The association between dietary saturated fatty acids (SFA) intake and type 2 diabetes (T2D) remains unclear. This study aimed at investigating the association between SFA intake and T2D risk based on (1) individual SFA (differing in carbon chain length), (2) food sources of SFA and (3) the substituting macronutrients.37,421 participants from the European Prospective Investigation into Cancer and Nutrition-Netherlands (EPIC-NL) cohort were included in this study. Baseline dietary intake was assessed by a validated food frequency questionnaire. T2D risks were estimated by Cox regression models adjusted for non-dietary and dietary covariates.893 incident T2D cases were documented during 10.1-year follow-up. We observed no association between total SFA and T2D risk. Marginally inverse associations were found for lauric acid , myristic acid , margaric acid , odd-chain SFA , and cheese derived SFA . Soft and liquid fats derived SFA was found related to higher T2D risk . When substituting SFA by proteins, carbohydrates and polyunsaturated fatty acids, significantly higher risks of T2D were observed (HRs per 1 energy% ranging from 1.05 to 1.15).trans fatty acids or limited intake variation in polyunsaturated fatty acids and vegetable protein.In this Dutch population, total SFA does not relate to T2D risk. Rather, the association may depend on the types and food sources of SFA. Cheese-derived SFA and individual SFA that are commonly found in cheese, were significantly related to lower T2D risks. We cannot exclude the higher T2D risks found for soft and liquid fats derived SFA and for substituting SFA with other macronutrients are influenced by residual confounding by The online version of this article (10.1007/s00394-018-1630-4) contains supplementary material, which is available to authorized users. Dietary guidelines recommend that intake of total saturated fatty acids (SFA) should not exceed 10% of energy per day . DietaryFirst, individual SFA that differ in carbon chain length may have different associations with T2D risk. So far, only three studies have addressed the associations between intakes of individual SFA through self-reported dietary measurements and T2D , 9, 10. Second, the association between SFA and T2D risk can be modulated by its food sources. As far as we know, no study has investigated SFA derived from different food sources in relation to T2D risk. However, a recent cross-sectional study found that SFA from meat was positively associated with more insulin resistance and less insulin secretion, whereas SFA from dairy and plant sources were beneficially related to these markers . In addiThird, the association may also depend on macronutrients that substitute SFA in the diet. The Nurses’ Health Study and Iowa’s Women Study found that replacing energy from SFA with polyunsaturated fatty acids (PUFA) was related to lower T2D risk , 15. HowThe aim of this study was to investigate the associations between intake of SFA and risk of incident T2D in a Dutch population based on (1) individual SFA (differing in carbon chain length), (2) the food sources of SFA and (3) the macronutrients that substitute SFA.The European Prospective Investigation into Cancer and Nutrition-Netherlands (EPIC-NL) cohort consists of the MORGEN (Monitoring Project on Risk Factors for Chronic Diseases) and the Prospect cohorts. The design and rationale of EPIC-NL were described in detail elsewhere . Brieflyn = 1046); prevalent diabetes cases at baseline (n = 679); participants with missing information of dietary intake (n = 180) and other co-variables, such as education information and physical activity level, (n = 339); participants with implausible energy intake based on the ratio of reported energy intake to estimated basal metabolic rate, i.e., the top or bottom 0.5% of the ratio (n = 346); leaving a total of 37,421 men and women for analyses.For this study, participants were excluded when one of the following criteria was fulfilled: participants who withheld permission for linkage with disease registries (Food intake was assessed by a self-administered Food Frequency Questionnaire (FFQ) on the usual consumption frequency of 79 main food categories during the year preceding enrolment. The FFQ was described in detail elsewhere . The FFQThe intakes for all micronutrients and macronutrients except individual SFA were calculated based on the Dutch food composition table 1996 . The intIntake of total SFA was calculated as the sum of SFA derived from the predefined food groups mentioned above. Considering relative low intakes of single individual SFA, two additional groups were made to add efficiency of the study: short- and medium-chain SFA  and odd-chain SFA (pentadecylic plus margaric acids).Energy reporting status of participants was estimated by comparison between self-reported energy intake and BMR . AccordAlcohol consumption was categorized as follows: 0, 0.1–6.0, 6.1–12.0, 12.1–24.0, and > 24.0 g/day for women and 0, 0.1–6.0, 6.1–12.0, 12.1–24.0, 24.1–60.0, and > 60.0 g/day for men. Glycaemic index (GI) of foods, a measure of the extent to which foods raise the blood glucose level, was obtained from international table complied by Foster-Powell et al. . To adjuThree sources of ascertaining incidence T2D were included in this study: self-report, hospital discharge diagnoses and urinary glucose strip tests . Verification of potential diabetes cases detected by any of these methods was carried out with information from participants’ general practitioner or pharmacist. Verification information was available for 89% of the potential diabetes cases, and 72% of these cases were verified as T2D and subsequently used for the analysis. The process of ascertainment and verification of incident and prevalent diabetes cases is described in detail elsewhere .At baseline, information on demographic characteristics, lifestyle characteristics, the presence of chronic diseases, and risk factors of chronic diseases was obtained with general questionnaires. Smoking status was categorized as current, past or never smoked. Family history of diabetes was defined as at least one parent with diabetes. Education was classified as low, middle and high education. Physical activity was measured with a validated questionnaire, from which the Cambridge Physical Activity Index was calculated and categorized as inactive, moderately inactive, moderately active and active .2). Systolic and diastolic blood pressures were obtained as the mean of two measurements in the supine position on the left arm using a random zero sphygmomanometer (MORGEN), or on the right arm using a BOSO Oscillomat  (Prospect). Hypertension was defined when one or more of the following criteria were fulfilled: diastolic blood pressure > 90 mmHg, systolic blood pressure > 140 mmHg, self-reported presence of hypertension or use of hypertensive medication. Hyperlipidaemia was defined through self-reported diagnosis and/or use of medication.A baseline physical examination was performed according to standard operating procedures. Body weight, height, waist circumference and blood pressure were measured. BMI was calculated as weight divided by height squared  and median (interquartile range) for continuous variables, or percentage for categorical variables. Correlations between dietary intakes of total SFA, individual SFA and SFA from different food sources, as well as correlations between intakes of SFA from different food sources, total fatty acids from that food source and the food source itself, were calculated using Pearson correlations coefficients.Person-years were calculated as the time between the date of the study entry and the dates of diabetes diagnosis, death, loss to follow-up or end of follow-up (1 January 2006), whichever came first. The endpoint of analysis was incident T2D.trans fatty acids, protein , dietary cholesterol, vitamin E and fibre. Model 5 was additionally adjusted for BMI (continuous) and waist circumference (continuous). Similar models were used for intake of SFA in grams per day. Intakes of individual SFA or SFA from specific food sources were separately evaluated per 1 SD of intake and entered into Cox models as continuous variables. We used the same adjustment models as described above, except that we additionally adjusted for the sum of all other consumed SFA.Cox proportional hazard regression models were used to calculate HRs with 95% CIs for the association between SFA intake and risk of T2D. Total SFA intake was evaluated per 1% of energy and entered as a continuous variable into Cox regression models. In addition to a crude model which was adjusted for total energy intake (Model 1), four models were constructed to adjust for potential confounding. Model 2 was adjusted for sex (men or women) and age at baseline (continuous). Model 3 was additionally adjusted for lifestyle and demographic factors: alcohol consumption level (six categories), smoking (three categories), physical activity (four categories), and education level (three categories). Model 4 was additionally adjusted for dietary factors (continuous): energy-adjusted intake of trans fatty acids, total carbohydrates, animal protein, and vegetable protein) [Cox models were converted into substitution models to estimate the risk of T2D when energy intake from total SFA was substituted with an equal amount of energy from each of the other macronutrients , protein) . We choo2), family history of T2D (without family history and with family history), energy reporting status  and sex (men and women) were estimated using a likelihood ratio test and by including continuous interaction terms in the multivariable adjusted model (model 5). The analysis was repeated in the same population after excluding the first 2 years of follow-up to minimize the possibility of reverse causation. The analysis was also performed in the population after excluding prevalent cases of cancer or cardiovascular disease or hypertension or hyperlipidaemia and participants who had a stroke or myocardial infarction, due to their possible dietary changes caused by the illness. Furthermore, in additional analyses, we estimated the associations between non-SFA fatty acids derived from different food sources (per 1 SD of intake) and T2D risk, to see whether we can actually tease SFA apart from other fatty acids derived from the same food source in relation with T2D.The proportional hazards assumption was tested by calculating Schoenfeld residuals and visual inspection of log–log plots, no deviation was detected. Interactions of total SFA intake (en%) with BMI categories , and P-values < 0.05 (2-sided) was considered as statistically significant.trans fatty acids, animal protein, cholesterol, vitamin E, vitamin D and calcium, and lower intakes of vegetable protein, carbohydrates, fibre and alcohol. Dietary GI and intake of PUFA were similar across SFA intake levels.In this study population , participants with a relatively high SFA intake were on average older women with a lower education level and were more likely to be a smoker and less devoted to physical activity. They also tended to have a higher BMI and waist circumference, a higher blood pressure, but were less likely to have hyperlipidaemia Table . Furtherr = 0.92), myristic and pentadecylic acids (r = 0.92), myristic and margaric acids (r = 0.85), pentadecylic and margaric acids (r = 0.88) . SFA from specific food source was highly correlated with total fatty acids from that food source, as well as the intake of the food source itself .The mean ± SD baseline intake of total SFA in the study population was 14.0 ± 2.6 percentage of energy. More than 95% of the population surpassed the upper intake limit of 10% of energy per day recommended by the Health Council of the Netherlands. The long-chain SFA palmitic acid (51.1%) and stearic acid (25.2%) represented most of the SFA intake . Cheese (17.0%), meat (16.9%), milk and milk products (15.8%) were the main food sources of SFA . High Pearson’s correlations were observed between palmitic and stearic acids . This result was similar when intake of SFA was modelled in grams per day  , no significant association between intake of energy from total SFA with risk of T2D was observed , myristic acid , margaric acid  and odd-chain SFA . No significant associations were observed for intakes of short- and medium-chain SFA (butyric through capric), palmitic, pentadecylic and stearic acids.Table per SD increase: 0.90; 95%I 0.83, 0.98) after multivariable adjustment (model 5), but not for milk and milk products derived SFA . Higher intake of soft and liquid fats derived SFA was