Chromatographic Fingerprint Research Articles

Comprehensive Identification of Chemical Fingerprint and Screening of Potential Quality Markers of Aloe vera (L.) Burm. f. from Different Geographical Origins via Ultra-High-Performance Liquid Chromatography Hyphenated with Quadrupole–Orbitrap-High-Resolution Mass Spectrometry Combined with Chemometrics

An integrated strategy was developed for the systematic chemical fingerprint and chemometrics analysis for the quality assessment of Aloe vera (L.) Burm. f. The ultra-performance liquid chromatography fingerprint was established, and all common peaks were tentatively identified by using ultra-high-performance liquid chromatography hyphenated with quadrupole-orbitrap-high-resolution mass spectrometry. Afterwards, the datasets of common peaks were subjected to hierarchical cluster analysis, principal component analysis and partial least squares discriminant analysis to holistically compare the differences. The results revealed that the samples were predicted to fall into four clusters, which were related to four different geographical locations. Using the proposed strategy, aloesin, aloin A, aloin B, aloeresin D and 7-O-methylaloeresin A were rapidly determined to be the potential characteristic quality markers. Finally, five screened compounds in 20 batches of samples were simultaneously quantified, and their total contents were ranked as follows: Sichuan province > Hainan province > Guangdong province > Guangxi province, which suggests that geographical origins may be an important factor affecting the quality of A. vera (L.) Burm. f. This new strategy can not only be used to explore possibly the latent active substance candidates for pharmacodynamic studies, but it is also an efficient analytical strategy for other complex traditional Chinese medicine systems.

Screening Antibacterial Constituents of Scutellaria Radix Based on Spectrum-Effect Relationships Between HPLC Fingerprints and the Inhibition of Oral Bacteria.

Scutellaria Radix (SR) is a widely used traditional Chinese medicine in clinics for the therapy of upper respiratory tract infectious diseases. Modern pharmacological investigations indicate that SR exerts a significant bacteriostatic effect on different oral bacteria, but few studies have systematically investigated the main active constituents of SR causing this activity. Spectrum-effect correlation analysis was applied to screening anti-oral-microbial constituents from SR. The aqueous extract of SR was divided into fractions of different polarity and the active fraction was screened using the agar diffusion method. Eighteen batches of SR were further prepared and the chromatography fingerprint was established using high-performance liquid chromatography. The antibacterial activities of these constituents were examined against different oral bacteria. Finally, the spectrum-effect relationship between the fingerprint and those antibacterial effects was analyzed by gray correlation analysis and partial least squares regression. Five active constituents were screened out and their antibacterial activity was systematically confirmed by a knockout/in strategy combined with a biofilm extraction method, which indicated that these five compounds were responsible for the antibacterial activity of SR. These results form the basis for further development and improved quality control of SR in the treatment of oral diseases.

Pattern Recognition of GC-FID Profiles of Volatile Compounds in Brandy de Jerez Using a Chemometric Approach Based on Their Instrumental Fingerprints

Brandy de Jerez is a unique spirit produced in Southern Spain under Protected Geographical Indication “Brandy de Jerez” (PGI). Two key factors for the production of quality brandies are the original wine spirit and its aging process. They are significantly conditioned by specific variables related to the base wine and the distillation method employed to produce the wine spirit used to obtain a finally aged brandy. This final beverage is therefore strongly influenced by its production process. The chromatographic instrumental fingerprints (obtained by GC FID) of the major volatile fraction of a series of brandies have been examined by applying a chemometric approach based on unsupervised (hierarchical cluster analysis and principal component analysis) and supervised pattern recognition tools (partial least squares–discriminant analysis and support vector machine). This approach was able to identify the fermentation conditions of the original wine, the distillation method used to produce the wine spirit, and the aging process as the most influential factors on the volatile profile.

