Self-assembly variation of glycyrrhetinic acid epimers: Assembly mechanism and antibacterial efficacy between 18 α-GA and 18 ß-GA.

Studies have confirmed that the spatial arrangement of chiral molecules has a decisive influence on supramolecular assembly. However, the effect of epimerization caused by the change of a single chiral center on the self-assembly of chiral molecules has not been reported. Herein, we explored a pair of epimers 18 α-glycyrrhetinic acid and 18 ß-glycyrrhetinic acid from Glycyrrhiza uralensis Fisch, which had completely different medicinal activities. In this study, we found that these epimers of glycyrrhetinic acid showed distinct self-assembly properties under the condition of the deionized water and a small amount of DMSO solution. Interestingly, the cis-configured 18 ß-GA could form a 'head-to-tail' interlaced structure and further self-assembled to form nanoparticles, while trans-configured 18 α-GA was connected in a "head-to-head" manner, and due to the excessive steric hindrance that made it difficult to assemble. This work not only clearly demonstrates the impact of epimerization due to changes in a single chiral center on the self-assembly of chiral molecules as well as biological activity, but also provides new insights into the self-assembly of natural organic molecules in the development of nanomedicines and biofunctional materials.

Improved Mobility-Stretchability Properties of Diketopyrrolopyrrole-Based Conjugated Polymers with Diastereomeric Conjugation Break Spacers.

Stretchable conjugated polymers with conjugation break spacers (CBSs) synthesized via random terpolymerization have gained considerable attention because of their efficacy in modulating mobility and stretchability. This study incorporates a series of dianhydrohexitol diastereomers of isosorbide (ISB) and isomannide (IMN) units into the diketopyrrolopyrrole-based backbone as CBSs. It is found that the distorted CBS (IMN) improves the mobility-stretchability properties of the polymer with a highly coplanar backbone, whereas the extended CBS (ISB) enhances those of the polymer with a noncoplanar backbone. Additionally, the different configurations of ISB and IMN sufficiently affect the solid-state packing, aggregation capabilities, crystallographic parameters, and mobility-stretchability properties of the polymer. The IMN-based polymers exhibit the highest mobility of 1.69 cm2 V-1 s-1 and crystallinity retentions of (85.7, 78.6)% under 20% and 60% strains, outperforming their ISB-based or unmodified counterparts. The improvement is correlated with a robust aggregation capability. Furthermore, the CBS content affects aggregation behavior, notably affecting mobility. This result indicates that incorporating CBSs into the polymer can enhance backbone flexibility via movement and rotation of the CBS without affecting the crystalline regions.

Enhancement of selectivity, 25-hydroxyvitamin D3 level, alkaline phosphatase activity and reproductive performance in gilts and primiparous sows using 14-epimer of 25-hydroxyvitamin D3.

Selecting breed-worthy gilts as sow replacements is essential for continuity of pig production cycle. Though vitamin D3 (VD3) is known to enhance reproductive performance of multiparous sows, there is still a knowledge gap on its impact in developing gilts and primiparous sows. This study was aimed to quantify plasma 25-hydroxyvitamin D3 (25(OH)D3), serum alkaline phosphatase (ALP), and examine the reproductive performance of primiparous sows fed diets supplemented with regular VD3, and its 25(OH)D3 epimers. The study sample comprised 10-week-old replacement gilts (50 % Landrace x 50 % Yorkshire, N = 180) assigned in a randomized complete block design to three treatments [2,000 IU/kg of VD3 (T1), 25 µg/kg of 14­epi-25(OH)D3, half dose (T2), and 50 µg/kg of 25(OH)D3 (T3)] equilibrated to 2,000 IU/kg in base diets. Selections occurred at 22, 27 and 35 weeks of age, respectively. Plasma 25(OH)D3, serum alkaline phosphatase (ALP), bone structure and reproductive performance were analyzed. Dietary treatments influenced carpus (P = 0.023), fore view stance (P = 0.017), infantile vulva (P = 0.014), inverted (P = 0.048), and prominent teat (P < 0.001). Post-partum 25(OH)D3 concentration and ALP activity were elevated by day 25 (P < 0.001). Treatment diets also influenced total born (P < 0.001), born alive (P = 0.048), and still born (P = 0.049). Two factors affect circulating 25(OH)D3 and ALP activity: physiological changes in sows during lactation, and dietary 25(OH)D3 intake. 14­epi-25(OH)D3 is a potent metabolite for improving maturation of reproductive organs in developing gilts. It also reduces still birth in primiparous sows.

