[Evaluation and optimization of content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia].

This study discovered that the resolution of 3,5-O-dicaffeoylquinic acid(isochlorogenic acid A) in the content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia(ChP)(2020 edition) was poor, which affected accurate quantification. We tested the method in ChP with chromatographic columns of seven brands to clarify the problems in the existing method, optimized the chromatographic conditions by adjusting the mobile phase composition and elution ratio and replacing the chromatographic column packing, and carried out the reproducibility assay for the new method. The two methods were compared for the content determination results of Chrysanthemi Flos prepared from six different varieties. As evaluated by the resolution based on different chromatographic columns of seven brands, the existing method failed to separate isochlorogenic acid A and isochlorogenic acid D well. The peaks of the two components were not completely separated on three chromatographic columns, and isochlorogenic acid A and isochlorogenic acid D generated a co-effluent peak in the other four columns. Isochlorogenic acid A and isochlorogenic acid D could be completely separated under the optimized chromatographic conditions. The difference in the peak areas of isochlorogenic acid A+isochlorogenic acid D obtained by the optimized method and the method in ChP was not significant, with deviation less than 3.0%, which further proved that the result measured by the method in ChP was the co-effluent of isochlorogenic acid A and isochlorogenic acid D. The optimized method can ensure the accurate quantification of isochlorogenic acid A. The existing content determination method of Chrysanthemi Flos has the problem of poor resolution. It is recommended to revise the chromatographic conditions for the content determination method of Chrysanthemi Flos to improve the resolution of isochlorogenic acid A and ensure its accurate quantification.

Evaluation and optimization of content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia / 中国中药杂志

This study discovered that the resolution of 3,5-O-dicaffeoylquinic acid(isochlorogenic acid A) in the content determination method of Chrysanthemi Flos in Chinese Pharmacopoeia(ChP)(2020 edition) was poor, which affected accurate quantification. We tested the method in ChP with chromatographic columns of seven brands to clarify the problems in the existing method, optimized the chromatographic conditions by adjusting the mobile phase composition and elution ratio and replacing the chromatographic column packing, and carried out the reproducibility assay for the new method. The two methods were compared for the content determination results of Chrysanthemi Flos prepared from six different varieties. As evaluated by the resolution based on different chromatographic columns of seven brands, the existing method failed to separate isochlorogenic acid A and isochlorogenic acid D well. The peaks of the two components were not completely separated on three chromatographic columns, and isochlorogenic acid A and isochlorogenic acid D generated a co-effluent peak in the other four columns. Isochlorogenic acid A and isochlorogenic acid D could be completely separated under the optimized chromatographic conditions. The difference in the peak areas of isochlorogenic acid A+isochlorogenic acid D obtained by the optimized method and the method in ChP was not significant, with deviation less than 3.0%, which further proved that the result measured by the method in ChP was the co-effluent of isochlorogenic acid A and isochlorogenic acid D. The optimized method can ensure the accurate quantification of isochlorogenic acid A. The existing content determination method of Chrysanthemi Flos has the problem of poor resolution. It is recommended to revise the chromatographic conditions for the content determination method of Chrysanthemi Flos to improve the resolution of isochlorogenic acid A and ensure its accurate quantification.

Optimization of pectin extraction from selected Philippine fruit peel wastes using Box-Behnken design

Background@#Pectin is a pharmaceutically relevant excipient that can be upcycled from selected Philippine fruit peel wastes. Method optimization of pectin extraction leads to maximizing yields from limited resources, while also reducing environmental wastes, and providing local alternative sources. @*Objectives@#This study aimed to optimize the method of extracting pectin from selected Philippine fruit peel wastes using the Box-Behnken design, by varying the acid extraction solvent, treatment time, and working temperature. @*Methodology@#The three-level (-1, 0, 1) Box-Behnken design (15 set-ups) was used to optimize the pectin extraction in each of the fruit peel samples (C. maxima; A. heterophyllus; ripe and unripe M. indica; D. zibethinus; and H. undatus). The three experimental factors were the type of 3N acid used as extracting solvent (HNO₃, H₂SO₄, and HCl); duration of treatment in minutes (60, 90, and 120); and temperature of treatment in C 60, 75, and 90). The %yield was computed in each set-up, and the projected yields were generated using multiple linear regression. The pectin samples obtained from the optimized conditions were subjected to the physicochemical characterization, with apple pectin as the standard. Degree of esterification (DE), equivalent weight (EW), methoxy content (MC), alkalinity of ash (AA), and anhydrouronic acid content (AUA) were performed. @*Results@#Maximum yields were extracted from C. maxima (28.96%), A. heterophyllus (20.12%), ripe M. indica (26.23%), and unripe M. indica (25.89%), using 3N H₂SO₄, for a treatment duration of 60 minutes, at a working temperature of 90 C, and H. undatus (25.03%) at 60 C, for a treatment duration of 120 minutes. @*Conclusion@#Optimum conditions were identified to extract pectin in each of the fruit peel samples. The 3N H₂SO₄ produced the highest pectin yields in all of the set-ups, while the treatment time and working temperature vary per fruit peel sample. Pectin extract from C. maxima, A. heterophyllus, and M. indica was comparable to the standard.