A strategy for antioxidant quality evaluation of Aster yunnanensis based on fingerprint-activity relationship modeling and chemometric analysis

In this study, a fingerprint-activity relationship between ultrahigh performance liquid chromatography (UHPLC) fingerprints and antioxidant activity was established to evaluate the quality of Aster yunnanensis Franch.(AYF) from different collecting spots. First, the fingerprint of AYF was established by UHPLC, and the similarity analysis was analyzed based on twenty-one common peaks. Then the chemical constituents from AYF were analyzed and identified by UHPLC-quadrupole time-of-flight tandem mass spectrometry (QTOF-MS/MS). Next, the antioxidant activity of twelve batches of AYF was assessed in vitro. Subsequently, eleven chemical markers were screened out by fingerprints and antioxidant activity utilizing grey relational analysis (GRA) and partial least squares (PLS). Finally, the contents of eleven chemical markers in twelve batches of AYF were detected by UHPLC, and the antioxidant quality of AYF was evaluated using chemometric analysis, such as principal components analysis (PCA) and technique for order preference by similarity to ideal solution (TOPSIS). The results showed that the antioxidant efficacy was associated with the total content of eleven compounds of AYF. Moreover, this method to discover quality markers was reasonable by fingerprint-activity relationship combination with chemometric analysis. The present study will certify quality markers associated with therapeutic effects, and provide a powerful strategy for evaluating the resource quality of AYF.

Effective components and mechanism of Qijiao Shengbai Capsules based on fingerprinting and network pharmacology

Qijiao Shengbai Capsules(QJ) can invigorate Qi and replenish the blood, which is commonly used clinically for adjuvant treatment of cancer and leukopenia due to chemoradiotherapy. However, the pharmacological mechanism of QJ is still unclear. This work aims to combine the high-performance liquid chromatography(HPLC) fingerprints and network pharmacology to clarify the effective components and mechanism of QJ. The HPLC fingerprints of 20 batches of QJ were established. The similarity evaluation among 20 batches of QJ was performed by using Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine(version 2012), resulting in a similarity greater than 0.97. Eleven common peaks were identified by reference standard, including ferulic acid, calycosin 7-O-glucoside, ononin, calycosin, epimedin A, epimedin B, epimedin C, icariin, formononetin, baohuoside I, and Z-ligustilide. The "component-target-pathway" network was constructed by network pharmacy, and 10 key components in QJ were identified, such as ferulic acid, calycosin 7-O-glucoside, ononin, and calycosin. The components were involved in the phosphoinositide 3 kinase-protein kinase B(PI3K-Akt), mitogen-activated protein kinase(MAPK), and other signaling pathways by regulating potential targets, including EGFR, RAF1, PIK3R1, and RELA, to auxiliarily treat tumors, cancers, and leukopenia. The molecular docking conducted on the AutoDock Vina platform confirmed the high binding activity of 10 key effective components with core targets, with the binding energy less than-5 kcal·mol~(-1). In this study, the effective components and mechanism of QJ have been preliminary revealed based on HPLC fingerprint and network pharmacology, which provided a basis for quality control of QJ and a refe-rence for further study on its mechanism.

Bioengineered dual fluorescent carbon nano dots from Indian long pepper leaves for multifaceted environmental and health utilities.

In this article, we present the synthesis of Piper longum leaves-derived ethanolic carbon dots (PLECDs) using the most simplistic environmentally friendly solvothermal carbonization method. The PLECDs fluoresced pink color with maximum emission at 670nm at 397nm excitation. Additionally, the dried PLECDs dissolved in water showed green fluorescence with higher emission at 452nm at 370nm excitation. The UV spectra showed peaks in the UV region (271.25nm and 320.79nm) and a noticeable tail in the visible region, signifying the efficient synthesis of nano-sized carbon particles and the Mie scattering effect. Various functional groups (-OH, -N-H, -C-H, -C = C, -C-N, and -C-O) were identified using Fourier transform infrared spectroscopy (FTIR). Its nanocrystalline property was revealed by the sharp peaks in the X-ray diffraction (XRD). High-resolution transmission electron microscopy (HRTEM) photomicrograph displayed a roughly spherical structure with a mean size of 2.835nm. The energy dispersive X-ray (EDAX) and X-ray photoelectron spectroscopy (XPS) revealed the elemental abundance of C, O, and N. The high-performance thin-layer chromatography (HPTLC) fingerprint of PLECDs showed an altered pattern than its precursor (Piper longum leaves ethanolic extract or PLLEE). The PLECDs sensed Cu2+ selectively with a limit of detection (LOD) and limit of quantification (LOQ) of 0.063μM and 0.193μM, respectively. It showed excellent cytotoxicity toward MDA-MB-231 (human breast cancer), SiHa (human cervical carcinoma), and B16F10 (murine melanoma) cell lines with excellent in vitro bioimaging outcomes. It also has free radical scavenging activity. The PLECDs also showed outstanding bacterial biocompatibility, pH-dependent fluorescence stability, photostability, physicochemical stability, and thermal stability.

