Exploration of underutilized crop diversity of Capsicum peppers in their primary center of diversity in Bolivia and Peru

The genus Capsicum is a highly diverse complex of domesticated and wild species that displays abundant variation in its main center of domestication and diversity in Bolivia and Peru but that remains under-researched. New collecting expeditions undertaken in 2010 by the Instituto Nacional de Innovación Agraria (INIA) in Peru and the Centro de Investigaciones Fitoecogenéticas de Pairumani (CIFP) in Bolivia have significantly increased the size of the collections. INIA Peru now maintains 712 accessions of the five domesticated species, making it one of the largest and most diverse national collections of native Capsicum pepper varieties in the world. The collection in Bolivia contains 492 accessions, including the five domesticated species, four wild species, and one wild botanical variety of a domesticated species. We report on the identification of promising native Capsicum germplasm for potential use in the development of differentiated products. Identification of promising material representative of native Capsicum diversity in both collections followed several steps: (1) Identification of a core collection of nearly 100 accessions per country representing the different species and their geographic distribution. Dried samples of these accessions were biochemically screened for commercially interesting attributes including capsaicinoid content, polyphenols, antioxidant capacity, carotenoids, lipid content and color; (2) Based on results of the biochemical screening, sub-sets of 44 Bolivian and 39 Peruvian accessions were selected, representing the different species and variation in biochemical attributes; and (3) The selected materials were grown in different environments to identify the agro-ecological conditions were they best express the special properties of commercial interest. The biochemical screening and agromorphological characterization and evaluation revealed that Capsicum accessions from Bolivia and Peru have unique combinations of functional attributes, confirming that a wealth of commercially valuable properties can be found in Capsicum’s primary center of diversity. This study was financed by GIZ

Fatty acids are crucial to fuel NK cells upon acute retrovirus infection

Natural killer (NK) cells are cytotoxic innate immune cells, able to recognize and eliminate virus-infected as well as cancer cells. Metabolic reprogramming is crucial for their activity as they have enhanced energy and nutritional demands for their functions during an infection. Fatty acids (FAs) represent an important source of cellular energy and are essential for proliferation of immune cells. However, the precise role of FAs for NK cells activity in retrovirus infection was unknown. Here we show that activated NK cells increase the expression of the FA uptake receptor CD36 and subsequently the uptake of FAs upon acute virus infection. We found an enhanced flexibility of NK cells to utilize FAs as source of energy compare to naïve NK cells. NK cells that were able to generate energy from FAs showed an augmented target cell killing and increased expression of cytotoxic parameters. However, NK cells that were unable to generate energy from FAs exhibited a severely decreased migratory capacity. Our results demonstrate that NK cells require FAs in order to fight acute virus infection. Susceptibility to severe virus infections as it is shown for people with malnutrition may be augmented by defects in the FA processing machinery, which might be a target to therapeutically boost NK cell functions in the future

Oxylipin metabolism is controlled by mitochondrial β-oxidation during bacterial inflammation

Oxylipins are potent biological mediators requiring strict control, but how they are removed en masse during infection and inflammation is unknown. Here we show that lipopolysaccharide (LPS) dynamically enhances oxylipin removal via mitochondrial β-oxidation. Specifically, genetic or pharmacological targeting of carnitine palmitoyl transferase 1 (CPT1), a mitochondrial importer of fatty acids, reveal that many oxylipins are removed by this protein during inflammation in vitro and in vivo. Using stable isotope-tracing lipidomics, we find secretion-reuptake recycling for 12-HETE and its intermediate metabolites. Meanwhile, oxylipin β-oxidation is uncoupled from oxidative phosphorylation, thus not contributing to energy generation. Testing for genetic control checkpoints, transcriptional interrogation of human neonatal sepsis finds upregulation of many genes involved in mitochondrial removal of long-chain fatty acyls, such as ACSL1,3,4, ACADVL, CPT1B, CPT2 and HADHB. Also, ACSL1/Acsl1 upregulation is consistently observed following the treatment of human/murine macrophages with LPS and IFN-γ. Last, dampening oxylipin levels by β-oxidation is suggested to impact on their regulation of leukocyte functions. In summary, we propose mitochondrial β-oxidation as a regulatory metabolic checkpoint for oxylipins during inflammation

Metabolic dysregulation of the lysophospholipid/autotaxin axis in the chromosome 9p21 gene SNP rs10757274