Biotransformation of docosahexaenoic acid into 10R,17S-dihydroxydocosahexaenoic acid as protectin DX 10-epimer by serial reactions of arachidonate 8R- and 15S-lipoxygenases.

Protectins, 10,17-dihydroxydocosahexaenoic acids (10,17-DiHDHAs), are belonged to specialized pro-resolving mediators (SPMs). Protectins are generated by polymorphonuclear leukocytes in humans and resolve inflammation and infection in trace amounts. However, the quantitative production of protectin DX 10-epimer (10-epi-PDX, 10R,17S-4Z,7Z,11E,13Z,15E,19Z-DiHDHA) has been not attempted to date. In this study, 10-epi-PDX was quantitatively produced from docosahexaenoic acid (DHA) by serial whole-cell biotransformation of Escherichia coli expressing arachidonate (ARA) 8R-lipoxygenase (8R-LOX) from the coral Plexaura homomalla and E. coli expressing ARA 15S-LOX from the bacterium Archangium violaceum. The optimal bioconversion conditions to produce 10R-hydroxydocosahexaenoic acid (10R-HDHA) and 10-epi-PDX were pH 8.0, 30 °C, 2.0 mM DHA, and 4.0 g/L cells; and pH 8.5, 20 °C, 1.4 mM 10R-HDHA, and 1.0 g/L cells, respectively. Under these optimized conditions, 2.0 mM (657 mg/L) DHA was converted into 1.2 mM (433 mg/L) 10-epi-PDX via 1.4 mM (482 mg/L) 10R-HDHA by the serial whole-cell biotransformation within 90 min, with a molar conversion of 60% and volumetric productivity of 0.8 mM/h (288 mg/L/h). To the best of our knowledge, this is the first quantitative production of 10-epi-PDX. Our results contribute to the efficient biocatalytic synthesis of SPMs.

Can the heptapeptide ASSIVSF of the ß2-adrenoceptor recognize ephedrine and pseudoephedrine epimers in a complex system?

Epimer separation is crucial in the field of analytical chemistry, separation science, and the pharmaceutical industry. No reported methods could separate simultaneously epimers or even isomers and remove other unwanted, co-existing, interfering substances from complex systems like herbal extracts. Herein, we prepared a heptapeptide-modified stationary phase for the separation of 1R,2S-(-)-ephedrine [(-)-Ephe] and 1S,2S-(+)-pseudoephedrine [(+)-Pse] epimers from Ephedra sinica Stapf extract and blood samples. The heptapeptide stationary phase was comprehensively characterized by scanning electron microscopy, X-ray photoelectron spectroscopy, and Fourier transform infrared spectroscopy. The separation efficiency of the heptapeptide column was compared with an affinity column packed with full-length ß2-AR functionalized silica gel (ß2-AR column). The binding affinity of the heptapeptide with (+)-Pse was 3-fold greater than that with (-)-Ephe. Their binding mechanisms were extensively characterized by chromatographic analysis, ultraviolet spectra, circular dichroism analysis, isothermal titration calorimetry, and molecule docking. An enhanced hydrogen bonding was clearly observed in the heptapeptide-(+)-Pse complex. Such results demonstrated that the heptapeptide can recognize (+)-Pse and (-)-Ephe epimers in a complex system. This work, we believe, was the first report to simultaneously separate epimers and remove non-specific interfering substances from complex samples. The method was potentially applicable to more challenging sample separation, such as chiral separation from complex systems.

Localizing Isomerized Residue Sites in Peptides with Tandem Mass Spectrometry.