associated with a higher risk of T2D . No significant associations were observed for intake of SFA from other food sources  or carbohydrates  was associated with a significantly higher risk of developing T2D. Surprisingly, we also observed an increased T2D risk when substituting SFA with vegetable protein  or PUFA . Substituting SFA with carbohydrates differing in GI values resulted in differences in T2D risk: substitution with high GI carbohydrates associated with a higher risk of T2D , whereas substitution with low- or medium GI carbohydrates did not relate to T2D risk. No significant association was observed with substitution of SFA with MUFA.Table P values: 0.13–0.99) and energy reporting status  was found. Moreover, no meaningful effect modifications by baseline BMI and sex were observed for the relations of SFA and T2D , but interactions existed between sex and substituting SFA with animal protein and PUFA . Stratification for sex in substitution models indicated a significant higher risk of T2D in women when substituting SFA with animal protein or PUFA, whereas this association was not present in men . Our results did not materially change after excluding the first 2 years of follow-up , or restricting analyses by including only baseline healthy participants . Associations between non-SFA fatty acids from specific food sources and T2D risk were fairly similar to associations between SFA from that food sources and T2D risk .No significant effect modification of family history , it is difficult to conclude whether the observed inverse association of cheese derived SFA is due to (particular) SFA, cheese as a whole product, or other constituents present in cheese such as calcium [per SD increase: 1.10; 95% CI 1.02, 1.19) is also in line with existing evidence [per SD increase: 1.03; 95% CI 0.95, 1.11).The associations we observed for individual SFA correspond to our findings for SFA derived from different food sources. To our knowledge, this is the first observational study that investigated SFA from different food sources in relation to incident T2D. We observed that dietary SFA derived from cheese was inversely related to T2D risk. This finding is supported by previous studies that found intake of cheese to be associated with lower T2D risk , 32, alt calcium . Moreove calcium . Additio calcium . Additio calcium , 35. Ourevidence , 36. Whetrans fatty acids at the beginning of the study. During the mid-1990s, there was a notable reduction of trans fatty acids content in margarines. This may have led to residual confounding by underestimated trans fatty acids intake, consequently affecting the observed association of substituting SFA with PUFA. This may also explain why we found higher T2D risk with SFA from soft and liquid fats. Furthermore, compared to the Nurses’ Health Study , the intake range of PUFA is relatively small in our study . As we only found adverse association for substituting SFA with animal protein or PUFA in women in the gender-stratified analysis, the result could be largely driven by the large proportion of women in our study. Altogether, the increased T2D risks when substituting SFA by proteins, carbohydrates, and especially PUFA, needs to be interpreted with caution.As far as we know, no previous studies have investigated substitutions of SFA with protein, carbohydrate nor MUFA in relation to T2D risk. We found substituting SFA with carbohydrate, animal protein, vegetable protein or PUFA was associated with higher T2D risk, but not with MUFA. Previous studies suggest that animal protein is related to a higher T2D risk –39, whictrans fatty acid intake. Further investigation is still warranted, especially in populations with other dietary patterns contributing to SFA intake, and with more variation in intake of PUFA, to draw further conclusions of the relations of SFA with T2D risk.In conclusion, our findings emphasize the importance of distinguishing between the types and food sources of SFA in relation to T2D risk. Our study suggests that individual SFA that exist to a large extent in cheese, and SFA derived from cheese, relate to a lower T2D risk. Regarding substitution of SFA with other macronutrients, we cannot exclude the possibility that our results reflect inverse associations of dairy-derived fatty acids with T2D and/or residual confounding by Below is the link to the electronic supplementary material.Supplementary material 1 (DOCX 255 KB)


There is still a good reversibility with cycling efficiency of 88.8 % after 2,000 cycles, demonstrating its excellent long-term cycling stability and might be the promising candidates as the excellent electrode material.In this paper, for the first time, we report the synthesis of nanoscale cuprous chloride (CuCl) cubic structure by a facile hydrothermal route. A possible mechanism for the growth of those nanostructures is proposed based on the experimental results. It is discovered that the existence of HCl could affect the surface of CuCl nanocubes. This unique cube-like nanostructure with rough surface significantly enhances the electroactive surface areas of CuCl, leading to a high special capacitance of 376 mF cm The increasing demand for energy in the 21st century has triggered tremendous research efforts for energy storage and conversion from clean and renewable energy sources . The sup2 nanostructures assembled by many ultrathin nanosheets. They thought that their excellent supercapacitor performances could be ascribed to their unique morphology and the fast ion and electron transfer characteristics [3 nanowires and investigated their electrochemical performances [In the last decade, supercapacitor technology has undergone an increasing development owing to the discovery of new electrode materials, especially metal oxide nanomaterials, and the design of new hierarchical nanostructures –23. For eristics . Gu et aormances . Mai’s gormances . HoweverCuCl is an I-VII semiconductor with a direct band gap of ~3.4 eV. It has a large exciton binding energy (~190 meV), which suggests the possibility of the fabrication of exciton-based blue/UV optoelectronic devices . CuCl is2 pieces and immersed into acetone solution, ultrasonically cleaned for 30 min, and rinsed with deionized water. 5.5 mmol CuCl2 powder was dissolved into 20 mL deionized water with constant stirring. Another 20 mL deionized water was added into 8 ml concentrated HCl (its density is 1.18 g cm−3) in a 50 mL glass beaker. The mixed solution was then put into a 100 ml sealed Teflon-lined autoclave, followed by hydrothermal reaction at 140 °C for 12 h. Afterward, the autoclave was naturally cooled to room temperature. The substrate was taken out, rinsed with copious deionized water, and dried at 100 °C for 12 h in air.All the chemicals were of analytic grade and used as received without further purification. In a typical procedure, Cu substrate was cut into 1 × 1 cmλ = 0.1542 nm, 40 kV, 150 mA).The detailed morphologies of the samples were characterized by scanning electron microscope . The chemical and elemental compositions of the prepared products were verified by energy dispersive spectroscopy (EDS), attached with SEM. The crystallinity of the prepared nanocubes was examined by X-ray diffractometer , of the electrode material was calculated from the galvanostatic discharge according to the following equation:I is the discharge current (A), Δt is the discharge time (s), ΔV is the voltage change (V) excluding IR drop in the discharge process, and S is the geometrical area of the electrode. The electrochemical impedance spectroscopy (EIS) measurements were performed by applying an AC voltage with 5 mV amplitude in a frequency range from 0.01 Hz to 100 kHz.Electrochemical characteristics of the as-obtained products were studied on an CHI660 electrochemical work station  using cyclic voltammetry (CV) and electrochemical impedance test by configuring the samples into a three-electrode cell, where the substrate was used as the working electrode, Pt foil as the counter electrode, and an Ag/AgCl electrode as the reference electrode. The electrolyte used was 1 M NaThe general morphologies of the prepared CuCl products were investigated by SEM, and the results are demonstrated in Fig. To further investigate the growth mechanism, controlled experiments were conducted by varying the reaction time. Figure 2O and Cl− provided by HCl, and copper(I) chloride complexes have been carried out according to the reaction , which includes the inherent resistances of the electroactive material, the bulk resistance of electrolyte, and the contact resistance at the interface between the electrolyte and electrode. Its range also corresponds to the charge-transfer resistance caused by the Faradic reaction, which was correlated with the intercalation and deintercalation of ion. The charge transfer resistance (Rct), which results from diffusion of electrons, can be calculated from the diameter of the semicircle in the high-frequency range. The Warburg resistance (Rw), which describes the diffusion of redox species in the electrolyte, can be reflected from the slope of the EIS curve in the low-frequency range [Qc represents the constant phase element accounting for a double-layer capacitance [Rs) manifests the good conductivity and the very low internal resistance of CuCl electrode, and consistent interfacial contact between CuCl and the Cu substrates. The enhanced electrochemical performance could be ascribed to the following structural features. First, separate cube-like structure leads to large open spaces to facilitate the electrolyte penetration and shorten the diffusion paths for both electrons and ions, resulting in reduced internal resistance. Second, Cu substrate can provide fast electronic transfer channels to improve the electrochemical performance. Moreover, Cl− from electrolyte can successively induce CuCl and then form combination of CuCl–Cl together with the occurrence of electrons migration through the bond to access the underlying copper substrate [−2 at current densities of 1.0, 1.5, 2.0, 3.0, and 5.0 mA cm−2, respectively. The enhanced electrochemical performance could be attributed to the following structural features. First, CuCl can absorb electrolyte anions (Cl+) on the electrode surface from electrolyte: (CuCl) surface + Cl− + e− ←→ (CuCl–Cl−) surface [− from the electrolyte is fully utilized in CuCl electrode. Figure −2. An areal capacitance of 376 mF cm−2 and a good reversibility with cycling efficiency of 88.8 % after 2,000 cycles are shown in Fig. −2, the specific capacitance is 27.6 mF cm−2, maintaining 10 % of that at the current density of 1 mA cm−2.To highlight the merits of the obtained unique CuCl architectures and further explore their potential applications in the supercapacitors, the electrochemical performances of CuCl nanocubes as the integrated electrode were evaluated in three-electrode configuration with 1 M Nacy range . Its ranacitance . The intubstrate . The cal surface , providi−2 at the current density of 1 mA cm−2 and a good reversibility with cycling efficiency of 88.8 % after 2,000 cycles. The synthesized CuCl nanocubes may have potential applications in energy storage and other electrochemical devices.In summary, a mild hydrothermal method was used to fabricate CuCl nanocubes. The as-prepared product possesses a specific capacitance of 376 mF cm