A Comparative Analysis of the Chloroplast Genomes of Three Lonicera Medicinal Plants.

Both Lonicerae japonicae flos and Lonicerae similis flos are important components in traditional Chinese medicine (TCM) with precious medicinal value. However, the absence of studies on their chloroplast genomes and chromatography has considerably hindered the study of their evolutionary and phylogenetic relationships. In this study, the complete chloroplast (cp) genomes of Lonicera acuminata Wall. and Lonicera similis Hemsl. were sequenced using the Illumina sequencing platform and compared with that of Lonicera japonica Thunb., which has been previously reported. Furthermore, the chromatographic fingerprints of the three plants were constructed using HPLC and the content of quality marker (Q-Marker) was calculated. The annotation results showed that the two chloroplast genomes were typical quadripartite structures with lengths of 155,330 bp (L. acuminata) and 155,207 bp (L. similis). A total of 126 different genes were annotated, containing 82 protein-coding genes, 36 tRNA genes, and 8 rRNA genes. The expansion and contraction of the inverted repeat (IR) regions suggested that the boundary regions of IR/SC were comparatively conserved in the three species, and six regions (trnH-GUG-psbA, rps2-rpoC2, rbcL-psaI, trnN-GUU-ndhF, rps15-ycf1, and infA) with nucleotide diversity values (Pi) of variable sites higher than 1% were identified. Phylogenetic relation indicated that L. similis had a closer genetic relationship with L. japonica than L. acuminata. Additionally, the chromatographic fingerprints showed that the characteristic peaks of the three medicinal plants were similar, including Neochlorogenic acid, Chlorogenic acid, 4-Dicaffeoylquinic acid, Sweroside, Secoxyloganin, Luteoloside, Isochlorogenic acid A, Isochlorogenic acid B, and Isochlorogenic acid C. The content of chlorogenic acid and total phenolic acid in L. acuminata (7.4633 ± 0.4461%, 14.8953 ± 0.0728%) and L. similis (14.1055 ± 0.2566%, 21.9782 ± 0.1331%) was much higher than that of L. japonica (3.9729 ± 0.0928%, 6.0964 ± 0.1228%), respectively. This study provides appropriate information for species identification, phylogeny, quality assessment, and rational use of three medicinal plants of the genus Lonicera.

Quality Assessment of Bee Pollen-Honey Mixtures Using Thin-Layer Chromatography in Combination with Chemometrics.

The aim of this study is to develop a rapid, effect-directed screening method for quality assessment of bee pollen-honey mixtures. The comparative antioxidant potential and phenolic content of honey, bee pollen, and the bee pollen-honey mixtures, was performed using spectrophotometry. The total phenolic content and antioxidative activity of bee pollen-honey mixtures with 20 % bee pollen share were in the range 3.03-3.11 mg GAE/g, and 6.02-6.96 mmol TE/kg, respectively, while mixtures with 30 % bee pollen share contained 3.92-4.18 mg GAE/g, and 9.69-10.11 mmol TE/kg. Chromatographic fingerprint of bee pollen-honey mixtures was performed by high-performance thin-layer chromatography with conditions developed by authors and reported for the first time. Fingerprint analysis hyphenated with chemometrics enabled authenticity assessments of honey in mixtures. Results indicate that bee pollen-honey mixtures represent a food with highly, both, nutritious characteristics and health-promoting effect.