Background - Common chromosome 9p21 SNPs increase coronary heart disease (CHD) risk, independent of 'traditional lipid risk factors'. However, lipids comprise large numbers of structurally related molecules not measured in traditional risk measurements, and many have inflammatory bioactivities. Here we applied lipidomic and genomic approaches to three model systems, to characterize lipid metabolic changes in common Chr9p21 SNPs which confer ~30% elevated CHD risk associated with altered expression of ANRIL, a long ncRNA. Methods - Untargeted and targeted lipidomics was applied to plasma from Northwick Park Heart Study II (NPHSII) homozygotes for AA or GG in rs10757274, followed by correlation and network analysis. To identify candidate genes, transcriptomic data from shRNA downregulation of ANRIL in HEK293 cells was mined. Transcriptional data from vascular smooth muscle cells differentiated from iPSCs of individuals with/without Chr9p21 risk, non-risk alleles, and corresponding knockout isogenic lines were next examined. Last, an in-silico analysis of miRNAs was conducted to identify how ANRIL might control lysoPL/lysoPA genes. Results - Elevated risk GG correlated with reduced lysophosphospholipids (lysoPLs), lysophosphatidic acids (lysoPA) and autotaxin (ATX). Five other risk SNPs did not show this phenotype. LysoPL-lysoPA interconversion was uncoupled from ATX in GG plasma, suggesting metabolic dysregulation. Significantly altered expression of several lysoPL/lysoPA metabolising enzymes was found in HEK cells lacking ANRIL. In the VSMC dataset, the presence of risk alleles associated with altered expression of several lysoPL/lysoPA enzymes. Deletion of the risk locus reversed expression of several lysoPL/lysoPA genes to non-risk haplotype levels. Genes that were altered across both cell datasets were DGKA, MBOAT2, PLPP1 and LPL. The in-silico analysis identified four ANRIL-regulated miRNAs that control lysoPL genes as miR-186-3p, miR-34a-3p, miR-122-5p, miR-34a-5p. Conclusions - A Chr9p21 risk SNP associates with complex alterations in immune-bioactive phospholipids and their metabolism. Lipid metabolites and genomic pathways associated with CHD pathogenesis in Chr9p21 and ANRIL-associated disease are demonstrated

Facile preparation of embedded polar group-containing pentafluorophenyl stationary phases for highly selective separations of diverse analytes

Pentafluorophenyl (PFP) stationary phase is one of the most important phases after the C18 phase in terms of its applications. Three embedded polar groups (EPG)-containing stationary phases were newly synthesized to act the EPGs as additional interaction sites. The silica surface was initially modified with (3-aminopropyl)trimethoxysilane (APS). The APS-modified silicas were coupled with 2,3,4,5,6-pentafluorobenzoic acid, 2,3,4,5,6-pentafluorophenylacetic acid, and 2,3,4,5,6-pentafluoro-anilino(oxo)acetic acid to obtain Sil-PFP-BA, Sil-PFP-AA, and Sil-PFP-AN phases, respectively. The new phases were characterized by elemental analysis, ATR-FTIR, scanning electron microscopy (SEM), and thermogravimetric analysis (TGA). The phases were evaluated with the Tanaka and Neue tests in reversed-phase liquid chromatography (RPLC). In addition, they were characterized as hydrophilic phases by the Tanaka test protocol used in hydrophilic interaction chromatography (HILIC) separation mode. The Sil-PFP-AA phase showed the highest molecular shape selectivity in RPLC, while Sil-PFP-AN achieved the highest separability in HILIC compared to the commercial PFP reference column. The Sil-PFP-AA phase was successfully applied for the analysis of capsaicinoids from real samples of fresh chili peppers (Capsicum spp.) in RPLC and the Sil-PFP-AN phase for vitamin C (ascorbic acid) in HILIC.Abul Khayer Mallik highly acknowledges the Alexander von Humboldt Foundation for the award and partial support to carry out the research work. We also acknowledge Matthias Epple, University of Duisburg-Essen, Germany for giving us the opportunity to measure the ATR-FTIR and SEM.Peer reviewe

Comparison of derivatization/ionization techniques for liquid chromatography tandem mass spectrometry analysis of oxylipins