The electrospinning process has gained popularity due to its ease of use, simplicity and diverse applications. The properties of electrospun fibers can be controlled by modifying either process variables  or polymeric solution properties . However, many variables affecting electrospinning are interdependent. An optimized electrospinning process is one in which these parameters remain constant and continuously produce nanofibers consistent in physicochemical properties. In addition, nozzle configurations, such as single nozzle, coaxial, multi-jet electrospinning, have an impact on the fiber characteristics. The polymeric solution could be aqueous, a polymeric melt or an emulsion, which in turn leads to different types of nanofiber formation. Nanofiber properties can also be modified by polarity inversion and by varying the collector design. The active moiety is incorporated into polymeric fibers by blending, surface modification or emulsion formation. The nanofibers can be further modified to deliver multiple drugs, and multilayer polymer coating allows sustained release of the incorporated active moiety. Electrospun nanofibers prepared from polymers are used to deliver antibiotic and anticancer agents, DNA, RNA, proteins and growth factors. This review provides a compilation of studies involving the use of electrospun fibers in biomedical applications with emphasis on nanoparticle-impregnated nanofibers. Electrospinning is a process of forming micro/nanometer-sized polymeric fibers, either hollow or solid, with the application of the electric force on the polymeric solution at the tip of a conducting tube. It is one of the most commonly used techniques to obtain continuous fibers in the nanometer size range ,2,3. EleThe electrospinning process involves the use of a very high voltage source (of either positive or negative polarity) to charge the polymer solution or melt, a grounded collector, and a syringe pump . It is acV  is reached. At cV, the jets (electrospinning) and sprays (electrospraying) begin, which are represented by Equation (1):H is the separation between capillary and the collector, L is the length of capillary, R is the radius of capillary and γ is the surface tension [V is the required voltage, γ is the surface tension and r is the radius of the pendant drop. Even though viscosity and conductivity are vital in the electrospinning process, they are missing from the equation. The use of applied voltage and the surface tension gives a representative equation for slightly conducting, medium- to low-conductivity solutions [When the electric field/voltage is increased gradually and the surface of the drop becomes convex at a certain voltage,  32) 0.11πγRwhere tension ,14:(2)V=The electrospinning process is simple and does not involve heavy machinery. The processing parameters and solution parameters affect the size, porosity and uniformity of the fibers. These parameters have been discussed individually; however, the observations made in any particular study by a team of researchers are not universal. The modification of a parameter in one polymer may produce a totally different result with another polymer. In addition, none of the parameters act independently during the electrospinning process, and the final fibers are the result of a combination of several parameters.The primary factor influencing the formation of fibers is the strength of the applied DC voltage. The size of the fiber, formation of beads and absence of jet formation are all dependent on the applied DC voltage. With an increase in the applied voltage of a polyethylene oxide (PEO)/water system, the originating site for the jet changes from the tip of the pendant drop to the tip of the capillary, and the volume of the pendant drop decreases gradually . When thThe fibers formed during the electrospinning process transport the charge to the grounded collector plate to close the circuit. This allows the electric current associated with the process to be measured; it is small . When conductivity, dielectric constant and flow rate through the pump remain the same, an increase in current indicates an increase in the mass of the fibers formed. During the electrospinning process, the increase in current is gradual at first and then increases sharply from one voltage point, while sharp, steep increases are observed during electrospraying. The point of a sharp change in current could be indicative of the defect/change in the bead density [Studies on the reversal of polarity obtained by grounding the solution in capillary and charging the collects resulted in inconsistent findings. Kilic et al. studied the effect of polarity on the electrospinning process of 7.5 wt% poly/water solution on production efficiency and nanofiber morphology. They concluded that due to the lack of columbic force acting on the polymer jet, the new reversed setup resulted in less nanofiber production. They also reported that the diameter and pore size of the web layer were much finer and more homogeneously distributed in the conventional setup . On the The polymer flow rate has a direct impact on the size, shape and porosity of electrospun fibers. A study on polystyrene/tetrahydrofuran (THF) solutions by Megelski et al. reported an increased fiber diameter and pore size with flow rate. However, at high flow rates, bead defects and flat, ribbon-like structures were observed due to insufficient drying . In a diThe distance between the capillary and the collector is another factor which plays a significant role in controlling the size and morphology of the nanofibers . This diw/v solutions of poly(desaminotyrosyl-tyrosine ethyl ester carbonate) (poly(DTE carbonate)) were electrospun at a voltage of 10 kV to 25 kV at 10 cm distance. Beaded fibers were observed during electrospinning at a lower concentration solution until the voltage reached 20 kV, and the density of beads decreased until the voltage reached 15 kV; average fiber diameter increased, while increasing the voltage from 20 kV to 25 kV. At higher solution concentrations, the smooth fibers obtained showed an increasing diameter and decreasing fiber density with the increase in the electric field from 10 kV to 25 kV [Viscosity and surface tension, which determine the spinnability of a solution, should be taken into consideration in determining the concentration of solution/melt for electrospinning. Surface tension is a dominating factor in a low concentration solution , and at such concentration, drops will be formed instead of a continuous fiber. At a higher concentration (viscosity >20 poise), the flow of the solution cannot be controlled and maintained. In the PEO/water study discussed above in to 25 kV .6 g/mol), exhibited a variety of fiber morphologies even with a minute change in the concentration within the range 0.3–3.0 wt%. Beaded and smooth fibers were formed at concentrations between 0.3 to 0.7 wt%, whereas smooth fibers with ribbons coexisted at 0.7 to 2 wt%. Above 2.0 wt%, only ribbons were formed, which were either helical or zigzag with triangular beads on them [The molecular weight of polymer influences the solution viscosity and thus has an important effect on fiber morphology. For example, decreasing the molecular weight of poly while maintaining other parameters constant resulted in the formation of bead-like structures. On the other hand, higher molecular weight resulted in smooth fibers initially followed by ribbon-like fibers upon further increase in molecular weight . Ultra-h on them . Additio on them .Viscosity determines the solution’s ability to form fibers. Smooth continuous fibers can be obtained at optimum viscosity for a particular polymer solvent combination. The viscosity, molecular weight of the polymer and polymer concentration are interrelated, and one cannot be independently judged. Low viscosity fails to form the fibers, while high viscosity requires a higher electric field for electrospinning, making it hard to operate ,32. BaumSurface tension is the measure of cohesive forces between the molecules in solution form and is dependent upon the solution composition, polymer and solvent(s) used. Yang et al. studied the influence of solvents on the formation of nanofibers with poly(vinyl pyrrolidone) and concluded that lower surface tension with high viscosity formed smooth nanofibers with ethanol as the solvent. They proposed the use of a multi-solvent system to obtain optimum surface tension and viscosity parameters for better electrospun fibers . Like viThe impact of conductivity and surface charge density of solution also plays an important role in the process of electrospinning. It is important to have high conductivity for a greater charge-carrying capacity. A highly conductive solution experiences a stronger tensile force in an electric field compared to the less conductive solution, making the former preferable for electrospinning. An increase in the conductivity of the solution causes a substantial decrease in the diameter of the nanofibers. The radius of the fiber jet varies inversely as the cube root of the conductivity of solution . For preThe distance between the tip of the capillary and the collector is only a few centimeters , and the path taken by the jet to reach the collector is a few folds more. The fibers formed during the process are porous and dry fast, depending on the choice of the solvent used for solubilization of the polymer. The insufficiently dried fiber may attach to itself mid-air , form riThe type of electrospinning process can have a significant impact on fiber formation in addition to the process and solution parameters. Two main aspects to deal with in the type of electrospinning are solution vs. melt electrospinning and nozzle configuration .Electrospun fibers are generally obtained from polymer solutions or melts or emulsions. Melt spinning has high-throughput rate and process safety; solutions have the advantage of using a large variety of polymeric materials, lower energy consumption and superior mechanical, optical and electrical properties of prepared fibers. Emulsion electrospinning is needed for high melting polymers to prepare flame-retardant fibers. A comparison of different spinning methods using poly(lactic acid) (PLA) as a model polymer has been reported by Gupta et al. .The melt of the polymer with other additives is extruded through the capillary, resulting in a thin fiber that cools and solidifies rapidly during its time in the air before depositing onto the collector . Take-upThe solution spinning method is ideal for polymers or blends of polymers that are thermally unstable or degrade upon melting. Based on how the solvent used for preparing the polymer solution is removed from the fibers, this method can be divided into two different types, namely, dry spinning and wet spinning. The dry spinning process uses hot air or inert gas on the polymer jet to facilitate the solvent’s evaporation and fiber solidification. For example, Postema et al. used a solvent combination of chloroform and toluene to produce PLA fibers of high strength by using dry spinning and hot-drawing processes. They were able to obtain an optimal tensile strength of 2.2 GPa by electrospinning at 25 °C . The wetIn the emulsion electrospinning process, finely ground polymers, insoluble and non-melting, are mixed with another polymer solution with a catalyst and emulsifying agents. The formed emulsion is then electrospun by either the dry or the wet spinning method. This technique has been used in preparing fibers from fluorocarbons with high melting points, ceramics, and polymer blends with flame-retardant properties . PersanoNozzle configuration relates to the number and the arrangement of capillary tubes from which the jets of fibers emerge. A single nozzle configuration is the simplest and most common configuration, wherein the charged solution flows through a single capillary . This paf~Qc0.18 where df represents the inner diameter of the fiber and Qc represents the flow rate of core solution [A second type of nozzle configuration that has recently been introduced is coaxial configuration. Two separate polymer solutions flow through two different capillaries, where the small capillary is within the larger capillary. This configuration can easily encapsulate a small fiber within a larger fiber, forming core-shell morphology . In one solution  This metsolution ,62,63. Csolution , flurbipsolution , and ibusolution  and antisolution  were preThe multi-jet electrospinning based on multiple capillaries arranged in circular geometry is yet another modification of the nozzle that can both increase the throughput and facilitate scale up and commercialization. This modification increases the mat thickness, deposits in a larger area and can mix fibers of different materials for better strength and versatility of use ,67. In tThe collector plate can be configured based on the application of polymeric fibers. Commercially, the most common collectors are stationary plates  and rotating plates. Both these types of collectors can be subdivided further into continual or patterned. A continual collector is simple and can be used to obtain fibers with a random internal structure on a stationary type, while some degree of directional control can be gained with the higher speed of a rotating collector. The patterned collectors have thin conductive wires separated from one another by an air gap. During the process, the fibers are deposited either between or perpendicular to the wires, individually, achieving some degree of alignment. Properly aligned polymer fibers are mostly useful in tissue engineering. In the rotating type of patterned collector, operation at lower speeds deposits fibers between the conductive wires, while deposits at the higher speed are dependent on electrostatic and mechanical forces increasing the degree of alignment. For