcel). Extraction and determination of Cl followed the same procedure described for leaf Cl content, but with the proportion of 10 g of soil to 50 mL of ultrapure water. The stocks were determined by multiplying the element concentration by the mass of soil in each layer.The analytical standard Tomato leaves (NIST 1573A), with 0.66% of Cl, was used in both soil and plant material analysis. The mean recovery of Cl was higher than 92%, assuring that the extraction and determination used for Cl were effective for both soil and plant material.Chemical analysis of the beans and coffee sensorial analysis−1) was determined after 5 h of soaking [−1 g−1) was determined according to Loeffler et al. [−1) was done according to Carvalho et al. [−1 g−1) was determined according to Carvalho et al. [After benefiting the coffee samples, the beans were stored in paper bags in a cold chamber until the chemical and sensorial analyses. The chemical analyses were performed at the Laboratory of Coffee Quality Analysis in the Empresa de Pesquisa Agropecuária de Minas Gerais (EPAMIG). Potassium leaching  was performed at the Laboratory of Agricultural Products Processing in the Universidade Federal de Lavras following the Specialty Coffee Association of America (SCAA) protocol. Three professionals with skills to differentiate fragrances, characteristics, and flavors participated in the cup test. The evaluation was based on scores given to the following attributes: fragrance/aroma, uniformity, clean cup, sweetness, flavor, acidity, body, aftertaste, balance, defects, and overall. The coffees were classified as the SCAA  accordinStatistical analysesp < 0.05), the response variables were submitted to Tukey’s test (p < 0.05) on the R 3.3.1 environment [After model validation and analysis of variance indicating differences among treatments (ironment . Princip2SO4 blends influenced coffee yield and the optimum proportion was 25% of KCl and 75% of K2SO4. The highest score in the cup quality test was observed with 100% K2SO4. However, other blends showed close scores. The decision for the fertilizer should consider the cost of the K source. KL and EC can indirectly show that the Cl can damage the coffee beans and reduce the selective permeability of the cell membrane, with possible negative consequences to the coffee beverage.The activity of the polyphenol oxidase enzyme and the cup quality indicate that the ion Cl- reduces the quality of the coffee beverage. The increased application of the Cl- ion increases KL, EC, and TTA, indicators of the loss of coffee quality. K content in the leaves was not influenced by the application of blends of K fertilizer while Cl content increased linearly with KCl applied. The application of KCl and K


Objective To describe the clinical results of patients admitted and managed as cases of placenta accreta spectrum (PAS) at a Central American public hospital and the influence of the prenatal diagnosis on the condition.Materials and Methods A retrospective analysis of PAS patients treated at Hospital Bertha Calderón Roque, in Managua, Nicaragua, between June 2017 and September 2021. The diagnostic criteria used were those of the International Federation of Gynecology and Obstetrics . The population was divided into patients with a prenatal ultrasonographic diagnosis of PAS (group 1) and those whose the diagnosis of PAS was established at the time of the caesarean section (group 2).Results: During the search, we found 103 cases with a histological and/or clinical diagnosis of PAS; groups 1 and 2 were composed of 51 and 52 patients respectively. Regarding the clinical results of both groups, the patients in group 1 presented a lower frequency of transfusions (56.9% versus 96.1% in group 2), use of a lower number of red blood cell units (RBCUs) among those undergoing transfusions , and lower frequency of 4 or more RBCU transfusions (29.4% versus 46.1% in group 2). Group 1 also exhibited a non-significant trend toward a lower volume of blood loss , and lower requirement of pelvic packing (1.9% versus 7.7% in group 2).Conclusion Establishing a prenatal diagnosis of PAS is related to a lower frequency of transfusions. We observed a high frequency of prenatal diagnostic failures of PAS. It is a priority to improve prenatal detection of this disease. Placenta accreta spectrum (PAS) is a condition associated to massive hemorrhage and polytransfusion,p < 0.05. The analyses were performed using the STATA  software, version 14.A retrospective analysis of medical records was carried out in search for patients with PAS treated at Hospital Bertha Calderón Roque, in Managua, Nicaragua, between June 2017 and December 2021. The diagnostic criteria used was those of the International Federation of Gynecology and Obstetrics .During the study period, 114 women with a histological and/or clinical diagnosis of PAS were found: 51 patients had a prenatal PAS diagnosis by ultrasound (group 1), and 63 were only diagnosed when they underwent laparotomy (group 2).Less than half of our cases (44.7%) had a prenatal PAS diagnosis (group 1). Patients in group 1 had a lower frequency of transfusions and those among them who received blood components required a lower number of RBCUs. Although some expert groups have reported excellent performance of the PAS ultrasonographic diagnosis,One of the advantages of knowing the PAS diagnosis is the possibility of “scheduling” the participation of the interdisciplinary groups during surgery.Our hospital is a reference center for the most critical obstetric conditions in the country, but like many other hospitals with similar characteristics, it does not have a PAS team.The present study has limitations. Its retrospective design makes it more susceptible to bias. The absence of histological confirmation in 30.7% of the cases enabled the inclusion of non-PAS cases; however, patients whose medical record described macroscopic findings compatible with the FIGO definition were included. The present is the first evaluation of the clinical results of PAS management in Nicaragua, and one of the few that have been carried out in Central America.The results shed light on the need to design improvement plans at the national level, with the need for multicenter prospective studies to confirm our observations and evaluate the effect of interventions already implemented, such as the creation of a PAS team that is provided with specific training in the management of this disease, the proposal of including PAS screening in routine prenatal care appointments for women with risk factors, and the project of including a section about PAS in the national guidelines for the treatment of postpartum hemorrhage.The presence of a prenatal diagnosis of PAS is related to a lower frequency of RBCU transfusions. We observed a high frequency of failures within the prenatal PAS diagnostic steps. It is a priority to improve the prenatal detection of this disease.