Multi-wavelength HPLC fingerprint similarity metric for cold-hot nature identification of Chinese herbal medicines

Cold-hot nature theory is the core basic theory of the nature of Chinese herbal medicines (CHMs). It is found that the material basis of cold-hot nature is CHM ingredients. In view of this, our group proposed a scientific hypothesis that “CHMs with similar nature should have similar materialbasis”. To demonstrate this hypothesis, we developed a novel multi-wavelength high performance liquid chromatography (HPLC) fingerprint similarity metric scheme for cold-hot nature identification. We explored a multi-wavelength distance metric learning model to compute the similarity of CHM ingredients, and developed an improved k-nearest neighbor algorithm based on multi-wavelength HPLC fusion (KMHF) to predict cold-hot nature of CHMs. Firstly, multi-wavelength HPLC fingerprints were used to extract the characteristic information of CHM ingredients. Secondly, we defined the similarity of CHM ingredients as semantic relevance and fingerprint similarity. We studied a multi-wavelength distance metric to measure the similarity of CHM ingredients. The learned distance metric could discover complementary characteristics of different wavelength HPLC through an optimization algorithm. Finally, an improved multi-wavelength k-nearest neighbor algorithm KMHF was proposed to analyze the relationship between cold-hot nature and CHM ingredients. Numerous experiments were designed to test the feasibility of the proposed KMHF algorithm. Experimental results indicate that the performance of our KMHF algorithm outperforms that of the compared algorithms. Experimental results demonstrate that the hypothesis that CHMs with similar cold-hot nature have similar material basis. The KMHF model is evaluated to be feasible for nature identification.

Qualitative and quantitative method for quality control of Itea ilicifolia based on antioxidant Q-markers.

Itea ilicifolia Oliv is a folk medicine with antioxidant potential. In this study, the fingerprints of 14 batches of I. ilicifolia were established by HPLC with 17 common peaks. The similarities evaluated by Similarity Evaluation System for Chromatographic Fingerprint of Chinese Materia (version 2012) were >0.89. Ten compounds were identified with definite structures by comparing the retention time and characteristic UV spectral pattern with those of reference substances. The antioxidant capacities of 14 batches of I. ilicifolia were evaluated based on O2 ·- , DPPH and ABTS·+ radical scavenging assays in combination with ferric reducing antioxidant power assay. Via multivariate statistical analyses of gray relation analysis, bivariate correlation analysis and partial least squares regression analysis, a study on the spectrum-effect relationship was then performed to screen eight peaks as the antioxidant Q-markers of I. ilicifolia. The contents of representative antioxidant Q-markers (isoorientin, orientin, vitexin, isovitexin and iteafuranal A) in samples were accurately determined to be 0.054-0.118%, 0.034-0.080%, 0.018-0.055%, 0.031-0.091% and 0.033-0.140%, respectively. The qualitative and quantitative analytical method based on Q-markers helps to control the antioxidant quality of I. ilicifolia, which will lay the foundation to promote the rational utilization of I. ilicifolia in curing diseases related to oxidative stress.

Analysis on quality value transmitting of Qinggusan reference sample of classical prescription based on quantitative fingerprint technology