research purposes, different designs of collectors have been used, including: mesh , pin 7373, gridsElectrospinning is easy and cost effective, and it offers great flexibility in selecting materials, high loading capacity and high encapsulation efficiency, which makes it suitable for medical and drug-related research. There are various methods of drug loading in the polymeric solution for electrospinning.Blending is the primary method for incorporating drugs into the polymer solution by dissolving or dispersing the drug and then subsequently electrospinning. Though this method is simple and easy, the physicochemical properties of the drug and the polymer need to be precisely considered, as these affect the encapsulation efficiency, the drug distribution in the fiber and the kinetics of the drug release. Lipophilic drugs  should be dissolved in a lipophilic polymer and hydrophilic drugs  in a hydrophilic polymer for better encapsulation. When the drug is not dissolved properly in the polymer solution, a dispersion is obtained, which might lead to burst release if the drug migrates to the fiber surface . To obtaSurface modification is the technique in which the therapeutic agent is bound or conjugated to the fiber surface to make it structurally and biochemically similar to the tissue. The drug release in this case will be attenuated, and the functionality of the biomolecules will be protected . The burAnother approach is the process of forming an emulsion for electrospinning, where the drug or the protein solution is emulsified within a polymer solution. The latter acts as an oil phase, and spinning such an emulsion produces a well-distributed fiber for a low molecular weight drug  and a coMulti-drug delivery is a recent approach in which multiple drugs with or without similar therapeutic effects are combined and electrospun with suitable polymer(s) ,101. WanYet another innovative method of incorporation and delivery of drug combines the large surface area of electrospun fibers with polyelectrolyte multilayer structures . Such a The electrospinning process has been used in drug delivery for treating various diseases. Electrospun fibers are administered mainly via oral and topical routes and as implantable systems.trans retinoic acid) and vitamin E (α-tocopherol) in cellulose acetate polymer-based electrospun fibers and solvent cast films [® L and Eudragit® S electrospun nanofibers for treating oral mucositis. The rapid evaporation of solvent during electrospinning prevented the drug molecules from forming crystalline aggregates within the nanofibers. In addition, the amount of ketoprofen released from nanofibers was significantly higher than that released from corresponding solvent-casted films. Furthermore, a marked reduction in inflammatory infiltrate was seen in mucositis-induced rabbits with developed nanofibers [Centella asiatica was incorporated into the electrospun fibers based on cellulose acetate polymer [The transdermal drug delivery system (TDDS) delivers the drugs locally or systemically via the skin. This is mainly suitable for drugs that cannot be taken by the oral route, either because of extensive degradation in the GIT or because the drug undergoes extensive first pass metabolism . Electrost films . The resst films . They obst films . The fibst films . This stnofibers . Apart f polymer . Suwanto polymer .Staphylococcus aureus MRSA252 and also inhibited the formation of new biofilms [Escherichia coli in apple cider.In recent years, antibiotics and antibacterial agents have been the most common drug molecules that are encapsulated, using different polymers and their combinations as carriers. Different polymers, such as PLA, PLGA and PCL, are primarily used in the polymeric electrospun fibers for biodegradability, and other natural and synesthetic hydrophilic or hydrophobic polymers are used to control the release pattern of the drug. Kenaway et al. used tetracycline hydrochloride with poly(ethylene-co-vinyl acetate) (PEVA), PLA and their 50:50 blend to deliver the drug for treating periodontal disease . They rebiofilms . Wang etbiofilms . Direct Bombyx mori silk fibroin with poly(ε-caprolactone) (PCL) electrospun fibers [Wound dressings protect the wound from external microorganisms and absorb/adsorb the exudate from the wound, providing an acceptable cosmetic appearance. The use of different components in the wound dressing prevents infection of the wound and accelerates the healing process . The comn fibers . The PCLw/o) emulsion, with water soluble drugs in aqueous phase and polymeric solutions of PEG-PLA in chloroform as oily phase, was prepared and electrospun to obtain fibers [Anticancer agents, such as doxorubicin, paclitaxel, cisplatin and dichloroacetate, have been incorporated into electrospun fibers with polymers such as PLA, PLGA and PLLA for postoperative chemotherapy. A water-in-oil  was encapsulated with BSA as a carrier protein into the nanofibers of PCL and poly(ethyl ethylene phosphate) (PCLEEP). The protein released in a sustained manner for more than three months from the electrospun fibers and showed partial retention of bioactivity . The samElectrospun nanofibers containing nanoparticles is another area of research that has been widely investigated for various applications, including surface-enhanced Raman scattering , antimicP. aeruginosa and methicillin-resistant S. aureus when compared to the pure CTS nanofibers [3 was added to form the pre-electrospinning solution. The solution was then treated using helium atmospheric plasma to reduce AgNO3 into metallic silver nanoparticles. Finally, the solution obtained was electrospun to form nanofibers containing embedded Ag nanoparticles. SEM images revealed the smooth and continuous nature of the nanofibers. The nanofibers demonstrated a sustained release of silver ions and also showed higher antibacterial activity against Gram positive (B. cereus) and Gram negative bacteria (E. coli) when compared to the untreated nanofibers [A study conducted by Lee et al. investigated the efficacy of chitosan (CTS) nanofibers containing silver nanoparticles for topical wound care. Silver nanoparticles were generated directly in the chitosan solution by chemical reduction using sodium borohydride. The nanoparticle solution obtained was then poured into sodium hydroxide resulting in the formation of CTS/AgNPs composite, which was further subjected to electrospinning to form nanofibers. The prepared fibers showed excellent antibacterial activity against nofibers . Similarnofibers . Anothernofibers .Nie and Wang studied the complex of DNA and poly lactide-co-glycolide (PLGA)/hydroxyapatite (HAp) composite scaffolds fabricated by electrospinning for their use in bone tissue engineering. DNA was incorporated into the scaffolds in the form of naked DNA or DNA/chitosan nanoparticles before/after fiber fabrication. The results revealed that the scaffolds were non-woven and predominantly composed of PLGA with a dispersion of HAp nanoparticles. The addition of HAp nanoparticles showed an increase in the release rate of DNA from the scaffolds containing both naked and encapsulated DNA. Moreover, the scaffolds with encapsulated DNA/chitosan nanoparticles also showed a higher cell attachment, greater cell viability and desired transfection efficiency in human marrow stem cells (hMSCs) . A simill-α-phosphatidylcholine encapsulating horseradish peroxidase (HRP) by the extrusion technique. This study compared the activity of HRP in nanofibers developed by coaxial or blend electrospinning with and without liposomes. Blend nanofibers were prepared by electrospinning unilamellar liposomes dispersed in an aqueous solution of PVA and nanofibers, while the coaxial nanofibers were developed using PVA-core/PCL-shell with embedded liposomes. The results indicate that the blending of liposomes could not conserve the intact liposomes, while the nanofibers obtained by coaxial electrospinning retained liposomes. In addition, the core/shell nanofibers not only preserved the activity of encapsulated HRP enzyme but also enhanced the proliferation of mesenchymal stem cells (MSC) [Liposome-enriched nanofibers were investigated for delivering and preserving horseradish peroxidase (HRP) enzymatic activity. Liposomes were prepared using soybean derived ls (MSC) .l-lactic acid-co-ε-caprolactone) (PLACL) nanofibers containing magnesium oxide (MgO) nanoparticles in synergy with aloe vera (AV), curcumin (CUR) and β-cyclodextrin (β-CD) were prepared and evaluated for treating breast cancer. In vitro toxicity of electrospun nanofibers composed of PLACL; PLACL with AV; PLACL with AV and MgO; PLACL with AV, MgO and CUR; and PLACL with AV, MgO, and β-CD was assessed in MCF-7 cells using the MTT assay. The results showed that PLACL nanofibers containing CUR showed the greatest cell death among all other nanofibrous scaffolds. However, PLACL nanofibers without MgO nanoparticles exhibited greatest tensile strength [Poly (strength . Sperm-sstrength .Electrospun nanofibers composed of poly (ɛ-caprolactone) (PCL) and zero valent zinc nanoparticles were investigated by Sezer et al. to promote neuroglial cell proliferation. Chemical characterization studies indicated that the nanoparticles did not interact chemically with the PCL matrix. In addition, the nanofibers enhanced the tensile strength of the PCL matrix and promoted cell proliferation depending on the amount of zinc nanoparticles embedded inside the PCL matrix . AnotherAlthough the benefits of electrospinning have been largely demonstrated in various fields of science, there is still a great need to implement the production in an efficient way. Several challenges relating to the electrospinning process are yet to be addressed. These include: (a) large-scale manufacturing; (b) accuracy and reproducibility during all the fabrication steps; and, (c) safety and environmental aspects of electrospinning . The majClogging of the spinneret due to the solution gelation is highly disruptive and causes production losses. This problem is more apparent with solutions of higher concentration, as they are more viscous  and withAnother important concern is regarding the solvent used in the electrospinning process. This particular issue is highly important not only for safety reasons during fabrication, but also for the final products, as solvent residues might be trapped inside the electrospun nanofibers. Accurate control over solvent residues becomes crucial in the case of large-volume, solvent-based electrospinning for biomedical and pharmaceutical applications . HoweverThe recent development of needleless electrospinning offers the possibility for fabricating electrospun nanofibers on a large scale and addresses the issue of clogging at the spinnerets . NeedlelThe electrospinning process has generated a lot of interest in various medical applications due to its ease of use, adaptability and flexibility in controlling the fiber diameter from the micrometer down to the nanometer range. Though this method has been in use for a few decades now, the techniques and the equipment used in the electrospinning process are ever-evolving. Electrospinning began with a single nozzle configuration and evolved into multi-nozzle configurations through coaxial and emulsion spinning configurations. Further studies are being carried out to modify the nozzle configuration and collector design in order to significantly improve fiber properties and simplify the manufacturing process. This review summarizes several key aspects of electrospinning in the use of electrospun fibers in drug delivery with a special emphasis on electrospun nanofibers impregnated with nanoparticles. Studies involving biomedical applications of electrospun nanofibers are also discussed. By careful selection of polymers, it is now possible to deliver various antibacterial agents and anticancer drugs in a required manner using electrospun nanofibers. In order to make further progress, particularly in the field of drug delivery, it is necessary to identify ways that allow large-scale fabrication of nanofibers with desired morphological and mechanical properties in a reproducible manner. Despite the relentless efforts being made by academic and industrial scientists, much of the research conducted with electrospun fibers is in vitro. Further progress in the field of electrospun nanofibers will require continued assessment in vivo. Scientists working in this field have to identify ways to use nanofibers for immunotherapy, gene therapy and regenerative medicine in order to meet the current, as well as any future, demands in drug delivery.