Three-dimensional (3D) woven composites have attracted much attention in the lightweight research of protective armor due to their high specific strength and good impact resistance. However, there are still many gaps in terms of the performance and influencing factors of three-dimensional deep-angle-interlock (3DDAI) Kevlar/EP armor materials. Therefore, in order to prepare 3DDAI Kevlar/EP armor materials with excellent ballistic resistance and mechanical properties, this paper studies the bending performance of 3DDAI Kevlar/EP armor materials and the influence of the number of stacking layers, resin content, laying method, and weft density. Finally, we compare it with the traditional two-dimensional (2D) plain laminated Kevlar/EP armor material. The results showed that when the 3DDAI Kevlar/EP armor material was subjected to bending load, the upper and bottom layers of the material had a great influence on the initial stiffness and fracture strength of the material, respectively; when the material’s warp and weft density are quite different, the utilization rate of the yarn and the strength of the material are negatively affected; the fracture energy of the 3DDAI Kevlar/EP armor material prepared by the orthogonal laying method was about 20% higher than that of the 3DDAI Kevlar/EP armor material with the unidirectional layering method; and the bending performance of the 3DDAI Kevlar/EP armor material in the weft direction was better than that of the 2D plain laminated Kevlar/EP armor material, with the 3DDAI Kevlar/EP armor material having better delamination resistance. The research results will lay the foundation for structural optimization and engineering applications of such materials. Fiber-reinforced composite materials are widely used in aerospace, transportation, military protection, and other fields due to their low density and high strength ,2,3. In Three-dimensional (3D) woven composite materials outperform the traditional two-dimensional (2D) fabrics laminated to prepare laminate armor , in termDue to the limitations of weaving technology, the thickness of a single 3D fabric is often limited to 1–6 mm ,18,19, aTherefore, to prepare three-dimensional deep-angle-interlock (3DDAI) Kevlar/EP armor materials with excellent mechanical and ballistic properties, in this paper, 3DDAI Kevlar fabric was used as the reinforcement and epoxy resin was used as the matrix to study the influence of the bending properties. These include the number of stacking layers, resin content, fabric weft density, and laying method. The work presented in this study lays a foundation for subsequent research on the ballistic resistance of 3DDAI Kevlar/EP armor materials.The 3D deep-angle-interlock Kevlar fabrics ; the bending strain was calculated using Equation (4); and the bending modulus was calculated using Equation (5).The 3DDAI fabric with a weft density of 50 picks/cm was used as the reinforcement, and armor materials (resin content 44% ± 1%) of different fabric pieces were prepared through the superimposition of the unidirectional laying method, including 3DDAI Kevlar/EP armor materials made of single, two, three, and four fabric laminates. The warp and weft directions of the material were tested for bending performance, respectively.The bending stress-strain curves of 3DDAI Kevlar/EP armor materials with different fabric layers are shown in It can be seen from In order to further study the bending characteristics of multilayer 3DDAI armor materials, taking 4 layers of 3DDI armor materials as an example, the damage morphology was analyzed by a LEICASAPO stereo microscope. Among them, the slope of the curve was calculated with a strain of 0.002 mm/mm as the limit, and the test was stopped at the point of sudden change of the slope to get the failure morphology of the material at different stages, which is shown in The bending properties of the 3DDAI Kevlar/EP armor materials with different superimposed layers are listed in It can be seen from It can be seen from The 3DDAI fabric with a weft density of 50 picks/cm was the reinforcement, and the armor material, with different contents of epoxy resin, was prepared by the symmetrical laying method, including resin contents of 34.27%, 36.75%, 40.39%, 44.61%, and 48.59% of the 3DDAI Kevlar/EP armor materials were tested in the warp and weft directions, respectively.The bending properties of 3DDAI Kevlar/EP armor materials with different resin content are listed in As can be seen from As can be seen from In the literature on woven and woven-composite ballistic materials, it was found that the ballistic performance of quasi-isotropic materials in the macroscopic plane was better than that of anisotropic materials in the macroscopic plane ,29,30. TThe 3DDAI fabric, with a weft density of 50 picks/cm, was used as the reinforcement and was combined with the resin system to prepare armor materials with different laying methods. 3DDAI Kevlar/EP armor materials (resin content 44% ± 1%) was prepared using the unidirectional laying method, the orthogonal laying method, the symmetrical laying method, and the 2/2 laying method, and bending performance tests were conducted along the warp and weft directions, respectively. From The bending properties of the 3DDAI Kevlar/EP armor materials with different laying methods are listed in CAQ is the Coefficient of ascension, CDQ is the coefficient of difference, Q1 is the bending performance of the material along the warp direction, Q2 is the bending performance of the material along the weft direction, warpQ is the bending performance of the unidirectional laying material along the warp direction, weftQ is the bending performance of the unidirectional laying material along the weft direction.In order to make the material appear quasi-isotropic in the macroscopic plane, the mechanical properties of the material in different directions are improved by changing the laying method. However, as was described above, there were still anisotropic features in the material when bearing the bending load after changing the method of laying. Therefore, we used Equation (6), the Coefficient of Ascension (CA), to express the comprehensive bending performance of the material relative to the unidirectional laying material, and the difference between the bending properties of the material in different directions was expressed by Equation (7), the Coefficient of Difference . A comparison of the bending coefficients of the 3DDAI Kevlar and EP armor materials in different laying methods was made, as shown in As shown in As noted above, the preparation of 3DDAI Kevlar/EP armor materials in an orthogonal laying method was conducive to maximizing the potential to improve the comprehensive bending performance of the material, while reducing the difference in bending performance between different directions of the material. Therefore, armor materials were mainly prepared by the orthogonal laying method to study the effect of different fabric weft densities on the bending properties of the materials.The 3DDAI fabrics with weft densities of 43, 46, and 50 picks/cm were used as reinforcements to prepare armor materials (resin content 46% ± 1%), and bending performance tests were carried out along the material’s warp and weft direction. The bending properties of the 3DDAI Kevlar/EP armor materials with different weft densities are listed in As can be seen from As can be seen from Many studies ,9,10,34 For further investigation, the bending failure morphology of the 2D plain laminated Kevlar/EP armor material in This indicates that the 3DDAI Kevlar/EP armor material has better in-plane performance in the weft direction. Additionally, since the 3DDAI Kevlar/EP armor material along the weft direction of the axial yarn (weft yarn) was actually larger than the 2D plain weave laminated Kevlar/EP armor material, the bending performance of the 3DDAI Kevlar/EP armor material was superior to that of the 2D plain laminated Kevlar/EP armor material.(1)When 3DDAI Kevlar/EP armor material is subjected to bending load, the upper and bottom layers of the material become the main carrying bodies, which has a greater impact on the initial stiffness and breaking strength of the material, respectively. Wherein the bending response of the 3DDAI Kevlar/EP armor material was nonlinear, the damage of the upper layer in the axial yarn and the damage of the matrix leads to the phenomenon of bending softening, and the fracture of the axial yarn in the bottom layer is the main cause of the material failure.(2)Due to the particularity of the 3DDAI fabric structure, when the material’s warp and weft density are quite different, the utilization rate of the yarn and the strength will decrease. Furthermore, its loose structure needs to be appropriately increased in the resin content to prepare stable armor materials, where the appropriate range of resin content is 40%–49%. In addition, the 3DDAI Kevlar/EP armor can be prepared by the orthogonal laying method to improve the macroscopic mechanical properties of the material and effectively increase the fracture energy of the material.(3)The 3DDAI Kevlar/EP armor material was in-plane anisotropic, and its bending performance along the weft direction was better than the 2D plain laminated material. Additionally, due to the penetration of the yarns in the thickness direction in the 3DDAI structure, even the lamination can effectively slow down the delamination of the material.Based on the potential of 3DDAI Kevlar/EP armor material in the field of protection, the bending mechanics response of the 3DDAI Kevlar/EP armor material was carried out in terms of the number of fabric superimposed layers, resin content, laying method, and fabric weft density, and then compared with the bending performance of 2D plain laminated Kevlar/EP armor material. The following conclusions were obtained through research:


Spodoptera littoralis larvae (an important crop pest) in continuous light (conditions abolishing rhythms in insects by interfering with the clock) and altered the performance of their oscillator using molecular biology tools. We have shown that the susceptibility of S. littoralis larvae to insecticides varies daily, being under the control of a molecular oscillator. This oscillator operates in the fat body, midgut, and Malpighian tubules where it is likely to control the activity of detoxifying enzymes. Our results indicate the role of a molecular oscillator in the metabolism of xenobiotics in S. littoralis larvae. These findings support the rational use of insecticides at specific times of the day.The oscillator, the core element of the biological clock, is a mechanism operating at the molecular level that is responsible for the generation of rhythms. It is produced by clock genes that code for proteins, regulating expression of its own and executive genes. Oscillator activity manifests itself at the level of the cells, organs, and the whole organism, thus enabling organisms to perform life functions at the most proper time. An example is the intensification of detoxification processes when the body is exposed to xenobiotics. In this study, we bred S. littoralis larvae by injecting the dsRNA of clock genes into their body cavity and culturing the larvae under continuous light. As a result, the daily susceptibility of larvae to insecticides was abolished and the susceptibility itself increased (in most cases). In the fat body, midgut, and Malpighian tubules (the main organs metabolizing xenobiotics) of the larvae treated with injected-dsRNA, the daily activity profiles of enzymes involved in detoxification—cytochrome P450 monooxygenases, Glutathione-S-transferase, and esterase—have changed significantly. The presented results prove the role of the molecular oscillator in the regulation of larvae responses to insecticides and provide grounds for rational use of these compounds (at suitable times of the day), and may indicate clock genes as potential targets of molecular manipulation to produce plant protection compounds based on the RNAi method.The molecular oscillator is the core of the biological clock and is formed by genes and proteins whose cyclic expression is regulated in the transcriptional-translational feedback loops (TTFLs). Proteins of the TTFLs are regulators of both their own and executive genes involved in the control of many processes in insects . We disrupted the clock operation in  Drosophila melanogaster, which consists of two interlocked TTFLs [period (per) and timeless (tim) genes during the solar day. The transcripts are then translated into proteins, PER and TIM, which bind to each other and enter the cell nuclei where they bind to CLK:CYC complexes to suppress their own expression after dusk. This system is reset by light via the cellular photoreceptor CRYPTOCHROME 1 (CRY1), which directs TIM down the degradation pathway , thus leading to CLK-CYC dependent re-initiation of transcription of per and tim after dawn. In the second loop, clk gene transcription is regulated by two factors: positively by PAR-Domain Protein 1 (PDP1) and negatively by VRILLE (VRI). In response, CLK-CYC positively regulates the transcription of the Pdp1 and vri genes. The supporting role of the molecular oscillator system is played by the clockwork orange factor (CWO), which is a transcriptional repressor that competes with CLK-CYC for binding sites in clock-controlled gene regulatory sequences, including the temporary inhibition of its own expression [The biological clock has been revealed to have activity on several levels of biological organization from the molecular to organismal level, thus controlling multiple aspects of life ,2. The ced TTFLs ,6. The fpression .Drosophila’s CRY1) plays the role of the cellular photoreceptor, while CRY2  acts as the major repressor of transcription in the first TTFL of the oscillator [per, tim, and cry2 genes. Much less is known about the second TTFL of the molecular oscillator in lepidopterans. Most of the available data comes from transcriptomic analyses, which indicate the expression of vri and pdp1 in certain tissues and also the rhythmic nature of this expression [vri and pdp1 genes code for transcription factors, attention is focused not only on their role in the molecular oscillator system itself, but also on the regulation of physiological and developmental processes, both in terms of circadian control and ontogenesis [cwo gene has also been associated with the lepidopteran response to exogenous factors altering homeostasis [The molecular oscillator of lepidopterans is structurally similar to the oscillator of the fruit fly. However, it also has its own characteristics. It comprises two CRY proteins (CRY1 and 2) which have immensely distinct physiological functions, while still maintaining the functionality of the TTFL ,9,10,11.cillator ,13. In tpression ,15,16. Aogenesis ,18,19. Meostasis .In insects, expression of genes encoding factors acting in TTFLs was found in the cells of several organs, which indicates that a complex multi-oscillator system functions in these animals ,22,23. Lpdp1 and cyc D. melanogaster mutants has been shown to affect detoxification pathways by disorganizing rhythmic expression of genes for P450s and specific esterases, leading to changes of sensitivity to permethrin and malathion in such mutants [per and tim genes) with detoxification pathways was not observed. These findings are confirmed by earlier reports indicating the rhythmic activity of uridine 5’-diphosphoglucosyltransferase (UGT) and P450 (determined by indirect measurements of ethoxycoumarin-O-deethylase activity—ECOD), which were disturbed in insects kept under constant light (conditions abolishing functions of molecular oscillator) [A. aegypti [A very important aspect of the biological clock’s role in maintaining homeostasis is the control of the response to stresses, including xenobiotics. Insects have a tremendous impact on ecosystems and the economy. However, research on the daily (and circadian) control of xenobiotic metabolism in insects, including insecticides, is still scarce in comparison to studies done on vertebrates ,41,42,43 mutants . At the  aegypti . These tSpodoptera litura. This was the first report in which these issues were addressed specifically for lepidopterans [Spodoptera littoralis, one of the most destructive pests in vegetables, fruits, and other crops. We disrupted the operation of the clock in the last instar larvae of S. littoralis, by keeping them under continuous light followed by determination of susceptibility to commonly used insecticides. Then, using RNAi technique, we knocked down operation of the molecular oscillator in these larvae and analysed the daily activity of xenobiotic metabolizing enzymes in their fat body, midgut, and Malpighian tubules, as well as the daily susceptibility to insecticides.However, these studies are limited to two species classified under the same order, Diptera, of which only one is of economic importance. During the preparation of the present manuscript, data was published on the influence of a molecular oscillator on the expression of genes encoding digestion and detoxification enzymes in the lepidopteran representative, opterans . Our resS. littoralis were reared as described previously [2 atmosphere; in addition, insect dissections were performed in ice-cold buffer.Larvae of eviously , in cycleviously  for deta® GC Ultra Grade . Insects were treated with insecticides as described previously for D. melanogaster [50) was calculated using probit analysis using LeOra Software POLO-Plus [50 values for each time point and each treatment was performed in three replicates.Fipronil, deltamethrin, malathion, propoxur, acetamiprid, and imidacloprid were obtained from Greyhound  and dissolved in acetone—ROTISOLVnogaster  with sliOLO-Plus . Tests wperiod (per), timeless (tim), cryptochrome 2 (cry2), cycle (cyc), vrille (vri), par-domain protein 1 (pdp1), or plant  dsRNA in 4 μL of transcription buffer. Larvae injected with 4 μL of transcription buffer and untreated larvae (intacts) were used as additional controls. The solutions were injected by inserting a sharpened glass capillary  into the body cavity through one proleg of the second pair. Injections were started at Zt 0 and given every 4 h until the beginning of the next day (Zt 0 of the 3rd day of the last instar)  C. Ten toper, tim, cry2, cyc, vri, pdp1, and