Isomerized amino acid residues have been identified in many peptides extracted from tissues or excretions of humans and animals. These isomerized residues can play key roles by affecting biological activity or by exerting an influence on the process of aging. Isomerization occurs spontaneously and does not result in a mass shift. Thus, identifying and localizing isomerized residues in biological samples is challenging. Herein, we introduce a fast and efficient method using tandem mass spectrometry (MS) to locate isomerized residues in peptides. Although MS2 spectra are useful for identifying peptides that contain an isomerized residue, they cannot reliably localize isomerization sites. We show that this limitation can be overcome by utilizing MS3 experiments to further evaluate each fragment ion from the MS2 stage. Comparison at the MS3 level, utilizing statistical analyses, reveals which MS2 fragments differ between samples and, therefore, must contain the isomerized sites. The approach is similar to previous work relying on ion mobility to discriminate MS2 product ions by collision cross-section. The MS3 approach can be implemented using either ion-trap or beam-type collisional activation and is compatible with the quantification of isomer mixtures when coupled to a calibration curve. The method can also be implemented in combination with liquid chromatography in a targeted approach. Enabling the identification and localization of isomerized residues in peptides with an MS-only methodology will expand accessibility to this important information.

Development and validation of LC-MS/MS for quantifying omadacycline from stool for gut microbiome studies.

This study developed and validated a new liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify omadacycline and its epimerization in stool to facilitate microbiome studies. Omadacycline was extracted in a methanol-water-ethylenediaminetetraacetic acid (ETDA) solvent containing deuterated omadacycline as internal standard, followed by dilution. In an optimal gradient elution mode, omadacycline and its C4 epimer were separated within 5 min on reversed-phase C18 column. The method showed a broad working range of 0.1-200 ng/ml with a limitation of detection (LOD) of 0.03 ng/ml, little fecal matrix effect, good intra-day and inter-day accuracy (90-101 %), precision (2-15 %), and recovery rate (99-105 %). The method was sufficiently sensitive to quantify omadacycline in human fecal samples (n = 82) collected during a 10-day therapy course and at follow-up (day 13 and day 30) that ranged from 1 to 4785 µg/g. Further analysis revealed that ∼9 % of omadacycline was epimerized in fecal matrix control while, on average, 37.4 % was epimerized in human fecal samples. This study developed and validated a novel, simple, sensitive, and accurate method utilizing LC-MS/MS to quantify omadacycline its epimerization in the human gut. This has important implications for future studies of omadacycline and other tetracycline-class antibiotics as part of gut microbiome studies.

Differential antiangiogenic and anticancer activities of the active metabolites of ginsenoside Rg3.

Background: Epimers of ginsenoside Rg3 (Rg3) have a low bioavailability and are prone to deglycosylation, which produces epimers of ginsenoside Rh2 (S-Rh2 and R-Rh2) and protopanaxadiol (S-PPD and R-PPD). The aim of this study was to compare the efficacy and potency of these molecules as anti-cancer agents. Methods: Crystal violet staining was used to study the anti-proliferatory action of the molecules on a human epithelial breast cancer cell line, MDA-MB-231, and human umbilical vein endothelial cells (HUVEC) and compare their potency. Cell death and cell cycle were studied using flow cytometry and mode of cell death was studied using live cell imaging. Anti-angiogenic effects of the drug were studied using loop formation assay. Molecular docking showed the interaction of these molecules with vascular endothelial growth factor receptor-2 (VEGFR2) and aquaporin (AQP) water channels. VEGF bioassay was used to study the interaction of Rh2 with VEGFR2, in vitro. Results: HUVEC was the more sensitive cell line to the anti-proliferative effects of S-Rh2, S-PPD and R-PPD. The molecules induced necroptosis/necrosis in MDA-MB-231 and apoptosis in HUVEC. S-Rh2 was the most potent inhibitor of loop formation. In silico molecular docking predicted a good binding score between Rh2 or PPD and the ATP-binding pocket of VEGFR2. VEGF bioassay showed that Rh2 was an allosteric modulator of VEGFR2. In addition, SRh2 and PPD had good binding scores with AQP1 and AQP5, both of which play roles in cell migration and proliferation. Conclusion: The combination of these molecules might be responsible for the anti-cancer effects observed by Rg3.

An Overview of Different Vitamin D Compounds in the Setting of Adiposity.