Qinggusan is the 69th prescription in the first batch of "Catalogue of Ancient Chinese Classic Formulas". In modern clinical practice， Qinggusan is mainly used to treat noninfectious fever. However， because few studies on Qinggusan reference samples and their quality value transfer are available， the development and promotion of its compound preparations are restricted. Therefore， establishing an accurate and comprehensive quality control method to clarify the critical quality attributes of Qinggusan reference samples is of great importance. In this study， 15 batches of Qinggusan reference samples were processed to determine the range of their dry extract ratios. Quantitative high-performance liquid chromatography （HPLC） fingerprint analysis was performed using a Waters Symmetry Shield RP18 column （250 mm×4.6 mm， 5 μm） with acetonitrile-0.1% （v/v） formic acid aqueous solution as the mobile phase in gradient elution mode. The flow rate was 1.0 mL/min， the column temperature was 30 ℃， and the detection wavelength was 254 nm. The HPLC fingerprints of the Qinggusan reference samples were established under these conditions to evaluate their similarity. The established method was systematically validated and found to demonstrate good precision， repeatability， and sample stability. Subsequently， characteristic peaks were identified and attributed by HPLC-quadrupole-time-of-flight-mass spectrometry （HPLC-Q-TOF-MS） analysis. MS was performed in electrospray ionization mode， the data were collected in both positive- and negative-ion modes， and the detection range was m/z 50-2000. The contents and transfer rate ranges of the index components， namely， gentiopicrin， mangiferin， picroside Ⅱ， picroside Ⅰ， and glycyrrhizic acid， were determined to analyze the quality value transfer of the samples. The results demonstrated that the dry extract rate of the 15 batches of Qinggusan reference samples ranged from 24.10% to 26.88% and that their fingerprint similarities were generally greater than 0.95. Twelve common peaks were identified by reference identification， literature comparison， and high-resolution MS analysis. Twelve compounds， including six iridoid glycosides， two flavonoids， one alkaloid， one triterpenoid saponin， and two others. Among them， L-picein， androsin， picroside Ⅳ， picroside Ⅱ and picroside Ⅰ were from Picrorhizae Rhizoma， loganin acid， swertiamarin and gentiopicrin were from Gentianae Macrophyllae Radix， neomangiferin and mangiferin were from Anemarrhenae Rhizoma， dichotomine B was from Stellariae Radix， and glycyrrhizic acid was from Glycyrrhizae Radix et Rhizoma. The five key components presented good linear relationships in their respective linear ranges， and all correlation coefficients were higher than 0.999. The relative standard deviations （RSDs） of precision， stability， and repeatability were less than 1.3%. The average recoveries varied between 95.92% and 102.5%， with RSDs less than 3.9%； these values meet the requirements defined in the 2020 edition of the Chinese Pharmacopoeia. The contents of gentiopicrin， mangiferin， picroside Ⅱ， picroside Ⅰ， and glycyrrhizic acid in the 15 batches of reference samples were in the range of 17.92-27.55， 1.83-4.42， 23.08-36.44， 8.43-15.04， and 0.94-2.39 mg/g， respectively， and their transfer rates from the decoction pieces to the reference samples were 47.91%-63.95%， 22.96%-59.39%， 60.82%-77.82%， 64.25%-99.53%， and 15.30%-39.30%， respectively. In this study， the chemical components of Qinggusan reference samples were comprehensively identified and their quality value transfer was studied through the combination of HPLC fingerprinting and MS. Clarification of the critical quality attributes of Qinggusan reference samples could provide a basis for the quality control of Qinggusan compound preparations.

A Comparison of the Stability of Refined Edible Vegetable Oils under Frying Conditions: Multivariate Fingerprinting Approach.

The stability of highly consumed vegetable refined oils after discontinuous frying of potatoes was compared. Both those vegetable oils containing additives and those that did not were considered. Vegetable oil samples were evaluated using refractive index, anisidine and peroxide values, UV absorbance and dielectric constant-based determination of the content of total polar compounds. Chemical changes caused over the frying time were monitored and multivariate modelling of the data was carried out. A new gas chromatographic-mass spectroscopy method was intended to record a fingerprint of both polar and non-polar compound fractions. Multivariate models of chromatographic fingerprints were also developed, and the results obtained from both approaches were verified to be statistically similar. In addition, multivariate modelling also allows to differentiate among vegetable oils according to oxidation performance. Indeed, it was initially observed that olive oils presented the highest natural thermo-oxidative stability compared to other seed oils, although it should be noted that these differences were not significant when regarding olive pomace oils and seed oils containing synthetic additives.