SSU (plant) was produced as described previously [cyc and pdp1 genes were cloned , were injected into the body cavity of the larvae. Then the mRNA level of targeted genes were determined in the organs as described in the The next step was to abolish the oscillator’s operation in larvae using the RNAi technique followed by an analysis of detoxifying enzyme  activities in the fat body, midgut, and Malpighian tubules—the organs mainly responsible for the metabolism of xenobiotics. The dsRNAs produced by in vitro transcription, based on the coding fragments of selected g and the pellet was discarded. The solution was centrifuged a second time at 100,000× g and the supernatant was discarded. The pellet obtained with the microsomal fraction was suspended in the buffer previously used to homogenize tissues and protein concentration was determined. The reaction mixture containing the microsomal fraction (250 µg proteins), 0.4 mM 7-ethoxy-2H-1-benzopyran-2-one, and 1 mM NADPH  in a final volume of 0.5 mL PBS was placed in 1.5 mL tubes and incubated for 30 min at 30 °C. For triplicate measurements, 150 µL of the reaction mixture was applied to three wells of a Nunc™ F96 MicroWell™ Black Polystyrene Plate . The fluorescence was measured at 380 nm excitation and 450 nm emission using a VICTOR X3 Multilabel Plate Reader . The amount of 7-hydroxycoumarin formed by ECOD was determined with a standard curve established from known amounts of 7-hydroxycoumarin. The ECOD activity was expressed as the nmol of product formed per minute per mg of protein.The total cytochrome P450 monooxygenase activity was measured as described previously ,52, withg then 20 min at 10,000× g. The pellets were discarded and the concentration of proteins in the second supernatant was determined. Twenty µL of diluted supernatant (1:5 with PBS-K) was applied to the well of a Greiner UV-Star® 96 well plate (Greiner Bio-One GmbH) containing 170 µL of PBS-K and 20 µL 5 mM glutathione (GSH), followed by the addition of 10 µL of 1.2 mM 1- chloro-2,4-dinitrobenzidine (CDNB). Enzyme activity was measured using the TECAN Infinite M200 microplate reader  at 340 nm and 30 °C for 10 min using the kinetic mode. GST activity was calculated using the Beer–Lambert Law where ε340 = 9.6 mM–1 · cm–1. The obtained results were presented as the nmol of glutathione conjugate formed per minute per mg of protein.The GST activity was determined according to procedures described previously ,56. OrgaS. littoralis in the context of insecticide metabolism [g and protein concentration was determined. Equal volumes (120 µL) of supernatant and 0.1 mM α-naphthyl acetate on PBS-K were mixed in a 0.5 mL test tube and incubated for 30 min at 35 °C. For the controls, the protein extracts from the organs were replaced with an equal volume of PBS-K. Then, 120 µL of 0.3% Fast Blue RR Salt  dissolved in double distilled water containing 5% SDS was added to stop the reaction. Ten minutes after development of color, 100 µL of reaction mixture was pipetted into three consecutive wells of the 96 well polystyrene microplate (Greiner Bio-One GmbH) to measure esterase activity in each organ in triplicate. The plates were read at 595 nm in VICTOR X3 Multilabel Plate Reader and the concentration of products was determined from standard curves of α-naphthol. The esterase activity was expressed as the nmol of product formed per minute per mg of protein.Esterase activity was measured according to the procedure described previously and used for studies of lepidopterans, including tabolism ,58, withAs all enzymatic assays required determination of the total protein level in the samples, a commercial BCA Protein Assay Kit was used for this purpose in accordance with the manufacturer’s instructions . This procedure included the standard curve determination on quantitatively standardized bovine serum albumin solutions for estimation of protein concentration.The average LC50 for fipronil changed over 24 h for LD larvae, with the highest values observed in the evening (Zt 8), followed by a drop to the lowest value at the time of transition from night to day (Zt 20), and then an increase during the day A. It is For deltamethrin, the average LC50 values for the LD group also changed cyclically during the day, with a significant increase during the night hours, reaching the maximum value at the time of transition from night to day (Zt 20) followed by a decrease in the morning and reaching a minimum in the middle of the day (Zt 4) B. It shoAnother rhythm profiles the diurnal changes in malathion sensitivity in LD larvae . A signiThe highest average LC50 values for propoxur administered to LD larvae were also recorded at night D. LC50 wDaily differences in the average LC50 values for acetamiprid administered to LD group were significant. The highest LC50 values for this insecticide were recorded in the larvae treated in the evening (Zt 8), while the lowest values were seen when larvae were treated at the end of the night (Zt 20); the difference between these values was two and a half times E. AlthouThe difference between the minimum and maximum average LC50 for imidacloprid applied in larvae was nearly two times , and as with other insecticides tested, the larvae showed diurnal differences in susceptibility to this compound F. FurtheIt was observed in the fat body of larvae from the control groups that the expression of all clock genes studied changed every 24 h . The proper, cyc, and pdp1 genes, the mean level of fat body ECOD activity at each time point was very low throughout the entire day and significantly lower at most time points than in the tissues of animals from the control group  during the daytime when compared to the levels observed at the same daytime points in the control . In larvae injected with tim dsRNA, the level of GST activity changed rhythmically  and an end-of-night maximum (Zt 20) (cry2 dsRNA larvae recorded during daytime (between Zt 0/24 to Zt 12) were about five to seven times higher at the same time points when compared with insects from the control groups. In insects injected with both cyc and pdp1 dsRNA, the activity of GST was very low throughout the day and remained at the same level as in the daytime hours of insects from the control groups  H. These  (Zt 20) . In thisl groups I,J. The l groups .per, tim, and cry2 dsRNA, the mean esterase activity was markedly elevated when compared with that of the control group (at each time point of the 24 h daily cycle) . Conversely, when the larvae were treated with cyc and pdp1, the mean level of esterase activity in the fat body was lowered at most time points of the 24 h diurnal cycle when compared with the values found for the control insects  with a gradual increasing to a maximum (more than eight-fold) at the end of the night (Zt 20) K. Neverty cycle) K–M. These groups , althoug insects N,O. In t insects .per gene, the expression of all analyzed clock genes in the midguts of larvae from the control groups had features of the rhythm , decreasing by nearly nine times, with cyc dsRNA by more than thirteen times at midday (Zt 4), and with pdp1 dsRNA by more than twelve times in the evening (Zt 8)  the level of midgut ECOD activity. Additionally, in both cases, the activity remained at a comparable level for 24 h  A,D,E. Neg (Zt 8) . In turnfor 24 h B,C, whicfor 24 h .per dsRNA group, we observed an increase in the average GST activity in the midgut at almost all time points as compared to the control   and a maximum at the end of the night (Zt 20) . As shown by the cosinor analyses for the larvae of both groups, the daily changes in GST activity were rhythmic , we observed a decrease in activity in larvae treated with dsRNA, by nearly five times (at Zt 20 in cyc dsRNA larvae), and even more than five and a half times (at Zt 20 in pdp1 dsRNA larvae). We also observed an obvious flattening of the daily GST activity profile in the midgut in the larvae from these groups, which is reflected in the lack of rhythm of this process (The activity of GST in the midgut of larvae from control groups changed over a 24 h cycle F. Its avin Zt 4) F. The acin Zt 4) . In the  (Zt 20) G,H. In trhythmic . In the  process .per, tim, and cry2, the midgut esterase activity of these larvae was significantly higher  when compared to insects from the control group . The opposite situation was found in larvae treated with cyc and pdp1 dsRNAs when compared to the control, because in both cases, a decrease in the mean activity of esterase in midgut at almost every time point of the day was observed . Moreover, in each group of larvae treated with clock gene dsRNA, there was a flattening of the 24 h profile of esterase activity in the midgut, which made the process lose its rhythmic character .The average activity of esterase in the midgut of the larvae in the control groups changed in the 24 h mode, showing a clear minimum during the daytime (between Zt 4 and Zt 8), a successive increase in the evening (Zt 8) and throughout the night, until the first hours of the next day (Zt 24/0–0/24), at which point it reaches its maximum and then drops rapidly K. This iol group K–M. In tobserved N,O. The haracter . Such laper, cry2, cyc, and