A large body of research shows an association between higher body weight and low vitamin D status, as assessed using serum 25-hydroxyvitamin D concentrations. Vitamin D can be metabolised in adipose tissue and has been reported to influence gene expression and modulate inflammation and adipose tissue metabolism in vitro. However, the exact metabolism of vitamin D in adipose tissue is currently unknown. White adipose tissue expresses the vitamin D receptor and hydroxylase enzymes, substantially involved in vitamin D metabolism and efficacy. The distribution and concentrations of the generated vitamin D compounds in adipose tissue, however, are largely unknown. Closing this knowledge gap could help to understand whether the different vitamin D compounds have specific health effects in the setting of adiposity. This review summarises the current evidence for a role of vitamin D in adipose tissue and discusses options to accurately measure vitamin D compounds in adipose tissue using liquid chromatography tandem mass spectrometry (LC/MS-MS).

[Applying Serum Vitamin D Metabolites in the Assessment of Renal Impairment in Diabetic Kidney Disease of Type 2 Diabetes Mellitus Patients: A Retrospective Study].

Objective: Total 25(OH)D (t-25[OH]D), a marker traditionally used in the assessment of vitamin D (VitD) in the human body, includes 25(OH)D 2, 25(OH)D 3, and C 3-epimers-25(OH)D 3(C 3-epi). In this study, we analyzed the relationship between serum VitD metabolites and renal impairment in patients with diabetic kidney disease (DKD). Methods: We covered, in the study, 339 subjects, including 114 otherwise healthy controls (HC), 74 type 2 diabetes mellitus (DM) patients with no glomerular filtration dysfunction, and 151 DKD patients. According to the results of combined evaluation of estimated glomerular filtration rate (eGFR) and urine albumin-to-creatinine ratio (UACR), the DKD patients were further divided into four subgroups, stage 2 subgroup of patients of DM combined with stage-2 chronic kidney disease (CKD2), stage 3 subgroup of patients of DM combined with CKD3, stage 4 subgroup of patients of DM combined with CKD4, and stage 5 subgroup of patients of DM combined with CKD5. The levels of 25(OH)D 2, 25(OH)D 3, and C 3-epi were measured by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), and the activity level of 25(OH) 3 (AVitD 3), t-25(OH)D concentration, 25(OH)D 2/25(OH)D 3 ratio, C 3-epi/t-25(OH)D ratio, and C 3-epi/AVitD 3 ratio were calculated. Results: The levels of 25(OH)D 3, t-25(OH)D, and AVitD 3 were lower in the DKD group than those in the DM and HC groups (all P<0.05). C 3-epi/t-25(OH)D ratio and C 3-epi/AVitD 3 ratio were higher in the DKD group than those in the HC group (all P<0.05). The levels of 25(OH)D 3, t-25(OH)D, AVitD 3, and C 3-epi were lower in the stage 5 subgroup than those in the stage 2 and stage 3 subgroups (all P<0.05). The levels of 25(OH)D 3, t-25(OH)D, and C 3-epi were lower in the stage 4 subgroup than those in the stage 3 subgroup (all P<0.05). The 25(OH)D 3, t-25(OH)D, and AVitD 3 levels were lower in the stage 4 subgroup than those in the stage 2 subgroup (all P<0.05). Conclusions: UPLC-MS/MS can be used to perform accurate evaluation of VitD nutritional status in DKD patients. DKD patients have decreased levels of serum t-25(OH)D, 25(OH)D 3, and AVitD 3, all of which progressively decrease along with the rise in CKD staging. The trend of C 3-epi and 25(OH)D 3 changes were not consistent.

The Impact of Storage Temperature and Time on Ergot Alkaloid Concentrations.

Ergot sclerotia produce toxic secondary metabolites, ergot alkaloids, that infect cereal crops and grasses. Ergot alkaloids have two isomeric configurations: the C-8-R-isomer (R-epimer), and the C-8-S-isomer (S-epimer). Ergot contaminated matrices, such as cereal grains or grasses, may be stored for extended periods at various temperatures before being analyzed, utilized, or consumed. This study assessed the concentration of six common ergot alkaloids in both configurations found in naturally contaminated wheat over time (one, two, and four months) at different temperatures (room temperature, +4 °C, and -20 °C) using ultra-high-performance liquid chromatography-tandem mass spectrometry. The data indicate that the total ergot concentration within a natural contaminated sample varies over time at room temperature, +4 °C, and -20 °C. The total ergot concentration increased until month two, and decreased at month four, independent of temperature (p < 0.05). The total R-epimer concentration appeared to be less stable over time than the total S-epimer concentration. The changes in the total R and total S-epimer concentrations may have been caused by changes in the ergocristine and ergocristinine concentrations, respectively. Time and temperature should be considered when storing potentially contaminated matrices in a laboratory or practical agriculture situations. Quantification of ergot contaminated matrices should occur prior to their use to ensure the most reliable estimates of the concentration of ergot.