Identification of <i>Sida rhombifolia</i> from Its Related Plants Using Thin-Layer Chromatographic Analysis

Sida rhombifolia belongs to the Malvaceae family and is often used to treat gout in Indonesia. S. rhombifolia has many efficacies and contains many different chemical components. The abundance and variation of chemical content and chemical compounds in this medicinal plant are challenging factors in ensuring medicinal plants' safety and quality control. Thin-layer chromatography (TLC) fingerprint analysis derived from S. rhombifolia extract can also be used for the medicinal plant's quality control. This research aimed to develop the optimum condition for the chemical fingerprint analysis of S. rhombifolia using a TLC fingerprint analysis. A total of 11 bands were produced with optimum separation using silica gel 60 F254 TLC plate, a mixed mobile phase condition with chloroform, ethyl acetate, and methanol (6.5:2:1.5). This fingerprint analysis performed an excellent separation in the TLC plate at 366 nm with sulfuric acid as reagent derivatization. In general, the results of the analysis validation, including stability, specificity, precision, and robustness of TLC fingerprints, met the acceptance criteria. The TLC fingerprint of S. rhombifolia can be distinguished from 2 related plants with similar leaf shapes, Turnera ulmifolia L. and Hibiscus rosa-sinensis. The developed method was validated, so it could be used to control S. rhombifolia quality.

Comprehensive quality evaluation of processed Scrophulariae Radix from different regions of China using HPLC coupled with chemometrics methods.

Scrophulariae Radix (SR) has been extensively used in traditional Chinese medicine (TCM) for thousands of years. However, the processing methods and production areas of Scrophularia ningpoensis have undergone notable historic changes. Thus, their effects on the bioactive constituents of SR still need to be studied further. This study aimed to establish an objective and comprehensive method to identify the correlation of bioactive constituents of SR with variety, place of origin and processing method for evaluating their qualities. An accurate and rapid high-performance liquid chromatography-diode array detector (HPLC-DAD) method for the simultaneous determination of 11 marker components (aucubin, harpagide, 6-O-methyl-catalpol, harpagoside, verbascoside, isoverbascoside, angoroside C, cinnamic acid, l-tyrosine, l-phenylalanine, and l-tryptophan) was established to evaluate the quality of SR for the first time. In addition, the effects of different production areas and processed methods on the target compounds were studied by analysing 66 batches of SR samples with chemometrics methods, including similarity evaluation of chromatographic fingerprints of TCM, principal component analysis (PCA), and partial least squares-discriminant analysis (PLS-DA). Compared with "sweating", short-term "steaming" and "slice-drying" could largely preserve the bioactive constituents of SR. When using the model established through PLS-DA, five components were identified as the most significant variables for discrimination. Furthermore, the score plots of PCA and the similarity evaluation revealed that variety had a more notable influence on the quality of SR than the place of origin. An objective approach of HPLC fingerprint coupled with chemometrics analysis and quantitative assessment could be applied to discriminate different processed SR and evaluate the qualities of SR rapidly.

Gas Chromatography Fingerprint of Martian Amino Acids before Analysis of Return Samples

Within the perspective of the current and future space missions, the detection and separation of building blocks such as amino acids are important subjects which are becoming fundamental in the search for the origin of life and traces of life in the solar system. In this work, we have developed and optimized a strategy adapted to space experimentation to detect the presence of amino acid-like compounds using gas chromatography coupled to mass spectrometry (GC-MS). Selected derivatization methods meet the instrument design constraints imposed on in situ extraterrestrial experiments. Coupled to a fast selective extraction, GC analysis would be highly efficient for the detection of organic materials. In the future, the corresponding GC-MS TIC could facilitate simple and fast selection of sediments/dust samples onboard GC-MS-equipped rovers for sample return-to-Earth missions.