pdp1, their expression dropped to low values, which led to a more or less pronounced flattening of their profiles and loss of the rhythm of expression of these genes in the treated insects , and this state is maintained throughout the whole day  and increasing until the middle of the night, followed by a rapid decrease to a minimum by the next morning (Zt 20–24/0) A. The dihole day A,D,E. Th tubules . In cont tubules B,C. The t rhythm .per and cry2 dsRNA to larvae had a similar effect. Both treatments resulted in a nearly constant level of GST activity in the Malpighian tubules during the day  in pdp1 dsRNA insects. Moreover, the flattening of the daily GST activity profile observed in both cases resulted in the process being non-rhythmic  level in the morning and in the first half of the day (Zt 0/24–4), then dropping to a minimum in the evening (Zt 8), and rapidly increasing to the highest values in the first hours of the night (Zt 12–16) F. In the the day F,H. More the day . In turnaffected G. The praffected . A very th dsRNA I,J. The rhythmic .per, tim and cry2, led to two changes in the average esterase activity in Malpighian tubules: (1) a slight increase of this activity in the course of the day (since application of the dsRNAs), and (2) a significantly higher activity in the daytime when compared to the activity in the control   K–M. As a tubules . When cynd night N,O. Comp control .pdp1 and cyc dsRNA was radically reduced and non-rhythmic. Such a regular pattern of responses to specific dsRNAs was not found in the case of other enzymes. The disruption of expression of both per and cry2 appeared to have opposite effects on GST and esterase activity than pdp1 and cyc. After knockdown of the per and cry2 genes, GST and esterase activity increased in organs, but at some time points of the day, its value remained comparable to the control values. On the other hand, in the case of ECOD, it was difficult to find a pattern between the treatment of insects with dsRNAs and the enzymatic activity in their fat body, midgut, and Malpighian tubules. Knock down of tim and cry2 genes expression using dsRNA did not seem to affect the rhythmic activity of ECOD in fat body and Malpighian tubules, while dsRNA of the per gene abolished its rhythm and lowered its level in all of the examined organs. However, in the midgut, the dsRNA of the tim gene did not affect the activity of ECOD, and the cry2 dsRNA significantly increased it.The daily activity of the tested detoxification enzymes in all examined organs in insects treated with per dsRNA to larvae significantly reduced the average LC50 for fipronil at almost every time point of the day to a value close to the lowest value described for insects from the control group, ultimately abolishing the fipronil susceptibility rhythm (expressed as changes in the LC50) in such treated larvae , which was more than three and a half times higher than the lowest observed at the end of the night (Zt 20) , and which was comparable to the lowest value found for control insects at the end of night (Zt 20). In insects from the pdp1 dsRNA group, these differences were even more significant. The largest reduction in the LC50 value, in relation to the LC50 in control, was found during the evening hours (Zt 8), when it was nearly eleven times that of the control, and the lowest at the end of the night (Zt 20), when it was approximately twice that of the control  and the lowest value at the end of the night (Zt 20), and the difference between the two was more than three-fold A. As a rd larvae A. On the (Zt 20) A. As res (Zt 20) . Injecti control A. This ss groups .tim and cry2 dsRNA insects (same as in control) was there a rapid increase from the middle of the night (Zt 16) to the maximum at the end of the night (Zt 20)  when compared to the control, by more than four times, seven times, and 240 times for per, cyc, and pdpi1 dsRNA insects, respectively  in control larvae changed cyclically over 24 h with a trough in the evening (Zt 8) and a peak at the end of the night (Zt 20); the difference was more than three-fold B. The co (Zt 20) B. Such aectively B. Furthe50 value . This rhper, cyc, and pdp1 dsRNA at each time point of the day when compared to the control values. The greatest differences were recorded at the turn of the day and night (Zt 12) with a value that was almost sixteen, eighteen, and eleven times lower than in the control for the per, cyc, and pdp1 dsRNA-treated larvae, respectively  and the highest at the beginning of the night (Zt 12), and the difference was more than three-fold C. Moreovectively C. This ve larvae . The sit control C. Thus, ecticide .per, cyc, and pdp1 dsRNA, in addition to propoxur, the mean LC50 values for this insecticide were lower (without exception) at each time point when compared with the LC50 values for the control. and the greatest differences were noted at the beginning of the night (Zt 12), with nearly twenty-six-fold difference for per dsRNA, twenty-eight-fold difference for cyc dsRNA, and nearly nineteen-fold difference for pdp1 dsRNA larvae  and at the beginning of the night (Zt 12), LC50 values were lower than in other time points of the day  for propoxur, which would indicate diurnal differences in susceptibility to this compound  and the highest at midnight (Zt 16), with a difference of almost two-fold in both cases. In addition, cosinor analysis of the LD50 values from propoxur exposure indicated the maintenance of susceptibility rhythms in tim and cry2 dsRNA larvae to propoxur  and the highest at the beginning of the night (Zt 12), with a difference of a little more than two-fold D. The daA larvae D. While  the day D. Howevecompound . In larvpropoxur .per, tim, and cry2 genes resulted in a significant elevation of the LC50 value for this insecticide. The highest difference in values of LC50 for acetamiprid between the control and per and tim dsRNA insects were found at the end of the night (Zt 20), and in the morning for cry2 dsRNA insects (Zt 24/0). These differences were more than ten-fold, eleven-fold, and nineteen-fold, respectively  that of the control; while LC50 in the pdp1 dsRNA group was lower at all time points than in the control .Response of insects from the control group to acetamiprid varied over 24 h E. As expectively E. It shoectively . Similar control E. This dper, tim, and cry2 dsRNA resulted in a significant increase of the LC50 value for imidacloprid-treated larvae treated for most of the day, with exceptions for per and tim larvae treated in the middle of the day (Zt 4) and in the evening (Zt 8), for which LC50 for imidacloprid was similar to that of the control. The highest (about three times) increase of LC50 after the injection of per and tim dsRNA in relation to the control was found at the beginning of the night (Zt 12) and after cry2 dsRNA injection (more than five times) in the morning (Zt 0/24)  by more than four-fold and twenty-six-fold, respectively.Analysis revealed diurnal differences in the control larvae response to imidacloprid. The highest LC50 value from imidacloprid exposure was found in the evening (Zt 8) and the lowest at the beginning of the night (Zt 12), with the difference being slightly more than two-fold F. These Zt 0/24) F. No cycZt 0/24) . The com control F. The maS. littoralis larvae on their susceptibility to insecticides. When looking for the TTFL components of the oscillator that are particularly important in response to insecticides, pdp1 and cyc are noteworthy. The knockdown of the expression of these two genes resulted in a radical increase in the susceptibility of larvae to all tested insecticides. Such a strong effect in response to all insecticides was not found when the expression of the per gene is lost. However, it was evident with fipronil, deltamethrin, malathion, and propoxur. An analogous relationship need not necessarily be the case for the rest of the oscillator genes when they are knocked down. In some cases, as indicated by the studies shown above, the tendency was even opposite—disturbance in the expression of clock genes per, cry2, and tim led to a very significant decrease in insect susceptibility to selected insecticides—such a relationship was evident in the case of neonicotinoids (acetamiprid and imidacloprid). However, even if susceptibility to selected insecticides decreased after knockdown of clock genes, insects treated in this way have a disturbed rhythm of response to these compounds. The results indicated that the abolition of the smooth operation of the oscillator in S. littoralis larvae may significantly impair their metabolic response to xenobiotics.Drawing general conclusions, our observations indicated a clear influence of the molecular oscillator of S. littoralis larvae are less susceptible to insecticides if their biological clock is functioning properly. On the other hand, if we disrupted the larvae’s clock by exposure to constant light (numerous rhythmic processes in moths are abolished under these conditions [Locusta migratoria migratorioides larvae that were more susceptible to this insecticide when dosed during the night [D. melanogaster reared under constant light (LL) [D. melanogaster. Based on LC50 