Stereochemistry and antimalarial activity of C-10 carba analogues of artemisinin.

Artemisinin is an endoperoxide bond-containing sesquiterpene lactone showing potent antimalarial effect as well as antitumor and antivirus activities. Inspired by this unique pharmacorphore, researchers around the world developed numerous Artemisinin derivatives. Among these derivatives, the C-10 carba analogues of artemisinin are frequently reported. However, the stereochemistry of C-10 carba analogues of artemisinin is overlooked and the corresponding mixture of stereoisomers are used. Herein, we reported for the first time stereochemistry and antimalarial activity of C-10 carba analogues of artemisinin. We employed two approaches to obtain the pure isomer of C-10 carba analogues and presented an interesting observation about their antimalarial activities. The minor isomer with large-sized substitute and S configuration at C-10 position had much lower antimalarial effect than the major isomer with R configuration. The study will shed light on the development of effective antimalarial drugs based on ART.

Improved quantitative LC-MS/MS analysis of vitamin D metabolites in serum after one-pot double derivatization.

In this study, we report a one-pot double derivatization scheme, which used acetylation after a Diels-Alder reaction using 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) to improve separation efficiency and provide baseline separations of the five vitamin D metabolites 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), 24,25-dihydroxyvitamin D3 (24,25(OH)2D3), 3ß-25-hydroxyvitamin D3 (3ß-25(OH)D3), 3α-25-hydroxyvitamin D3 (3α-25(OH)D3) and vitamin D3 on a C-18 stationary phase. Vitamin D metabolites are often very challenging to measure quantitatively using mass spectrometry, due to their low serum concentration levels and low ionization efficiencies. Moreover, some of these species are isomers with virtually identical mass spectral dissociation behavior. To overcome the low ionization efficiency and unspecific fragmentation behavior, derivatization using Diels-Alder reactions with Cookson-type reagents such as PTAD are common. These derivatization reactions generally result in more complicated liquid chromatography separations, because both 6R- and 6S-isomers are formed during Diels-Alder reactions. It has been shown that separations have been particularly challenging for the 3α-25(OH)D3 and 3ß-25(OH)D3 epimers. Here, we optimized the PTAD derivatization and the esterification using acetic anhydride. By utilizing the esterification catalyst 4-dimethylaminopyridine, we avoided quenching and evaporation between the two derivatization steps, but were also able to perform the esterification at room temperature without heating. The optimized one-pot double derivatization LC-MS/MS assay was validated with respect to inter/intra-day precision, accuracy, recovery and linear dynamic range and applied to metabolic fingerprinting of vitamin D3 metabolites in serum samples. The metabolites 3α-25(OH)D3, 3ß-25(OH)D3 and 24,25(OH)2D3, were readily quantified in all investigated samples. The method was, in principle, also fit for purpose for quantification of the native vitamin D3 species; the relatively high blank concentration of the commercial vitamin D-depleted serum used for calibration, however, limited the limits of quantification for this metabolite. The method provided insufficient limits of quantification for serum levels of 1,25(OH)2D3.

Thiolactones and Δ8,9-Pregnene Steroids from the Marine-Derived Fungus Meira sp. 1210CH-42 and Their α-Glucosidase Inhibitory Activity.

The fungal genus Meira was first reported in 2003 and has mostly been found on land. This is the first report of second metabolites from the marine-derived yeast-like fungus Meira sp. One new thiolactone (1), along with one revised thiolactone (2), two new Δ8,9-steroids (4, 5), and one known Δ8,9-steroid (3), were isolated from the Meira sp. 1210CH-42. Their structures were elucidated based on the comprehensive spectroscopic data analysis of 1D, 2D NMR, HR-ESIMS, ECD calculations, and the pyridine-induced deshielding effect. The structure of 5 was confirmed by oxidation of 4 to semisynthetic 5. In the α-glucosidase inhibition assay, compounds 2-4 showed potent in vitro inhibitory activity with IC50 values of 148.4, 279.7, and 86.0 µM, respectively. Compounds 2-4 exhibited superior activity as compared to acarbose (IC50 = 418.9 µM).