Pharmacopoeial Standards for Venpucani Ilakam - A classical Siddha medicine

Belief on the therapeutic values of herbal medicine is in increasing trend and the consumption of traditional medicine is also in upward trend. The quality control and assurance of these traditional medicines is a prime criterion for any herbal drug manufacturer. Venpucani ilakam (VPI) is a classical poly herbal Siddha medicine under the category Ilakam (Legiyam in Ayurveda). This poly herbal medicine is a combination of 18 ingredients of plants and animal origins. An attempt was made to prepare the drug with authenticated ingredients and subjected to physicochemical analysis, thin layer chromatographic photo documentation and high performance thin layer chromatographic finger printing. The TLC photo documentation of defatted chloroform extract showed nine spots under UV 254, ten spots under UV 366 nm and eleven spots after derivatization with vanillin sulphuric acid; defatted success ethanol extract showed five spots under short wavelength, six spots under long wavelength and seven spots after derivatization. HPTLC finger printing recorded eleven peaks under 254 nm, seventeen under 366 nm and fifteen peaks after derivatization in defatted chloroform extract; nine peaks under 254 nm, eleven peaks under 366 nm and eight peaks after derivatization in defatted ethanol extract. About twenty three percent loss in weight was observed on heating at 105oC and drug was acidic nature. Not less than fourty percentage of the drug was soluble in alcohol and also in water. This drug has wide therapeutic use and the derived results may serve as reference standards.

Comparative Investigation Between Raw and Stir-Frying Processed Cuscutae Semen Based on HPLC Fingerprints Coupled With Chemometric Methods.

The curative effect of Cuscutae Semen (CS) could be adjusted after stir-frying, and it is difficult to distinguish between the raw CS and stir-fried processed Cuscutae Semen (SFCS) in clinical practice. The paper aims to establish the fingerprints of raw and fried Cuscutae Semen products by high-performance liquid chromatography (HPLC) so that the compounds in raw and stir-fried products could be analyzed qualitatively and quantitatively. At the same time, the chemometrics method was used to evaluate the difference between CS and SFCS to provide reference for the research of traditional Chinese medicine (TCM) CS. The raw and SFCS products' related substances were separated on a C18 column (250 mm × 4.6 mm, 5 µm) maintained at 30 °C (column temperature). The mobile phase comprised 0.1% formic acid aqueous solution (A) and acetonitrile (B), and a detection wavelength of 328 nm. The data were imported into statistical software for chemometric analysis. The developed HPLC method exhibits good linearity and has good systematic applicability. The content of these 12 compounds in the samples was further determined and the data analyzed by chemometrics. The results showed that the composition of CS changed on heating, and HCA showed that CS and SFCS could be clearly distinguished. PCA showed that six components caused differences, namely neochlorogenic acid, cryptophyllogenic acid, caffeic acid, quercetin, isorhamnetin, and kaempferol. This study unequivocally establishes a chromatographic fingerprint method intended for the extensive analysis of raw and stir-fried processed CS, which could substantially enhance the quality control of CS and the rational development and utilization of TCM resources. This method for the simultaneous quantification of multiple compounds in CS and SFCS revealed the components responsible for the differences between raw and processed products. This will provide support for quality control of this herbal medicine.

Liquid Chromatography Fingerprint Analysis of Released Compounds in Plasma Samples of Stroke Patients after Thrombolytic Treatment

Plasma samples obtained from stroke patients treated with recombinant tissue-type plasminogen activator (rt-PA) and not treated with rt-PA were evaluated with different HPLC methodologies to obtain information about the possible release of small molecules as a result of the thrombolytic treatment. Plasma samples, without derivatization and derivatized with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC), were evaluated with a HPLC gradient method, which consisted of a mobile phase of 10 mM ammonium acetate buffered solution (pH = 5.3) and acetonitrile. Three different detection methods were applied: UV, fluorescence, and ESI-MS. The results obtained showed that a group of new highly hydrophilic compounds appeared in most samples analyzed from treated patients, just after the administration of rt-PA. These compounds appeared shortly after the administration of the drug and were detected during the first 24 h after treatment, disappearing from plasma after this time. These new compounds were not detected either in controls or in non-treated stroke patients, which suggests that they were released into the plasma as a consequence of the thrombolytic effect of the drug. Our results suggest that these new compounds might be free glycans. The use of AQC as a derivatizing reagent has demonstrated that the new compounds detected cannot contain primary or secondary amine groups in their structure. The molecular mass determined by ESI-MS (821 Da) suggests that if these compounds are free glycans they might be a high-mannose type.