Epoxyeicosatrienoic acids (EETs) are metabolites of arachidonic acid via cytochrome P450 (CYP)/epoxygenase and are hydrolyzed by soluble epoxide hydrolase (sEH). Circulating and tissue levels of EETs are controlled by CYP (EET synthesis) and sEH (EET degradation). Therefore, both increases in CYP activity and decreases in sEH expression potentiate EET bioavailability, responses that prevail in the female sex as a function of estrogen. This mini review, based on subtitles listed, briefly summarizes studies focusing specifically on (1) female-specific potentiation of CYP/epoxygenase activity to compensate for the endothelial dysfunction; and (2) estrogen-dependent downregulation of sEH expression, which yields divergent actions in both systemic and pulmonary circulation, respectively.Estrogen-Potentiating EET Synthesis in Response to Endothelial Dysfunction: This section summarizes the current understanding regarding the roles of estrogen in facilitating EET synthesis in response to endothelial dysfunction. In this regard, estrogen recruitment of EET-driven signaling serves as a back-up mechanism, which compensates for NO deficiency to preserve endothelium-dependent vasodilator responses and maintain normal blood pressure.Ephx2/sEH Expression:Estrogen-Dependent Downregulation of  This section focuses on molecular mechanisms responsible for the female-specific downregulation of sEH expression.Roles of EETs in Systemic Circulation, as a Function of Estrogen-Dependent Downregulation of sEH: This section summarizes studies conducted on animals that are either deficient in the Ephx2 gene (sEH-KO) or have been treated with sEH inhibitors (sEHIs), and exhibit EET-mediated cardiovascular protections in the cerebral, coronary, skeletal, and splanchnic circulations. In particular, the estrogen-inherent silencing of the Ephx2 gene duplicates the action of sEH deficiency, yielding comparable adaptations in attenuated myogenic vasoconstriction, enhanced shear stress-induced vasodilation, and improved cardiac contractility among female WT mice, male sEH-KO and sEHI-treated mice.Roles of Estrogen-Driven EET Production in Pulmonary Circulation: This section reviews epidemiological and clinical studies that provide the correlation between the polymorphism, or mutation of gene(s) involving estrogen metabolism and female predisposition to pulmonary hypertension, and specifically addresses an intrinsic causation between the estrogen-dependent downregulation of Ephx2 gene/sEH expression and female-susceptibility of being pulmonary hypertensive, a topic that has never been explored before. Additionally, the issue of the “estrogen paradox” in the incidence and prognosis of pulmonary hypertension is discussed. It is established that oxidative metabolism of arachidonic acid through the cyclooxygenase (COX) and lipoxygenase pathways to biologically activate eicosanoids plays a critical role in the regulation of pathophysiological processes. To date, a dubbed “third pathway” of the cytochrome P450 (CYP)/epoxygenase system has come to the forefront of research with aims to evaluate the pathophysiological significance of its biologically active mediators, epoxyeicosatrienoic acids (EETs) . EETs poCytochrome P450 are encoded by a complex superfamily of genes; they are located in the endoplasmic reticulum and add an epoxide across one of the four double bonds of arachidonic acid to produce four EET regiosomers: 5,6-EET, 8,9-EET, 11,12-EET and 14,15-EET. The CYP2C and CYP2J families are responsible for the majority of EET generation in mammals . Specifiin vitro studies aiming to evaluate CYP activities were performed in the presence of inhibitors of endothelial nitric oxide synthase (eNOS) and COX. More intriguingly, the CYP/EET-evoked compensatory action exerts in a female favorable manner, as indicated by the evidence that in eNOS and COX-1 double knockout (KO) mice, EET-mediated responses via an EDHF-based event contribute significantly to the preservation of endothelium-dependent relaxation, coinciding with normal blood pressure in female animals , evokes a solely EET-mediated response that fully preserves shear stress-induced vasodilation  , reminis of SSID . Thus, i of SSID .Ephx2 gene and extensively expressed in multiple organs/tissues including vasculatures; it converts epoxides to diols by adding water to open the epoxide, thus inactivating EETs  or treatment with sEHIs in male mice reduced their blood pressure to the level comparable to that of wild type (WT) females; in the latter, disruption of the Ephx2 gene further reduced blood pressure but with significantly smaller decrement than in male counterparts  among female WT, male sEH-KO mice and male WT mice treated with sEHIs , there are other two unique features in the pulmonary circulation that differ from the systemic circulation: (1) hypoxia pulmonary vasoconstriction (HPV)  that is Thus, human studies show a female to male ratio of 4.3:1 among the total PH patients , and 4.1BMPR2 gene turns out to be one of the most important genetic-based alterations responsible for the sex-bias in IPAH  involving estrogen metabolisms and the female predisposition to PH . In gene in IPAH , as the in males . Female pression , followession  of estrogens in the incidence and prognosis of PH . Figure  failure , whereas failure  and rest failure . In mono failure . This poper se, but rather by alternative pathways. Indeed, in addition to targeting sEH, sEHIs are capable of binding with other enzymes due to the presence of multi-target ligands. For instance, PTUPB is a tight COX-2 binder  reveled that GSK alleviated endothelial dysfunction in COPD patients . Since H2 binder  and TPPU2 binder . As suchWe briefly summarized the pathophysiological significance of potentiating EET production and/or inhibiting EET hydrolysis, as a function of estrogen, in the regulation of systemic and pulmonary circulations. In the systemic circulation, increases in EETs afford better cardiovascular performance and lower incidence of ischemic diseases in women. In the pulmonary circulation, clinical development of PH appears to require “two hits” that can be triggered by either genetics (sex and gene polymorphisms), environmental factors (hypoxia and sex hormone dysmetabolism), or both . Thus, from a pros and cons point of view, targeting CYP and sEH may prove to be a double-edged sword with beneficial and adverse effects on systemic and pulmonary circulatory systems; this brings concerns surrounding the use of sEHIs as therapeutic regimens, or the consideration of sEHI-related clinical trials in female populations who bear hyper-responsiveness to acquired pathological insults (as second hits) to the respiratory system.AH and DS contributed equally to the literature search, figures, and writing, and gave their final approval of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Introduction: We present a case of unusual combination of sarcoidosis with refractory pyoderma gangrenosum. Pyoderma gangrenosum has been a reported with other autoimmune disorders like rheumatoid arthritis and ulcerative colitis, but it has hardly ever been reported with another rare disease like sarcoidosis. Here we present a case of 39 year old female, who was diagnosed and treated for sarcoidosis but later presented to our OPD with worsening skin lesions which were biopsied and diagnosed as pyoderma gangrenosum. Not only were they unusual, but they were even refractory to standard medical therapy.Case description: A 39 year old female, was referred to our OPD for further management once the diagnosis of sarcoidosis was confirmed. Symptoms started in December 2014, when she had complaints of fever along with cough. Further evaluation was done and a chest CT revealed multiple mediastinal lymph nodes. Patient was started on empirical anti tubercular drugs (four drug regimen) ,which was given for six months . She felt better during the treatment course but after a period of two months, she started having again low grade fever with myalgias and arthalgias. Patient was admitted under internal medicine and was extensively evaluated. Repeat chest CT was suggestive of multiple mediastinal lymph nodes, with no shrinkage in size as compared to the initial one. Her erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) were still elevated (76 and 54) while the angiotensin converting enzyme (ACE) levels were slightly high(86). Endoscopic ultrasound-guided fine needle aspiration cytology (FNAC) was done which was suggestive of granulomatous inflammation . Considering the diagnosis of sarcoidosis, she was started on intramuscular methylprednisolone acetate 80 mg weekly for four weeks along with hydroxychloroquine. She reported improvement in fever and myalgias in the next visit and the inflammatory markers had normalised as well. But a month later, she presented to our OPD with new skin lesions over the arms and legs, one of which had appeared after minor trauma to the skin. Patient was admitted, and a deep skin biopsy was done from the ulcer site which was suggestive of pyoderma gangrenosum. There were no associated gastrointestinal symptoms or arthritis (the most common causes of pyoderma gangrenosum). The inflammatory markers at this stage were still normal (ESR 24 and CRP 5.0). She was started on oral corticosteroids (1 mg/kg) along with azathioprine (as steroid sparing drug). The patient had partial improvement but while tapering steroids to 20 mg/day, the lesions worsened and the new lesions appeared over the breasts, biceps area and the thighs. She was already experiencing steroid side effects related to a high dose of long duration. Biopsy was rechecked to confirm the diagnosis, but the conclusion remained the same. A therapeutic trial more of several additional weeks was planned before switching the medications. On the fourth follow-up visit, the skin lesions had worsened and began to increase in size over the thighs, breasts and the legs. She received a prolonged course of antibiotics along with daily dressings but all to no avail. Considering refractory disease, azathioprine was discontinued and patient was switched to injectable cyclophosphamide, according to literature for refractory pyoderma gangrenosum. She was given six monthly cycles, during which the patient showed significant improvement. But since the lesions persisted, we added colchicine 0.5 mg twice daily and gave two more doses of injectable cyclophosphamide . The lesions began to heal after second dose but she needed prolonged injectable course to help the disease go in remission. The steroid could be tapered down to 5 mg/day. Currently, our patient is doing well with no new lesions. She has been kept on azathioprine maintenance therapy along with prednisolone 5 mg/day.Discussion: Pyoderma gangrenosum is a rare disorder causing multiple non-healing ulcers over the limbs, aggravated by trauma. It is a neutrophilic dermatosis which has been associated mostly with inflammatory bowel disease and haematological malignancies, although most cases remain idiopathic. The association with sarcoidosis is a rare occurence. We did a review of the medical literature and found only four previously reported cases . The pathogenesis of pyoderma gangrenosum is immune mediated with predominant neutrophil infiltration. There has been an association with TNF-alpha, IL-8 and I-23 especially the variant associated with immunological diseases. Sarcoidosis is also an immune mediated disease with increased accumulation of CD 4 + T cells at the site of inflammation. The cytokine response in granulomatous inflammation is mediated by IL-2, TNF-alpha and interferon gamma. Skin manifestations are a known complication in sarcoidosis with manifestations like lupus pernio, erythrema nodousum, maculopapular rashes and keloid scars due to underlying inflammation. Pyoderma gangrenosum has not been reported due to the rarity of the cases being diagnosed. Both conditions have a common denominator of increasing skin lesions after injury, hence there can be a relation between the two. Our patient had a refractory pyoderma gangrenosum, which kept recurring upon tapering steroids. Her lesions had a predilection for areas of trauma, as the ulcers would appear on the sites she got injury with delayed healing. We did a literature search where cyclophosphamide and TNF Blockers has been recommended for refractory pyoderma gangrenosum. She showed a dramatic response to cyclophosphamide as after the first dose the wounds started to heal. After three doses of cyclophosphamide, she showed a much better response with larger lesions healing up as well. This case does give us a relationship between sarcoidosis and pyoderma gangrenosum. Unless one has a high index of suspicion, these cases may be overlooked. A chest roentgenogram should be recommended in all patients with idiopathic pyoderma gangrenosum.Key Learning Points: This case does give us a relationship between sarcoidosis and pyoderma gangrenosum. Unless one has a high index of suspicion, these cases may be overlooked. A chest roentgenogram should be recommended in all patients with idiopathic pyoderma gangrenosum.Disclosure: N. Mendiratta: None. S. Bajad: None. M. Bindroo: None.