[Optimization of determination methods of astragaloside â £ and analysis of contents of astragaloside â £ in different origin and grade].

A rapid and accurate method for determination of astragaloside Ⅳ was established,which was further applied to determine the contents of astragaloside Ⅳ in 87 batches of different origin and different grade of Astragali Radix. The ROC curve was used to analyze the contents of astragaloside Ⅳ in different origin. Simultaneous contents of astragaloside Ⅳ in different grade were compared with chemometrics. HPLC-ELSD method was used to determine the contents of astragaloside Ⅳ. A Vensil MP C18 column( 4. 6 mm×250 mm,5 µm) was used with acetonitrile-water( 32 ∶68) as the mobile phase at a flow rateof 1 m L·min-1. The column temperature was 25 ℃ with ELSD parameters as follows: gas flow rate was 2. 5 L·min-1,the drift tube heating temperature was set to 105 ℃,and the gain value was 4. 0. The optimized method avoided the problem that the consumable quality unstable and the recovery rate was not high. The contents determined by the optimized method were higher than the pharmacopoeia method,with less time and high recovery rate. The ROC curve analysis showed that there was no significant difference of contents of astragaloside Ⅳ between the top grade of Shanxi wild-simulated Astragali Radix top and the first grade of Gansu cultivated Astragali Radix. The contents of astragaloside Ⅳ in the second,third and fourth grade of Shanxi wild-simulated Astragali Radix was significantly higher than those of produced from Gansu.There was a significant negative correlation between the contents of astragaloside Ⅳ and grade in Shanxi Astragali Radix. While there was no correlation for Gansu Astragali Radix. This study provided the basis for the quality grade standard of Astragali Radix.

Optimization of determination methods of astragaloside Ⅳ and analysis of contents of astragaloside Ⅳ in different origin and grade / 中国中药杂志

A rapid and accurate method for determination of astragaloside Ⅳ was established,which was further applied to determine the contents of astragaloside Ⅳ in 87 batches of different origin and different grade of Astragali Radix. The ROC curve was used to analyze the contents of astragaloside Ⅳ in different origin. Simultaneous contents of astragaloside Ⅳ in different grade were compared with chemometrics. HPLC-ELSD method was used to determine the contents of astragaloside Ⅳ. A Vensil MP C18 column( 4. 6 mm×250 mm,5 μm) was used with acetonitrile-water( 32 ∶68) as the mobile phase at a flow rateof 1 m L·min-1. The column temperature was 25 ℃ with ELSD parameters as follows: gas flow rate was 2. 5 L·min-1,the drift tube heating temperature was set to 105 ℃,and the gain value was 4. 0. The optimized method avoided the problem that the consumable quality unstable and the recovery rate was not high. The contents determined by the optimized method were higher than the pharmacopoeia method,with less time and high recovery rate. The ROC curve analysis showed that there was no significant difference of contents of astragaloside Ⅳ between the top grade of Shanxi wild-simulated Astragali Radix top and the first grade of Gansu cultivated Astragali Radix. The contents of astragaloside Ⅳ in the second,third and fourth grade of Shanxi wild-simulated Astragali Radix was significantly higher than those of produced from Gansu.There was a significant negative correlation between the contents of astragaloside Ⅳ and grade in Shanxi Astragali Radix. While there was no correlation for Gansu Astragali Radix. This study provided the basis for the quality grade standard of Astragali Radix.

Optimization and Validation of a Microscale In vitro Method to Assess α-Glucosidase Inhibition Activity.