analyses, D. melanogaster had the lowest susceptibility to propoxur during the day, whereas susceptibility was lowest for S. littoralis at night.The realization of the vital functions of each organism at the right time of the day is the key to the efficient course of physiological processes maintaining homeostasis. Such processes are aimed at optimizing the use of energy necessary for steady state maintenance, and are accomplished by an efficiently operating biological clock mechanism. Among the numerous biological clock-dependent processes, those related to the response to compounds that are not physiologically present in the organism and delivered from the environment (xenobiotics) have a particularly high adaptive value. It has been proven that the disturbance of the clock functions has an extremely destructive effect on the body through inefficient metabolism and excretion of xenobiotics, which, when remaining in a biologically active form, threaten the homeostasis of many organs and body systems. The results presented here are an example of the role of the biological clock in minimizing the negative impact of xenobiotics on the organism. We found that nditions ), their nditions . Analysihe night . Similarght (LL) . HoweverApis mellifera [S. littoralis, where daily profiles of larvae susceptibility to individual insecticides varied greatly throughout the day. Interestingly, susceptibility to diazinon (organophosphate) in cotton aphid Aphis gossypii was shown as changing during the day with a fashion similar to what we observed in S. littoralis treated with malathion  [S. littoralis. The response to pyrethoids seemed to be similar in S littoralis and in another moth, B. mori, to deltamethrin and permethrin, respectively [S. littoralis, B. mori also had the lowest susceptibility to these insecticides in the second half of the night, and the highest during the day. Crickets, Acheta domesticus, also showed the highest resistance to the pyrethroid ß-cyfluthrin at night, expressed as a high survival rate [Aedes aegypti [Anopheles gambiae. Cyclic changes of numerous genes expression which are involved in the metabolism of DDT and pyrethroids were shown, and both were determined at the level of the transcripts [Drosophila suzukii population to malathion (organophosphate), and lack of such a response rhythm to fenpropathrin (pyrethroid), while indicating that the expression of a number of genes for detoxification factors is rhythmic in this species [Cimex lectularius, susceptibility to pyrethroids and neonicotynoids was also studied in the context of the circadian regulation of genes for GSTs and P450s, linking observations with the daily activity of these enzymes [Triatoma infestans, a vector of Trypanosoma cruzi causing Chagas disease [S. littoralis, the application of the RNAi technique with the use of dsRNA turns out to be effective [per, tim, cry2, and cyc) and one so-called supporting TTFL (pdp1); cyc and pdp1 occupy a special position, as both encode factors involved in the regulation of cellular responses to xenobiotics. The use of dsRNA not only destroyed the rhythmic expression of the clock genes, but also very significantly lowered the level of transcripts in the studied organs. This effect on clock genes expression has an influence on the daily profiles of enzymatic activity related to detoxification  in all examined organs. Nevertheless, this effect very much depended on which gene expression was influenced and which organ was tested. We can conclude that dysfunction of per expression lowers the activity of P450 and increases GST and esterase in all examined organs. Disturbance of tim expression, in principle, has little or no effect on the function of P450 and GST, but essentially increases esterase activity in the fat body and midgut. On the other hand, knockout of cry2 expression does not change P450 activity in the fat body and Malpighian tubules, and increases it in the midgut. In the same larvae, an increase is also observed in the GST and esterase activity in the midgut and Malpighian tubules. We observed a very homogeneous effect after administering the cyc and pdp1 dsRNA to insects. Without exception, this led to a decrease in the activity level of each of the tested enzymes in each of the examined organs. Moreover, despite the knockout of per, tim, and cry2 expression, in some organs, the rhythmic activity of the tested enzymes was maintained, especially after injection of tim dsRNA into larvae. However, the enzymatic activities tested were not found to have signs of biological rhythm in any of the organs when the expression of cyc and pdp1 was disturbed. This may be explained by the high activity of enzymes around the clock in insects that were injected with dsRNA of genes encoding per and cry2 due to subordination of genes encoding these enzymes to PER and CRY2 proteins. This relationship would have the form of negative regulation. Thus, the lack of factors encoded by per and cry2 enables the expression of detoxification genes during this phase of the day when its level normally remains low. On the other hand, the persistently low level of enzyme activity in cyc and pdp1 larvae indicates that both TTFL elements considered as positive regulators of transcription are necessary to maintain gene expression for detoxification enzymes at an elevated level at certain times of the day. The clearly less significant effect of the tim dsRNA on the enzymatic activities studied proves the minor importance of this gene in the regulation of enzymes involved in detoxification, or more generally, in the metabolism of xenobiotics. This might not be so surprising considering the role of tim in the oscillator itself in lepidopterans, which is rather supporting and related to the information about photoperiodic status of the environment than regulation of genes expression that are dependent on the clock. To date, rhythm analysis of GST activity, esterase, and oxidase activity in the heads and whole bodies of A. gambiae indicated that only GST activity had features that allowed it to be considered rhythmic in both parts of the insect body, with a distinct peak extending for hours from late night to dawn [D. melanogaster, esterase activity, and GST were found to be non-rhythmic, but ECOD (P450) and uridine 5′-diphosphoglucosyltransferase (UGT) activities were rhythmic, with a peak in the first half of the day and the second half of the night, respectively [C. lectularius, the activity of ECOD (P450), GST, and esterase were clearly rhythmic with peaks in the evening hours, although the measurements of activity were made in protein extracts obtained from whole insects [S. litura, GST activity in the hemolymph was significantly lower when the clock was desynchronized by growing insects under constant light [A. aegypti strains was significantly higher when the insects were kept under the optimal photoperiod, and lower when transferred to permanent darkness, confirming the significant role of the oscillator in the body’s response to xenobiotics [S. litura treated with the dsRNA of clock genes [Over the last decade, studies on the daily changes in susceptibility to insecticides continue to be relevant. Oscillations in sensitivity to esfenvalerate, cyhalothrin, alpha-cypermethrin, beta-cyfluthrin, deltamethrin (pyrethroids), pyriproxyfen (derivative of pyridine), pirimicarb (carbamate), diazinon (organophosphate), indoxacarb (organochlorine), and teflubenzuron (benzoyl urea derivative) were shown in the honey bee ellifera ,63. In tosphate) . Moreoveectively . As we sval rate . The sco aegypti . Such ex aegypti ,69. In tnscripts ,71 and anscripts . Interes species . In the  enzymes . Variabl disease . Howeverffective ,50. Ther insects . Althougnt light . Additioobiotics . Our conobiotics . Among tck genes .The interest in using RNAi to protect crops by controlling insect pests is high and increasing, as reflected in the literature. The proposed target in this case are genes usually related to both development and metabolism in general ,80,81,82S. littoralis larvae to the insecticides we have tested. Several conclusions can be drawn from our results. First, the treatment of insects with per and cyc  and pdp1 (at every time point of the day) significantly lowered the LC50 for fipronil, malathion, and propoxur. Second, the susceptibility of insects (LC50) exposed to these insecticides did not change substantially over the course of the day following the injection of tim and cry2 dsRNA. Third, the effect of dsRNA treatment in insects exposed to deltamethrin was primarily observed almost exclusively during the hours when the LC50 value was highest in the control group . An exception is the administration of pdp1 dsRNA, which dramatically lowered the LC50 for deltamethrin, regardless of its administration time. Fourth, the reaction of larvae to neonicotinoids after treatment with dsRNA of genes encoding negative regulators in TTFL  was similar, i.e., the LC50 value was increased compared to that of the control larvae. However, when the dsRNA of positive regulators in TTFL (cyc and pdp1) were used, the LC50 value for these insecticides decreased or (less frequently and only in cyc dsRNA larvae) did