The data presented in this article are related to the research article entitled “New synthesis strategy for hollow NiO nanofibers with interstitial nanovoids prepared via electrospinning using camphene for anodes of lithium-ion batteries” [1]. Hollow NiO nanofibers were prepared by electrospinning process using camphene and subsequent heat-treatment process with various temperatures. The data presented in this manuscript showed the effect of the heat-treatment temperature of the as-spun fibers on the lithium ion storage properties of the hollow NiO nanofibers as anodes for lithium ion batteries. Each FE-SEM image, XRD pattern, cycle, and rate properties of the hollow NiO nanofibers obtained at various heat-treatment temperatures were investigated. The discharge capacities of T-300, T-350, and T-400 after 400 cycles were 554, 956, and 471 mA h g−1, respectively. The rate properties of the samples are shown in −1. The final discharge capacities of T-300 at 0.5, 1.5, 3.0, 5.0, 7.0, and 10.0 A g−1 were 807, 603, 450, 351, 290, and 233 mA h g−1, respectively. The final discharge capacities of T-350 at 0.5, 1.5, 3.0, 5.0, 7.0, and 10.0 A g−1 were 1050, 1024, 937, 796, 643, and 483 mA h g−1, respectively. The final discharge capacities of T-400 at 0.5, 1.5, 3.0, 5.0, 7.0, and 10.0 A g−1 were 921, 906, 824, 733, 642, and 528 mA h g−1, respectively.The lithium ion storage properties of the samples obtained at different heat-treatment temperatures were shown in


A related concern in the field is the over-reliance on a finite number of chosen preclinical models, leading to early clinical translation when the probe might not be optimized for high intertumor variation or intratumor heterogeneity. The ultimate potential may require multiple probes, as are used in molecular pathology, and a combination with ultrahigh-resolution imaging and image recognition systems, which capture the data at a finer granularity than is possible by the surgeon. Alternatively, one might choose a more generalized approach by developing the tracer based on generic hallmarks of cancer to create a more “one-size-fits-all” concept, similar to metabolic aberrations as exploited in fluorodeoxyglucose - positron emission tomography (FDG-PET)  or tumor acidity. Finally, methods to approach the problem of production cost minimization and regulatory approvals in a manner consistent with the potential revenue of the field will be important. In this area, some solid steps have been demonstrated in the use of fluorescent labeling commercial antibodies and separately in microdosing studies with small molecules.Molecular image-guided surgery has the potential for translating the tools of molecular pathology to real-time guidance in surgery. As a whole, there are incredibly positive indicators of growth, including the first United States Food and Drug Administration clearance of an enzyme-biosynthetic-activated probe for surgery guidance, and a growing number of companies producing agents and imaging systems. The strengths and opportunities must be continued but are hampered by important weaknesses and threats within the field. A key issue to solve is the inability of macroscopic imaging tools to resolve microscopic biological disease heterogeneity and the limitations in microscopic systems matching surgery workflow. A related issue is that parsing out true molecular-specific uptake from simple-enhanced permeability and retention is hard and requires extensive pathologic analysis or multiple  The lack of choice and features in these systems has been a problem in the past, but today there are many systems that have been steadily improving. This success has largely been due to resurgence in the use of indocyanine green (ICG) for tissue perfusion assessment, largely driven by Novadaq Tech. Inc. (now acquired by Stryker Corp.) pushing the field forward to evolve the commercial market for this type of direct visual guidance.,The key major opportunity for this field lies in the widespread adoption of molecular pathology and proteomic/genomic phenotyping of diseases in all major care centers.a priori, and the efficacy in vivo is being tested in these early phase trials. At this point, most of the more specific agents have not advanced beyond Phase 1 trial, but there is potential. However, some of the factors listed next will determine success or failure of these trials.The other major opportunity has been the creative growth in molecular probes for surgical guidance. A listing of those registered with clinicaltrials.org for patient recruitment is shown below in in vivo. Although bulk tissue removal can still aid the surgeon as guided by the fluorescence signal, there are diseases, such as brain or ovarian cancer, where microscopic pathology removal is more critical and macroscopic views of the fluorescence will be insufficient. Pathology analysis of immunohistochemistry slides routinely shows high morphological heterogeneity in tumor nodules and ducts, indicating that specific molecular probes to these targets would likely also be microscopically heterogeneous. A pathologist is trained to view this heterogeneity and make sense of it, knowing that staining patterns are usually complex, and it is this complexity that can be one of the keys to diagnosis. Yet a surgeon must perform resection at the macroscopic or perhaps mesoscopic resolution. The macroscopic view of a heterogeneous uptake blurs out the uptake and observed contrast, and so in vivo levels of tumor to background are routinely seen near ,The weaknesses of the field are the issues that will limit the successes if not solved. One of the most obvious and yet least discussed aspects of the field is that molecular-specific dyes have a localization that is often microscopically heterogeneous, and so this is not easily appreciated with surgical imaging in vivo, ignoring the tumor specificity it delivers in ex vivo analyses. As such, they might see a mismatch between in vivo and in vitro data and stop development of new molecular probes. So, while this problem can seem trivial, without a technological or logistical solution to it, there will remain a lack of clarity in target validation. What remains pivotal is comparing imaging data with the accepted gold standard in daily clinical care, which remains simple H/E histopathology. One practical issue is the comparison of frozen section fluorescence imaging to fixed H&E pathology images, which requires challenging and labor-intensive comparison studies.,,in vivo–ex vivo assay. Efforts to apply deep learning systems comparing in vivo fluorescence with ex vivo tumor delineation might be a solution in this complex equation. This is an important part of clinical trial testing, which without an appreciation for it could lead to a slowing of enthusiasm because of a perceived low contrast, stymying research and development in this field.One solution to imaging of the cancer heterogeneity issues, is to have a combination of: (1) a widefield high-resolution imaging system and (2) an automated software to recognize and classify the molecular morphology patterns. This would be similar to what is seen in immunohistochemistry, and automation of this is coming, but for now it remains one of the key weaknesses of the field, where the observed contrast levels seen are distinctly unimpressive. Alternatively, it is possible that fast magnification change imaging could be possible where the low power, low magnification can be quickly swept up to higher power and high resolution. The value of this is that the scoping in and out from macroscopic to microscopic imaging would allow for superior workflow for the surgeon, who desires the ability to see boundaries or fine structures better. Without these types of tools, many companies developing molecular probes may fail to see the high contrast ,in vivo is implausible, although some methods are being proposed now.Another major weakness of the field, which pervades all of molecular cancer research, is the over-reliance of preclinical testing in tumor cell lines that are monolithically positive for the molecular target of interest.in vivo pathology of each individual patient being imaged.Given this, the ability to know the expression level and the heterogeneity of expression may be the key to allowing success in use of the molecular probe.,Another weakness of the field is the regulatory processes that limit development and deployment of imaging probes to one at a time, with good manufacturing practices (GMP) production requirements, release testing, toxicity testing, and ongoing stability analysis for each molecule.The threats to FGS are perhaps the most troubling and require some careful thought before they derail progress in the field. Although each threat may not have immediate solutions, it is critical to appreciate them. The largest and most pressing threat is the fact that clinically observed contrast and uptake data are being misinterpreted at times in terms of the specificity of the signal. As previously mentioned in the weakness area, limitations of current macroscopic imaging systems can be a problem that leads to an underestimation of the specificity for the molecular probe to the target. But an even further problem that is more insidious is that macroscopic imaging tools simply cannot provide verified information about the specificity of uptake. It is very common for researchers to interpret the macroscopic contrast as the specificity of probe, and yet, factors such as vascular perfusion and lymphatic impairment can often have more effect upon the uptake than any specificity of binding or localization. This is well known as the enhanced permeability and retention (EPR) effect,,,,It is very common to simply compare two tumor lines, one positive for the target, and one negative for the target as a quantitative assay of specificity of uptake. Yet, while this seems reasonable, these two tumor lines can easily have much larger vascular perfusion and lymphatic flow differences, which will dwarf the effects of the probe binding. Preclinical testing of specificity in this manner, while seemingly logical, based just on the target, adds a high level of randomness to the field, and many published studies have data that are confounded by this issue. Solutions to this problem are complex and require more careful microscopic and pathologic analysis of uptake, comparing matched targeted and untargeted probes, to ensure that the measured bulk uptake is actually bound to the target or activated by the expected target. Solutions such as using a reference tissue to normalize are insufficient,in vivo toxicity testing in a way that allows the cost to match the benefit.,,,,,Another key threat is the lack of an obvious way to advance production and ,,per se, there are approaches such as in vivo pharmacokinetic imaging,in vivo optical imaging to assess activity and target bioavailability will likely be useful to confirm the value of agents.in vivo kinetics and clearance organ behavior of both the dye and the targeting moiety can alleviate some of the costs of preclinical work,,Toxicity and release testing will always be a necessary and expensive part of the process as well, which cannot be avoided. However, there are methods that can limit costs, if, for example, the imaging is done with therapeutic proteins that have already undergone substantial human use, and can be slightly modified without altering their pharmacokinetic behavior. In these cases, minimal testing might be feasible under negotiation with regulatory bodies.,The threat about economic viability for new molecular probes is less of an issue for small molecules than it is for larger biological agents, and it is likely that the two will have very different development paths. It is possible that once the safety of fluorescently labeled therapeutic antibodies has been better established, one could make the case that small labeling changes are relatively minor, and that these agents, if made under certain parameters, should inherently have FDA clearance if the antibodies are already in common human use.The perceived value of molecular guided surgery is exceptionally high. It comes with the promise of combining molecular pathology information into a nuclear medicine-like molecular guidance, which would improve the precision of surgical procedures. Increasing interest by different surgical specialties has been robust,