In vitro reversible dehydration in C3-substituents of zinc chlorophyll analogs by BchF and BchV enzymes: Stereoselectivity and substrate specificity in the dehydration.

In the biosynthetic pathway of bacteriochlorophyll(BChl)-a/b/c/d/e molecules, BchF and BchV enzymes catalyze the hydration of a C3-vinyl to C3-1-hydroxyethyl group. In this study, the in vitro reactions catalyzed by BchF and BchV partially afforded a C31-epimeric mixture of the hydrated products (secondary alcohols), with the primary recovery of the C3-vinylated substrate. The stereoselectivity and substrate specificity for the in vitro reverse enzymatic dehydration were examined using zinc chlorophyll analogs as model substrates by BchF and BchV, which were obtained from extracts of Escherichia coli overexpressing the respective genes from Chlorobaculum tepidum and used without further purification. Both BchF and BchV preferred dehydration of the (31R)-epimers over the (31S)-epimers. The (31R)-epimer was directly dehydrated by BchF and BchV to give the C3-vinylated product. By contrast, two reaction pathways for BchF and BchV dehydrations of the (31S)-epimer were proposed: (1) the (31S)-epimer would be directly dehydrated to C3-vinyl group. (2) the (31S)-epimer would be epimerized to the (31R)-epimer, and the resulting epimer was dehydrated. The results indicated that both BchF and BchV did function as a hydratase/dehydratase and could play a role in the C31-epimerization. An increase in the alkyl size at the C8-position gradually suppressed the BchF and BchV-catalyzed dehydration in vitro, while the C121- and C20-methylation only slightly affected the reaction. Using the BchF dehydration, a large amount of 3-vinyl-bacteriochlorophyllide-a was successfully prepared, with the retention of the chemically labile, central magnesium atom.

Dynamics of the vitamin D C3-epimer levels in preterm infants.

OBJECTIVES: The primary objective was to determine levels of C3-epi-25(OH)D in very low birth weight infants. The secondary objective was to evaluate the possible influence of preterm birth, intrauterine growth restriction (IUGR), and season of birth on the production of C3-epimers. METHODS: A total of 127 infants with birth weight less than 1,500 g met the inclusion criteria of the study. We examined 25-hydroxyvitamin-D [25(OH)D] levels and C3-epi-25(OH)D in maternal serum before labor, and in cord blood and infants' serum on days 14 and 28, and at discharge. RESULTS: The mean levels (±SD) of C3-epi-25(OH)D of the cord, on day 14, on day 28, and at discharge were 2.2 (2.9), 7.7 (5.5), 11.7 (7.6) and 14.9 (11.7) nmol/L respectively. The proportion of total 25(OH)D as the C3-epimer was 6.9% (cord), 16.3% (day 14), 22.4% (day 28) and 23.3% (discharge). A statistically significant correlation between 25(OH)D and C3-epi-25(OH)D can be demonstrated from birth. The severity of immaturity and IUGR did not affect the production of C3-epimers. In summer/autumn vs. winter/spring, the mean (SD) percentage of total 25(OH)D as the C3-epimer significantly differs only in maternal serum samples and umbilical cord samples (p value <0.001). CONCLUSIONS: The production of C3-epi-25(OH)D is functional even in the most immature newborns, has fetal origins, and is largely dependent on circulating 25(OH)D. At the end of the first month of life, C3-epimers make up more than 20% of 25(OH)D.

Steriodal saponins from the rhizomes of Tupistra chinensis Baker.

Chemical constituent investigation on the n-BuOH extract of the rhizomes of Tupistra chinensis Baker leads to the isolation of ten compounds including eight undescribed furostanol saponins, tupischinosides A - H, and two known ones. The structures of isolated compounds were determined by extensive spectral analysis and chemical evidences. Interestingly, tupischinosides A and B, C and D, E and F, G and H were identified as four pairs of epimers. The cytotoxicity of tupischinosides A - H against human cancer cell lines U87, SHG44, U251, LN229 and HepG-2 was evaluated by CCK-8 method. As a result, tupischinosides A and C exhibited significant proliferation inhibitory effect on the tested cancer cells. On the contrary, the corresponding epimers, tupischinosides B and D, which only differ in the configuration of C-23 didn't exhibit any cytotoxicity to cancer cells. These results indicated that the stereochemistry of C-23 was crucial to the activity of the compounds.