BACKGROUND: Microscale in vitro assays are fast, simple, and inexpensive, with reduced reagent quantities, waste, and experimental animal use. However, they have low reproducibility and low correlation with the results of in vivo models, possibly due to differences in precision and accuracy in methodologies between laboratories. OBJECTIVE: The objective was the optimization and validation of an in vitro assay, carried out on microscale, to assess the inhibition of α-glucosidase activity, which is indicative of antihyperglycemic activity. METHODS: The optimization was carried out using a fractional factorial design taking into account the best inhibition percentage and the absorbance of the controls. With the optimized experimental conditions in hand, we carried out method validation. RESULTS: The optimized conditions were as follows: enzyme concentration, 0.55 U/mL; substrate concentration, 111.5 µM; and 17.5 min incubation at 37°C. A linear range between 100 and 310.2 µg/mL of acarbose (r2 0.994) was established. The RSD was <2% and the % error was <3%. The Z factor was >0.96. This method was applied to four plant extracts, one of which was found to be very active. CONCLUSION: The method was found to be accurate, precise, selective, linear, and reliable in evaluating the antihyperglycemic activity of natural extracts in vitro.

Schinus terebinthifolius scale-up countercurrent chromatography (Part I): High performance countercurrent chromatography fractionation of triterpene acids with off-line detection using atmospheric pressure chemical ionization mass spectrometry.

'Countercurrent chromatography' (CCC) is an ideal technique for the recovery, purification and isolation of bioactive natural products, due to the liquid nature of the stationary phase, process predictability and the possibility of scale-up from analytical to preparative scale. In this work, a method developed for the fractionation of Schinus terebinthifolius Raddi berries dichloromethane extract was thoroughly optimized to achieve maximal throughput with minimal solvent and time consumption per gram of processed crude extract, using analytical, semi-preparative and preparative 'high performance countercurrent chromatography' (HPCCC) instruments. The method using the biphasic solvent system composed of n-heptane-ethyl acetate-methanol-water (6:1:6:1, v/v/v/v) was volumetrically scaled up to increase sample throughput up to 120 times, while maintaining separation efficiency and time. As a fast and specific detection alternative, the fractions collected from the CCC-separations were injected to an 'atmospheric pressure chemical ionization mass-spectrometer' (APCI-MS/MS) and reconstituted molecular weight MS-chromatograms of the APCI-ionizable compounds from S. terebinthifolius were obtained. This procedure led to the direct isolation of tirucallane type triterpenes such as masticadienonic and 3ß-masticadienolic acids. Also oleanonic and moronic acids have been identified for the first time in the species. In summary, this approach can be used for other CCC scale-up processes, enabling MS-target-guided isolation procedures.

Elemental composition of edible nuts: fast optimization and validation procedure of an ICP-OES method.

BACKGROUND: An inductively coupled plasma-optical emission spectrometry method for the speedy simultaneous detection of 19 elements in edible nuts (walnuts: Juglans nigra; almonds: Prunus dulcis; hazelnuts: Corylus avellana; Brazil nuts: Bertholletia excelsa; cashews: Anacardium occidentalle; pistachios: Pistacia vera; and peanuts: Arachis hypogaea) available on the Serbian markets, was optimized and validated through the selection of instrumental parameters and analytical lines free from spectral interference and with the lowest matrix effects. RESULTS: The analysed macro-elements were present in the following descending order: Na > Mg > Ca > K. Of all the trace elements, the tested samples showed the highest content of Fe. The micro-element Se was detected in all the samples of nuts. The toxic elements As, Cd and Pb were either not detected or the contents were below the limit of detection. One-way analysis of variance, Student's t-test, Tukey's HSD post hoc test and hierarchical agglomerative cluster analysis were applied in the statistical analysis of the results. CONCLUSION: Based on the detected content of analysed elements it can be concluded that nuts may be a good additional source of minerals as micronutrients.

Doehlert design-desirability function multi-criteria optimal separation of 17 phenolic compounds from extra-virgin olive oil by capillary zone electrophoresis.

In Brazil, the consumption of extra-virgin olive oil (EVOO) is increasing annually, but there are no experimental studies concerning the phenolic compound contents of commercial EVOO. The aim of this work was to optimise the separation of 17 phenolic compounds already detected in EVOO. A Doehlert matrix experimental design was used, evaluating the effects of pH and electrolyte concentration. Resolution, runtime and migration time relative standard deviation values were evaluated. Derringer's desirability function was used to simultaneously optimise all 37 responses. The 17 peaks were separated in 19min using a fused-silica capillary (50µm internal diameter, 72cm of effective length) with an extended light path and 101.3mmolL(-1) of boric acid electrolyte (pH 9.15, 30kV). The method was validated and applied to 15 EVOO samples found in Brazilian supermarkets.