not change. In our research, we showed that disrupting the function of a molecular oscillator by using RNAi to knock out clock genes (and therefore encoding factors that affect transcription) effectively changed the activity of detoxifying enzymes in dedicated organs (as it is mentioned above), but also affected the susceptibility of D. melanogaster mutants, and after RNAi treatment in S. litura and A. aegypti [S. littoralis larvae oscillator are particularly involved in the response to insecticides. The answer to this question is not easy, because the knockout of different genes resulted in different reactions of the larvae. However, attention may be drawn to the fact that both the susceptibility to insecticides and the activity level of the tested enzymes were decreased in each of the tested organs when pdp1 expression was disturbed; the same is true in most cases when insects were treated with cyc dsRNA. These observations suggest a very strong involvement of the second TTFL oscillator in the response to insecticides, and its dysfunction results in a decrease in insecticide resistance of the larvae. Such a suggestion is consistent with the postulate put forward by the authors of the work on D. melanogaster [pdp1 positive transcriptional regulators  produced a similar effect. Such indirect regulation dependent on factors positively regulating transcription in the TTFL seems to be a case described in S. litura, however, the RNAi knockout was performed on the cwo gene [cwo caused an increase in the expression of genes involved in detoxification, because CWO likely competes with clock transcription factors (CLK-CYC) at the binding site in regulatory regions of these genes. The undoubted importance of research in this area is also emphasized by the results of analyses conducted on mammals, indicating that the CLK and BMAL1  loop is an important point in the path of xenobiotic metabolism, coordinating efficient detoxification [per and cry2 genes—on the level of detoxification enzyme activity and insecticide susceptibility that we tested. The PER and CRY2 proteins function as a heterodimer in the TTFL loop of the oscillator. One would therefore expect a similar or even identical effect to be induced by per and cry2 dsRNA administrations. However, it must be taken into account that the organism’s response to the knock down of each of these genes does not have to manifest only in a change in the functioning of the molecular oscillator and the genes dependent on the oscillator. The PER and CRY2 proteins may also perform other functions in the body, which is the subject of numerous discussions. Especially the one concerning peripheral oscillators—e.g., the involvement of oscillator genes, especially per, in response to genotoxic stress [per and cry2 in the response to stress induced by xenobiotics. Undoubtedly, it is an excellent problem for planning research in the near future.Biological clock genes represent a specific class as, when encoding transcriptional regulatory factors, they are highly situated in the cascade of the organism’s response to noxious substances. Using the RNAi gene silencing technique, the significant role of genes that act in a such hierarchical system has also been demonstrated, which, by coding for factors regulating transcription are superior in the control of executive genes during detoxification ,91. Howe aegypti ,45,46. T aegypti ,92,93. Tnogaster . Lowerincwo gene . It has fication ,95,96. Sc stress . PerhapsThe conducted experiments led us to the conclusion that molecular oscillators of lepidopterans larvae  are responsible for the control of the xenobiotic metabolizing enzymes in organs strongly involved in detoxification, such as the fat body, midgut and Malpighian tubules. The activity of these enzymes, which changes throughout the day, is most likely responsible for the daily cycle of susceptibility of larvae to insecticides. Ultimately, we postulate that these results support the idea of the rational use of insecticides at strictly defined times of the day.


Bioscience Reports at the request of the Editor-in-Chief and the Editorial Board following receipt of a notification from several readers, alerting the Editorial Board to duplications in Figures 3C and 6D, after which a third duplication in Figure 6D was identified. Readers also identified that several panels from Figures 6C and D also appeared in the group’s previous publication . 2019 Oct 24;11(20):8951-8968. doi: 10.18632/aging.102359).This article is being retracted from The authors have contacted the Journal to correct the Article however, given they were unable to provide the original images and the extent of the issues raised, the Editorial Board stand by the decision to retract the article. The authors do not agree to the Retraction.


In the SP-ICP-MS determination of TiO2NPs in cosmetics, the background signal was recorded in each dwell time interval due to the signal of the Ti dissolved form in deionized water, and the background signal of the cosmetic matrix was compensated by dilution. A correction procedure for the frequency and intensity of the background signal is proposed, which differs from the known procedures due to its correction by the standard deviation above the background signal. Background signals were removed from the sample signal distribution using the deionized water signal distribution. Data processing was carried out using Microsoft Office Excel and SPCal software. The distributions of NP signals in cosmetic product samples were studied in the dwell time range of 4–20 ms. The limit of detection of the NP size (LODsize) with the proposed background signal correction procedure was 71 nm. For the studied samples, the LODsize did not depend on the threshold of the background signal and was determined by the sensitivity of the mass spectrometer.We discuss the features involved in determining the titanium dioxide nanoparticle (TiO The most effective UV radiation blockers are nanoparticles (NPs) in a size range of up to 100 nm [2 NPs used in the cosmetic industry are safe for direct contact with human skin [2 NPs exist [2 NPs results in various scenarios of NP interactions with the environment, for example, their release upon contact with water, changes under the influence of UV radiation, and others [Titanium dioxide , which limits its use for small TiO2 NPs [size. The LODsize depends on the detected isotopes, sample matrix, operational parameters, and other factors, and depends on the ability to distinguish the NP pulse signals from the background signals, as well as on the detector’s sensitivity to a specific element [size of TiO2 NPs was in the range of 95–130 nm, depending on the detected isotope and the threshold value of the standard deviation of the background signal. The LODsize can be improved by using a collision cell or ms/MS technology in combination with reaction cell gases to reduce matrix interference. The LODsize of TiO2 NPs of 37–55 nm was achieved by using a collision cell [size of TiO2 NPs to 30–35 [48Ti and 47Ti, respectively. In this case, the use of specialized software for data acquisition and processing is required, which allows the detection of a NP as a transient peak over several dwell times. The software allows automatic summarizing of the fragmented NP signal and it gives the total NP intensity. To date, open-source platforms are available for SP-ICP-MS analyses [On the other hand, SP-ICP-MS has a relatively high limit of detection of the NP size  is proportional to the concentration of the dissolved element, interferents in the sample matrix, and reagents used in the sample preparation. Besides TiO2 NPs, cosmetic products may contain Ti in dissolved form. In addition, cosmetic products often contain calcium compounds, and sulfur compounds are also found. 48Ca (abundance 0.19%) and 16O32S represent interfering influences in the analysis of 48Ti, increasing the continuous signal at m/z 48. Strict compliance with the conditions for registration of each single TiO2 NP separately from other NPs requires dilution of the cosmetic matrix suspension with deionized water to achieve optimal concentrations of NPs in the solvent. The effect of the suspension dilution on the background signal at m/z 48 was studied for samples 1 and 2 with typical cosmetic matrices without TiO2 NPs. The sample suspensions were successively diluted 10, 100, and 1000 times with deionized water. The signal intensity monitoring at m/z 48 showed that dilution of the sample suspension by 1000 times resulted in reducing the content of dissolved Ti and interferents in the cosmetic matrix and lowering a signal to 15 counts, which corresponds to the background signal of deionized water  allowed for characterizing the background signal more correctly . On the other hand, the increase in dwell time resulted in a broadening of the distribution. The standard deviation value of the signal σ was expected to increase exponentially with an increase in the width of the distribution, which led to an increase in the threshold of the background signal calculated by 3σ, 5σ, and 10σ criteria . The thrThe threshold calculated by 5σ correctly characterized the background signal in the dwell