Chiral Separation, Configuration Confirmation and Bioactivity Determination of the Stereoisomers of Hesperidin and Narirutin in Citrus reticulata Blanco.

Hesperidin and narirutin are a class of flavanone glycosides, which are the main active constituents in Citrus reticulata Blanco. In the present study, a chiral HPLC-UV method with amylose tris(3,5-dimethylphenylcarbamate) as a stationary phase under a normal-phase mode was used to achieve the stereoselective separation of the C-2 diastereomers of hesperidin and narirutin simultaneously. The single epimer was then successfully prepared by applying semi-preparative chromatography, whose absolute configuration (R/S) was characterized by combining the experimental electronic circular dichroism (ECD) detection with time-dependent density functional theory (TDDFT) calculations. The epimer composition of these two chiral flavanone glycosides in Citrus reticulata Blanco was then determined, which was found to be slightly different in the herbs from different production regions. The anti-inflammatory activity of each prepared single epimer was further evaluated, and some differences between one pair of epimers of hesperidin and narirutin were observed, which suggested that the presence of different epimers should be considered in the quality evaluation and control of natural medicine.

Linkage and Stereochemistry Characters of Phenolic Antioxidant Product Formation.

The present study developed a smart and novel strategy to elucidate the linkage and stereochemistry characters during phenolic antioxidant product formation. A series of phenolic isomers or analogues were treated with 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide radical, to create 16 antioxidant dimerization reactions in aqueous solution. The products were rapidly identified by ultraperformance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass-spectrometry. Through a systematic function-structure relationship analysis of these reactions and theoretical calculations, it is concluded that the phenolic antioxidant product is formed via linear linkage or furanocyclic linkage. The linear linkage is fulfilled via a radical coupling and controlled by the O-O linkage exclusion, meta-linkage exclusion, and catechol-activated principles. However, when an exocyclic π-bond conjugates with the phenolic core and is affixed at the -OH para-position, the furanocyclic linkage may occur via a subsequent intramolecular Michael addition. The intramolecular addition always lacks Re-attack to show "α,ß diastereoselectivity." The α,ß diastereoselectivity is the stereochemistry character of furanocyclic linkage during phenolic antioxidant product formation. All these novel findings can benefit not only the field food science but also other fields as well.

Differentiation of tetracyclines and their 4-epimers by mass spectrometry of the alkali metal adduct ions.

Tetracyclines (TCs) are a family of broad-spectrum antibiotics. During the manufacturing process or storage, epimerization of tetracyclines could occur, leading to 4-epimers which are nearly inactive. From an analytical point of view, isomers are often difficult to distinguish. Previously, four pairs of TCs (oxytetracycline, tetracycline, doxycycline, chlortetracycline and their respective 4-epimers) were differentiated by mass spectrometry (MS) through protonated ions. However, they do not follow common rules and so it is still quite difficult to differentiate between them. In order to solve this, the four pairs were differentiated in the current study by collision induced dissociation (CID) spectra of the alkali adduct ions, including lithium, sodium and potassium. In the spectra of the sodium adducts, all studied tetracyclines showed a tendency to form [M+Na-NH3]+ ions, while the 4-epimers liked to form [M+Na-NH3-H2O]+ ions. Meanwhile, energy resolved mass spectrometry (ERMS) showed that all four 4-epimers' sodium adducts had the tendency to fragment at higher energy points. In the CID spectra of lithium adducts of TCs, a similar trend was observed for three pairs, except for doxycycline. For potassium adducts, the fragmentation was found to be less discriminative. As was derived from the 3D model, the four pairs all interact with the alkali metal through the dimethyl amino group at the C-4 position. The lithium adduct species also bound through the hydroxyl group at the C-5 position. If the TCs did not have a hydroxyl group at the C-5 position, they bound with the hydroxyl group at the C-6 position. For the same TC, with an increase of the diameter of the metal ion, the loss of H2O decreased gradually. As sodium adduct ions are common during the ionization process, TCs and their 4-epimers could be differentiated rapidly by ERMS of the sodium adduct ions.