Astragalus membranaceus, that has been reported to have bioactivities against cardiovascular disease and kidney disease. There is limited information on the metabolism of astragaloside IV, which impedes comprehension of its biological actions and pharmacology. In the present study, an ultra-performance liquid chromatography coupled with quadrupole/time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS)-based approach was developed to profile the metabolites of astragaloside IV in rat plasma, bile, urine and feces samples. Twenty-two major metabolites were detected. The major components found in plasma, bile, urine and feces included the parent chemical and phases I and II metabolites. The major metabolic reactions of astragaloside IV were hydrolysis, glucuronidation, sulfation and dehydrogenation. These results will help to improve understanding the metabolism and reveal the biotransformation profiling of astragaloside IV in vivo. The metabolic information obtained from our study will guide studies into the pharmacological activity and clinical safety of astragaloside IV.Astragaloside IV is a compound isolated from the Traditional Chinese Medicine  The metabolism of drugs in organisms is an important biotransformation process. The study of the metabolite profile of a candidate compound plays an important part in demonstrating the basic information about a biological process as well as the pharmacology and toxicology of a drug . DetectiRadix astragali − molecular ion of metabolites had a weight of 697.4193 Da, which heralded a molecular formula of C37H62O12. Compared with the literature, the molecular weight of metabolites was 132 Da less than that of the parent drug, demonstrating that these metabolites were dexylcosylation metabolites of astragaloside IV. The fragment ion at m/z 471.2417 was formed by the loss of 46 Da (formic acid) along with 162 Da (hexose) and 18 Da water from the precursor ion (m/z 697.4193), which suggested the loss of glucose and dehydration. Hence, M2 and M3 were identified as isomers of dexylcosylation products of astragaloside IV. However, the detailed mechanisms for the formation of isomeric remain unknown. The mass spectra of M2 and M3 are shown in Metabolites M2 and M3 had retention times of 6.00 min and 6.77 min, respectively. The [M+HCOOH‒H]m/z 667.4044 Da  was eluted at a retention time of 6.78 min. Metabolite M4 had a molecular formula of C36H60O11, which was 162 Da less than that of the parent drug. Comparison with the literature revealed M4 to be a hydrolyzed metabolite with deglycosylation of the parent drug. The fragment ion of M4 at m/z 515.3008 was formed by loss of 152 Da from the precursor ion (m/z 667.4044). This finding suggested the loss of 132 Da (pentose) and 18 Da water and 2 Da hydration and dehydrogenation, which further confirmed the conclusion made above. Hence, M4 was identified to be a deglycosylation metabolite of astragaloside IV. The mass spectrum is shown in The molecular ion detected at − molecular ions of metabolite M5 and M6 had a weight of 665.4269 Da, which heralded a molecular formula of C37H62O8. The molecular weights of M5 and M6 were 164 Da less than that of the parent drug. Compared with literature, M5 and M6 were glucuronidation and deglycosylation metabolites of the parent drug ent drug . The fram/z 663.3753Da  was eluted at a retention time of 4.32 min. The metabolite M7 had a molecular formula of C36H56O11, which had a weight that was 2 Da less than that for M5 and M6. Comparison with the literature revealed M7 to be the glucuronidation, deglycosylation and dehydrogenation metabolite of astragaloside IV. The fragment ion of M7 at m/z 615.2835 was formed by loss of 46 Da (formic acid) along with 2 Da (hydrogen) from the precursor ion (m/z 663.3753). The ions at m/z 599.2879 and m/z 583.2925 were caused by deoxygenation. The ion at m/z 441.1942 was formed by the loss of glucuronate, which further confirmed the conclusion made above. Hence, M7 was identified as the glucuronidation, dehydroxylation, deglycosylation and dehydrogenation metabolite of astragaloside IV. The mass spectrum is shown in The molecular ion detected at − molecular ion of metabolite M8 had a weight of 651.4109 Da, which heralded a molecular formula of C36H60O10. The molecular weight of M8 was 178 Da less than that of the parent drug. The MS/MS fragment ion of M8 at m/z 519.2768 was formed by the loss of C5H8O4 (132 Da) from the precursor ion (m/z 651.4109). The fragment at m/z 519.2768 indicated the loss of xylose and a methyl group. Hence, M8 was identified as a deglycosylation and dehydroxylation metabolite of astragaloside IV. The mass spectrum of M8 is shown in Metabolite M8 had a retention time of 6.96 min. The [M+HCOOH−H]m/z 619.4197  was eluted at a retention time of 6.43 min. Metabolite M9 had a molecular formula of C36H60O8, which carried a weight 210 Da less than that of the parent drug. M9 was confirmed to be loss of 46 Da (formic acid) along with the deglycosylation and dehydrogenation metabolite of the parent drug. The fragment ion of M9 at m/z 453.2858 was formed by the loss of xylose, oxygen and dehydratation from the precursor ion (m/z 619.4197). The ion produced at m/z 407.2801 was formed by the loss of hydroxy and methyl groups. Ion information further confirmed our conclusion. Hence, M9 was identified as a deglycosylation and dehydrogenation metabolite of astragaloside IV. The mass spectrum is shown in The molecular ion detected at m/z 569.3162 Da  was eluted at a retention time of 7.34 min. Metabolite M10 had a molecular formula of C30H50O8S, which was a weight of 260 Da less than that of the parent drug. The fragment ion of M10 at m/z 551.2433 was formed by the loss of hydration from the precursor ion (m/z 569.3162). The ion at m/z 519.2256 was caused by the loss of two hydroxyl groups from the ion (m/z 551.2433). The fragment ion of m/z 485.2810 carried a weight that was 34 Da less than the ion at m/z 519.2256, which was caused by the loss of a hydroxy group and hydration. The fragment ion of M10 at m/z 389.2687 indicated the loss of a sulfate group from the precursor ion (m/z 485.2810). Hence, M10 was identified as a deglycosylation and sulfation metabolite of astragaloside IV. The mass spectrum is shown in The molecular ion detected at m/z 553.3212 Da  was eluted at a retention time of 7.74 min. Metabolite M11 had a molecular formula of C30H50O7S, which carried a weight that was 16 Da less than that of M10. The fragment ion of M11 at m/z 515.0021 was formed by the loss of an isopropyl group  from the precursor ion (m/z 553.3212). The ion at m/z 485.2805 was formed by the loss of methyl and hydroxy groups from ion m/z 515.0021. The fragment ion of M11 at m/z 391.2842 was the isoform of the ion that lost a sulfate group from the precursor ion (m/z 485.2810). Hence, M11 was identified as a deglycosylation and sulfation metabolite of astragaloside IV. The mass spectrum is shown in The molecular ion detected at m/z 539.3059 Da  was eluted at a retention time of 6.40 min. Metabolite M12 had a molecular formula of C29H48O7S, which is a weight of 14 Da less than that of M11, suggesting that it was probably produced by neutral loss of a methyl group. The fragment ion at m/z 405.2654 was caused by loss of sulfate and methyl groups as well as a hydroxyl group associated with dehydrogenation. Therefore, M12 was identified as a hydrolyzed metabolite that underwent deglycosylation, sulfation and demethylation of the parent drug. The mass spectrum is shown in The molecular ion detected at − molecular ions of metabolite M14 carried a weight of 535.3640 Da, which heralded a molecular formula of C31H52O7. The molecular weight of M14 was 294 Da less than that of the parent drug, suggesting that M14 was a metabolite formed by loss of 46 Da (formic acid) along with di-deglycosilation and deglycosylation of the parent drug. The fragment ion of M14 at m/z 389.2696 was formed by the loss of hydroxyl and methyl groups from the precursor ion (m/z 535.3640). Hence, M14 was identified as the hydrolyzed metabolite of astragaloside IV. The mass spectrum is shown in Metabolite M14 had a retention time of 8.05 min. The [M+HCOOH−H]29H48O6S. The molecular weight of M15 was 16 Da less than that of M12 and M13, suggesting that M15 was a dehydroxylation metabolite of M12 or M13. The fragment ion of M15 at m/z 389.2691 was caused by the loss of sulfate and methyl groups and a hydroxyl group associated with dehydrogenation from the precursor ion (m/z 523.3094), which further confirmed the above conclusion. Hence, M15 was identified as a deglycosylation, sulfation, demethylation and hydroxylation metabolite of astragaloside IV. The mass spectrum of M15 is shown in Metabolite M15 had a retention time of 8.93 min. The molecular weight of metabolite M15 was 523.3094 Da, which heralded a molecular formula of Cm/z 473.3263 Da  was eluted at a retention time of 6.89 min. Metabolite M16 had a molecular formula of C29H46O5, the weight of which was 356 Da less than that of the parent drug. The fragment ion of M16 at m/z 451.2698 was formed by dehydration and dehydrogenation. The fragment ion at m/z 405.2636 was formed by the loss of an isopropyl group. Hence, M16 