Screening the Synergistic Components of Acetylcholinesterase Inhibition from Phellodendron Bark Based on Fingerprint-Activity Relationship Modeling

Objective: Phellodendron chinense (PC) bark and Phellodendron amurense (PA) bark are two herbal medicines recorded in the Chinese Pharmacopoeia (2020 edition) that are easily mistaken for one another. In this study, the chemical constituents of PC and PA were compared using chromatographic fingerprints, and the potential synergistic acetylcholinesterase (AChE) inhibitor components were screened based on the correlation of fingerprint activity. Materials and Methods: Chromatographic fingerprints based on high-performance liquid chromatography were developed for the analysis and comparison of chemical compounds in PC and PA samples. The AChE inhibitory activity of PC and PA was determined using the Ellman method. Subsequently, the contribution of the characterized alkaloids in PC and PA to the overall AChE inhibition was modeled using partial least squares regression (PLSR). Results: The total alkaloid content in PC was higher than that in PA, which causes PC to have stronger anti-AChE activity. Overall, 13 and 20 common peaks were identified in the PC and PA samples, respectively. Among them, berberine (BER) was the dominant alkaloid in PC, which covered more than 65% of the total peak area in PC, but only approximately 25% of that in PA, indicating that the chemical composition is different between PC and PA. The spectrum–effect analysis based on PLSR and the correlation analysis showed that the BER-palmatine (PAL) and BER-jatrorrhizine (JAT) pairs have a synergistic inhibitory effect on AChE activity. Conclusions: A high-performance chromatographic fingerprint was established to distinguish PC and PA. The efficacy-associated markers were screened, including the pairs of BER-PAL and BER-JAT with anti-AChE activity, and the findings may assist with the quality control of PA and PC.

Chemometric Classification of Mangifera indica L. Leaf Cultivars, Based on Selected Phytochemical Parameters; Implications for Standardization of the Pharmaceutical Raw Materials.

Mangifera indica leaves are among the most common materials employed in manufacturing herbal medicinal products. Despite the phytochemical variation of M. indica cultivars, there are no monographs to guide the cultivation, processing, and authentication of the materials. This study characterized 15 Ugandan M. indica leaf varieties, with reference to extraction index (EI), total phenolic content (TPC), antioxidant activity (AOA), and mangiferin concentration (MC). In addition, HPLC fingerprints were established to evaluate the overall phytoequivalence of the materials. Then, using hierarchical clustering (HC) and principal component analysis (PCA), the materials were assigned quality grades. The mean EI was 9.39 ± 1.64% and varied among the varieties (P=0.001); the TPC varied significantly (P < 0.0001), from 183.29 ± 2.36 mg/g (Takataka) to 79.47 ± 0.58 mg/g (Apple mango). AOA ranged from 16.81 ± 2.85 μg/mL (Doodo red) to 87.85 μg/mL (Asante). MC varied significantly (P < 0.0001), from 105.75 ± 0.60 mg/g (Kate) to 39.53 ± 0.30 mg/g (Asante). HC gave four major grades: A to D (A, varieties with the highest TPC, MC, and AOA). These parameters reduced to below average from group B to group D. The chromatographic fingerprints were visually similar, but the number of peaks varied, from 19 (Kawanda green) to 29 (Kawanda wide), with 23.5 ± 2.9 average peaks. Whole fingerprints were less similar (r < 0.8) than common peak fingerprints (r > 0.9, P < 0.001). PCA grouped the fingerprints into five clusters; loading plots for PC 1 and 2 revealed two important compounds, one at Rt = 15.828 minutes (mangiferin) and the other at 6.021 minutes. Using the standardized common fingerprints, unknown field samples clustered closely with Koona, Kate, and Kawanda green varieties. The EI, TPC, MC, and AOA values can be utilized to monitor consistency in the quality of materials and the production process. The grades generated can be used to select materials for cultivation and manufacturing. Where minimum concentrations are set, materials of different concentrations are used to dilute or concentrate each other. The HPLC fingerprints can be utilized to authenticate the materials. More samples from different agroecological regions of the country should be tested to cater to climatic variations in order to develop GMP-compliant botanical identification methods.