time range of up to 3 ms and was overestimated at longer dwell times. The threshold of the background signal calculated by 10σ in the entire dwell time range was excessively overestimated relative to the real intensity. The selection of the threshold criteria for the discrimination of the background signals from the NP signals remains on the researcher, but it must be reasoned. Background signals should not be taken as false positive NP signals, and low-intensity NP signals should not be taken as false negatives when processing the results.size of TiO2 NPs. At dwell times exceeding 1 ms, the background signal was detected in each window . The obtained particle size distributions (hydrodynamic diameter) for samples 3, 4, and 5 were in the ranges 174–575, 174–437, 229–575 nm, respectively . Note th2 nanoparticles is absent. At the same time, the distribution of the sample signals has peaks with intensities exceeding the upper limit of the background signal intensity of 48 counts, which is clearly seen in the enlarged scale of the ordinate axis “Frequency” , and most of the registered NPs have been removed from the distribution  equal to the sample and background distributions.The number of background signals N on the sample and background spectra, the NP signal in one dwell time window on the sample spectrum conditionally replaces one dwell time window on the background signal spectrum. Respectively, the number of background signals in the signal distribution of the sample decreases by the number of NPs in this sample, which is not taken into account in Equation (1).With the initial equality of the total number of dwell time windows For reliable correction of the background signal, the number of background distribution signals corr”, which will contain only NP signals.Considering the assumptions made, subtracting the corrected distribution of background signals from the distribution of sample signals according to Equation (5) will result in obtaining the corrected distribution of sample signals “sample–bgd2 NP sizes, including those with NP sizes below the LODsize defined by the background signal, the number of background signals with certain intensities will exceed the number of NP signals. The corrected distribution of the background signal does not permit accounting for the signals of small NPs with intensities equal to the intensities of background signals at the upper limit. The calculated number of background signals corr” distribution has a number of signals in the negative range, but significantly less than before the correction   in the dwell time range of 16–20 ms, and a similar number of TiO2 NPs (808 and 774 particles)  . For samll time) . An incrrticles) .2 NPs and choose the optimal conditions for their detection.Thus, the signal distribution correction of the samples allows us to account for the contribution of the background signal to the size distribution of TiO2 NP size distribution after the correction was observed in a narrow range. For example, for sample 5 it was 206–216 nm versus 230–321 nm for distributions without background signal correction  was in the range of the background signal fluctuations.It should be noted that in each case the developed algorithm of background signal correction provided the same LODof 71 nm . The LOD2, and samples 3, 4, and 5 contained TiO2 NPs. Sample 6 did not contain TiO2 and was used as a control sample.The objects of the study were freely available cosmetic products . Accordit-octylphenoxypolyethoxyethanol, Sigma-Aldrich, St. Louis, MO, USA), 0.22-μm syringe filters  and deionized water  were used. Standard reference dispersion (SRD) of 60 nm silver NPs  was used to determine the transport efficiency of ICP-MS. A 0.1 mg/mL dissolved Ti standard solution  stabilized by high-purity nitric acid  was used for calibration.For experimental studies, a non-ionic surfactant Triton X-100 . The ICP-MS operating parameters were selected according to the manufacturer’s recommendations . The sigThe presence of NPs in the cosmetic product samples was confirmed on the IG-1000 nanoparticle size analyzers  at a voltage of 35 Vpp, a frequency of 500 kHz, and a time of 0.1 s.The samples were sonicated in a GRAD 40–35 ultrasonic bath .2 NPs, an indirect calibration procedure was used. Calibration of the SP-ICP-MS system for TiO2 NP size determination was performed according to the procedure described in [Due to the unavailability of reference monodisperse standards of TiOribed in ,23.d is the NP diameter, ρ is the NP density, d and density ρ.The transport efficiency of ICP-MS was determined using the SRD of 60 nm silver NPs with the known number of NPs in the volume of dispersion. The number of NPs was calculated based on the Ag total mass concentration of the SRD and the mass of a single 60 nm silver NP calculated by Equation (6):2 NPs was the same.The ratio between the number of NP peaks on the time-resolved spectrum and the number of NPs in the volume of standard dispersion introduced into the system allowed calculating the fraction of silver NPs that reached the detector considering the flow rate. The detection of the signal of the standard 60 nm silver NPs was performed under strictly optimized conditions, fulfilling the requirement to detect each single NP completely in a single dwell time . The tra2 NP size was performed using Ti standard solution. Solutions for constructing the calibration curve were prepared before measurement. The mass of Ti (m) that reached the detector and generated a signal of known intensity was calculated based on the concentration of the standard solution and the transport efficiency according to Equation (7):V is the sample flow rate, C is the Ti concentration in the standard solution.The instrument calibration for the determination of TiOm) was used in the determination of TiO2 NPs assuming that the behavior of the Ti ionic form of the Ti standard solution and TiO2 NPs was the same. The mass of Ti (m) corresponding to the signal intensity was recalculated into the mass of TiO2 NPs  was calculated by Equation (8) using the NP mass (3 for TiO2) and assuming the spherical geometry of the particle:The transformed calibration curve of the dependence of the signal intensity on the real detected mass of Ti (v/v) was added, and the solution was made up to the mark with deionized water. The mixture was shaken and sonicated for 30 min. Large particles and agglomerates of TiO2 NPs were filtered by a 0.22-μm syringe filter. The filtrate was diluted with deionized water and sonicated for 5 min to evenly distribute the NPs in the solution. All measurements were performed immediately after the suspension preparation to prevent agglomeration of TiO2 NPs in time.Preparation of the studied samples of cosmetic products for analysis was performed according to a well-known procedure . A 0.05-The obtained raw data were processed using Microsoft Office Excel software. The signal intensities from the time-resolved spectrum were represented by graphical distributions in «frequency–intensity» coordinates, where frequency is the number of signals of a definite intensity. The change in signal intensity in a wide range of dwell times was estimated in absolute unit–counts. The ratio of counts per second (cps) was recalculated considering dwell time. The absolute intensity value (counts) was used to represent signal distributions. Moreover, the capabilities of the SPCal software were used to calculate the background signal .2 NP sizes in cosmetic products. The analysis of TiO2 NPs in cosmetic products is a difficult task due to the high background signal and the requirements for optimal detection conditions by this method. It was found that the background signal in the analysis of TiO2 NPs in cosmetic product samples at the millisecond time resolution was determined by the background content of Ti in its dissolved form and interferents in deionized water, and the background signal of the cosmetic matrix can be regulated by the sample dilution.We considered the specifics of the millisecond time-resolved SP-ICP-MS determination of TiO2 NPs complicates the correct estimation of the NP signals. Using a σ-criteria for correcting the background signal leaves either a false positive background signal or accepts low-frequency signals of NPs as false negatives in the overlapped area. An alternative correction procedure for the frequency and intensity of the background signal was proposed, which allows the separating of the background and TiO2 NP signals in the overlapped area.The background signal was shown to make a significant contribution to the distribution of sample signals and depend on the detection conditions. Overlapping of the distributions of background signals and TiO2 NP signals in real cosmetic product samples were studied in the dwell time range of 4–20 ms. The limit of detection of the size (LODsize) of TiO2 NPs was 71 nm when the developed procedure for correcting the background signal was used. It has been shown that with SP-ICP-MS determination of the TiO2 NP sizes in real cosmetic products, the LODsize does not depend on the background signal and is determined by the ICP-mass spectrometer sensitivity.The distributions of TiO