was identified as a deglycosylation and dehydroxylation metabolite of astragaloside IV. The mass spectrum is shown in The molecular ion detected at 46H76O20. The molecular weight of M17 was 130 Da greater than that of the parent drug, suggesting that M17 was a metabolite formed by loss of 46 Da (formic acid) along with glucuronation of the parent. The fragment ion at m/z 485.2800 was formed by the loss of glucuronic acid, glucose, hydrogen and xylose from the precursor ion (m/z 959.487). The fragment ion at m/z 469.2490 was formed by dehydroxygenation of the ion at m/z 485.2800. Hence, M17 was identified as a glucuronation and demethylation metabolite of astragaloside IV. The mass spectrum is shown in Metabolite M17 had a retention time of 7.91 min. The molecular ions of metabolite M17 had a weight of 959.4852 Da, which heralded a molecular formula of C41H68O17S. The molecular weight of M18 and M19 was 50 Da larger than that of the parent drug, suggesting that M18 and M19 were isomers of the parent drug formed by loss of 46 Da (formic acid) along with sulfate conjunction metabolites. The fragment ion at m/z 651.0512 was formed by the loss of SO4 (96 Da) and xylose (132 Da) from the precursor ion (m/z 879). The fragment ion at m/z 651.0512 lost a glucose molecule (162 Da) and four hydrogen atoms (4 Da) to form the ion at m/z 485.2690. Hence, M18 and M19 were identified as sulfation metabolites of astragaloside IV. The mass spectra of the metabolites are shown in Metabolites M18 and M19 had retention times of 7.81 and 7.62 min, respectively. The molecular weight of metabolite M18 and M19 was 879.4373 Da, which heralded a molecular formula of C42H70O15. The molecular weight of M20 was 16 Da less than that of the parent drug, suggesting that M20 was a metabolite formed by loss of 46 Da (formic acid) along with methoxylated the metabolite of the parent drug. The fragment ion of M20 at m/z 775.5131 was formed by the loss of hydrogen atoms and hydroxyl groups from the precursor ion (m/z 813.4667). The fragment ion at m/z 387.2538 was formed by the loss of glucose, xylose, methyl and hydroxyl, which further confirmed the above conclusion. Hence, M20 was identified as a methoxylated metabolite of astragaloside IV. The mass spectrum is shown in Metabolite M20 had a retention time of 8.14 min. The molecular weight of metabolite M20 was 813.4667 Da, which heralded a molecular formula of Cm/z 811.4180 Da  was eluted at a retention time of 6.92 min. Metabolite M21 had a molecular formula of C42H68O15, which carried a molecular weight that was 18 Da less than that of the parent drug. The fragment ion of M21 at m/z 405.2648 was formed by the loss of glucose, glycuronic acid and isopropanol, with cleavage of the furan ring at sites 22 and 23 (406 Da) from the precursor ion (m/z 811.4180), which further confirmed the above conclusion. Hence, M21 was identified as a dexylcosylation and glucuronation metabolite of astragaloside IV. The mass spectrum is shown in The molecular ion detected at m/z 809.4262 Da  was eluted at a retention time of 6.86 min. Metabolite M22 had a molecular formula of C42H66O15, which carried a molecular weight that was 20 Da less than that of M21. The fragment ion of M22 at m/z 405 was formed by the loss of formic acid, glucose, glycuronic acid and isopropanol, along with cleavage of the furan ring at sites 22 and 23 (404 Da) from the precursor ion (m/z 809.4262), which further confirmed the above conclusion. Hence, M22 was identified as a dehydrogenation metabolite of M21. The mass spectrum is shown in The molecular ion detected at In general, there were nine phase I metabolites of astragaloside IV in rats. The main processes were deglycosylation, dehydrogenation and hydroxylation. The phase II metabolites of astragaloside IV in rats included one methylation product (M20), six glucuronides  and seven sulphation ones . The possible metabolic pathways of astragaloside IV are summarized in An UPLC-Q-TOF-MS/MS-based approach was applied to the metabolite profiling of astragaloside IV in plasma, bile, urine and feces samples from rats. A total of 22 metabolites was detected and tentatively identified, and 15 of them were reported for the first time. The major metabolic pathways of astragaloside IV in rats were deglycosylation, sulphation and glucuronidation. Physicochemical characteristics such as high molecular weight, high hydrogen-bonding capacity, high molecular flexibility, and poor membrane permeability may contribute to the poor bioavailability of saponins ,18. StudThe study protocol  was approved by the Ethical Committee of Nanjing University of Chinese Medicine .Astragaloside IV (purity >98% as determined by high-performance liquid chromatography (HPLC)) was purchased from the National Institute for the Control of Pharmaceutical and Biological Products . Formic acid, acetonitrile and methanol  were of HPLC grade. Water was produced by a Milli-Q Ultrapure Water system . Sodium carboxyl methyl cellulose (CMC-Na) was supplied by Sinopharm Chemical Reagents .Male Sprague-Dawley (SD) rats (250 ± 20 g) were acquired from SLAC . Rats were kept in controlled-environment room  for ≥1 week for adaptation. Animals were fasted overnight before experimentation.Astragaloside IV suspended in 0.5% CMC-Na was administered to three rats  on three consecutive occasions over 24 h. Before administration, blank blood samples were collected. The sampling time point of astragaloside IV in rat plasma was chosen as reported . Brieflyvia the oral route to three rats at a single dose of ≈80 mg/kg. Animals were then anesthetized by 1% pentobarbital sodium . An abdominal incision was made and the bile duct cannulated with PE-10 tubing  for collection of bile samples. A heating lamp was used for maintaining body temperature during experimental procedures to prevent hypothermic alterations. Drug-containing bile samples were collected for 24 h and stored at −70 °C until additional extraction and analyses. Meanwhile, blank bile samples were collected from an additional two rats.Astragaloside IV was administered Rats were placed in a metabolic cage for 24 h. Before experimentation, rats had been in metabolic cages for acclimatization for 2 days and given standard chow and water. Blank urine and feces samples were collected. Urine and feces samples were collected for 24 h. Rats were weighed and given astragaloside IV suspended in 0.5% CMC-Na (80 mg/kg body weight). Samples of urine and feces from three rats were collected for 48 h. All urine and feces samples were stored at −70 °C until additional extraction and analyses.18 SPE columns. After washing with 1 mL of water, analytes were eluted with 1 mL methanol. Eluents were centrifuged at 28,000 g for 10 min at 4 °C. An aliquot (2 μL) of supernatants was then injected into the UPLCQ-TOFMS/MS system.Plasma samples from individual animals were combined. After addition of 600 μL of methanol, plasma samples (200 μL) were vortex-mixed to precipitate plasma proteins, and then centrifuged at 3000 g for 10 min at 4 °C. Supernatants were loaded onto preconditioned C18 SPE columns directly. After washing with 1 mL of water, analytes were eluted with 1 mL methanol. Eluents were centrifuged at 28,000 g for 10 min at 4 °C. Supernatants (2 μL) were injected into the UPLCQ-TOFMS/MS system.Bile samples from individual animals were combined. Bile samples (500 μL) were loaded onto preconditioned C18 SPE column was preconditioned with 2 mL methanol and 2 mL water. Urine samples (500 μL) were added onto the SPE columns. After washing with 1 mL water, analytes were eluted with 1 mL methanol. Eluents were centrifuged at 28,000 g for 10 min at 4 °C. Supernatants (2 μL) were injected into the UPLC-Q-TOF-MS/MS system.Urine samples from individual animals were combined. The CFeces (0.5 g) from each blank feces sample and drug-containing sample were selected separately. Methanol (5 mL) was added and the suspension vortex-mixed for 5 min, followed by ultrasonic extraction for 30 min using a KQ5200 Ultrasonic Cleaner . The mixture was centrifuged at 3000 g for 10 min at 4 °C to precipitate insoluble content. The supernatant was dried with nitrogen gas at 37 °C. Subsequently, 1 mL methanol was added to each sample followed by ultrasonic extraction for 20 min and extracts centrifuged at 28,000 g for 10 min at 4 °C. Supernatants were passed through 0.45-μm membranes  and injected into the UPLC-Q-TOF-MS/MS system.UPLC separation was performed on an ACQUITY system  equipped with a binary solvent delivery system, auto-sampler and an ESI interface. Analytes were eluted on a ACQUITY BEH column . The column temperature and auto-sampler temperature were maintained at 30 °C and 4 °C, respectively.Analyses were undertaken on the ACQUITY system. The flow rate of the mobile phase was 0.3 mL/min and the volume injected was 2 μL. The mobile phase consisted of acetonitrile (A) and a 0.1% formic acid-water solution (B). Eluent gradients were: 5% A for 1 min; 5%–20% A from 1 min to 2 min; 20%–50% A from 2 min to 6 min; 50%–70% A from 6 min to 10 min; 70%–95% A from 10 min to 11 min; 95% A from 11 min to 11.5 min; 95% A from 11.5 min to 12 min.E. The conditions of ESI-Q-TOF-MS/MS analyses were: capillary voltage, –2.5 kV; sample cone, 30 V; extraction cone, 4.0 V; source temperature, 120 °C; desolvation temperature 450 °C; flow rate of cone gas, 50 L/h; flow rate of desolvation gas (N2), 800 L/h.A Xevo G2 Q-TOF Mass Spectrometer (Waters) was used in negative ESI mode for data acquisition using UPLC/MSTM interface. The [M−H]− of leucine-enkephalin infused at 20 μL/min was used as a reference lock mass (m/z 554.2615 Da) at 200 pg/μL. Centroided data were acquired for each sample from 100 Da to 1500 Da. Dynamic range enhancement was applied throughout the MS experiment to ensure accurate mass measurements over a wide dynamic range. Accurate mass and elemental composition for precursor ions and fragment ions were analyzed using Mass Lynx v4.1.The mass instrument was calibrated using sodium formate. Mass accuracy and reproducibility were maintained using a Lock SprayData analyses were carried out with MetaboLynx. Analytical results of drug-contained samples and control were compared using MetaboLynx with a list of potential metabolites. Metabolites were detected and identified automatically.in vivo, and information gained from the present study is relevant to the pharmacological activity of astragaloside IV.The metabolite of astragaloside IV in rat plasma, bile, urine and feces samples were studied by UPLC-Q-TOF-MS/MS method. After adminitstration, astragaloside IV suffered with phases I and II metabolic reactions and generated 22metabolites were detected, 15 of them were reported for the first time. This study has improved our understanding of the metabolic profiling of astragaloside IV