The performance of two derivatization and ionization techniques for the quantitative reversed phase liquid chromatography (LC)- mass spectrometry (MS) analysis of hydroxy fatty acids (OH-PUFA) in plasma was evaluated: One used AMPP (N-(4-aminomethylphenyl)pyridinium chloride) leading to a positive charged amid-derivate which can be detected by electrospray ionization (ESI)-MS. Second yielded penta fluorobenzyl bromide (PFB) ester derivates allowing detection in electron capture atmospheric pressure chemical ionization (ecAPCI)-MS. The sensitivity of detection of a comprehensive set of hydroxy fatty acids of n6- and n3- poly unsaturated fatty acids was investigated. On the SCIEX3200 MS the applied PFB derivatization led to poor limits of detection (LOD) of 10–100 nM (0.1–1 pmol/0.03–0.3 ng on column). By contrast, AMPP derivatization led to a similar sensitivity compared to the standard ESI(-) of non derivatized analytes (LOD about 1 nM (10 fmol/3 pg on column)). For several analytes, including 9-HETE, 11-HETE and 17-HDHA the AMPP derivatization improved sensitivity enabling their detection in human plasma. However, precision was reduced by AMPP derivatization and variation in IS recovery indicated a strong matrix influence on the MS-signal. In sum, with the instrumentation used, neither of these derivatization methods improves in our hands the LC–MS based quantification of oxylipins

Comparison of derivatization/ionization techniques for liquid chromatography tandem mass spectrometry analysis of oxylipins

The performance of two derivatization and ionization techniques for the quantitative reversed phase liquid chromatography (LC)- mass spectrometry (MS) analysis of hydroxy fatty acids (OH-PUFA) in plasma was evaluated: One used AMPP (N-(4-aminomethylphenyl)pyridinium chloride) leading to a positive charged amid-derivate which can be detected by electrospray ionization (ESI)-MS. Second yielded penta fluorobenzyl bromide (PFB) ester derivates allowing detection in electron capture atmospheric pressure chemical ionization (ecAPCI)-MS. The sensitivity of detection of a comprehensive set of hydroxy fatty acids of n6- and n3- poly unsaturated fatty acids was investigated. On the SCIEX3200 MS the applied PFB derivatization led to poor limits of detection (LOD) of 10–100 nM (0.1–1 pmol/0.03–0.3 ng on column). By contrast, AMPP derivatization led to a similar sensitivity compared to the standard ESI(-) of non derivatized analytes (LOD about 1 nM (10 fmol/3 pg on column)). For several analytes, including 9-HETE, 11-HETE and 17-HDHA the AMPP derivatization improved sensitivity enabling their detection in human plasma. However, precision was reduced by AMPP derivatization and variation in IS recovery indicated a strong matrix influence on the MS-signal. In sum, with the instrumentation used, neither of these derivatization methods improves in our hands the LC–MS based quantification of oxylipins

Supporting Information - Surface modification of silica with β-alanine derivatives for unique applications in liquid chromatography

More details are in the experimental, materials and method sections; additional physical properties of the stationary phase and other comparative results.Peer reviewe

Surface modification of silica with β-alanine derivatives for unique applications in liquid chromatography

Column purchasing cost is an important issue for an analyst to analyze complex sample matrices. Here, we report the development of an amino acid (β-alanine)-derived stationary phase (Sil-Ala-C12) with strategic and effective interaction sites (amide and urea as embedded polar groups with C12 alkyl chain) able to separate various kinds of analytes. Owing to the balanced hydrophobicity and hydrophilicity of the phase, it showed exceptional separation abilities in both reversed-phase high-performance liquid chromatography (RP-HPLC) as a hydrophobic phase and hydrophilic interaction chromatography (HILIC) as a hydrophilic phase. Remarkably, the baseline separation was achieved for the challenging β- and γ-isomers of tocopherol. Usually, three columns such as pentafluorophenyl or C30, C18, and sulfobetaine HILIC are required for the analysis of vitamin E, capsaicinoids, and vitamin C in chili peppers (Capsicum spp.), respectively. However, only Sil-Ala-C12 was able to separate these analytes. A single column can serve 3–4 purposes, which suggests that Sil-Ala-C12 had the potential to reduce column purchasing costs.Dr. Abul K. Mallik highly acknowledges Alexander von Humboldt Foundation for their fellowship and partial research grant for this research work.Peer reviewe

Capsaicinoids, flavonoids, tocopherols, antioxidant capacity and color attributes in 23 native Peruvian chili peppers (Capsicum spp.) grown in three different locations

Twenty-three Peruvian chili pepper accessions, belonging to the four domesticated species Capsicum annuum, Capsicum baccatum, Capsicum chinense and Capsicum frutescens, were grown under different meteorological conditions and agricultural practices in three Peruvian locations (Chiclayo, Piura and Pucallpa). Results are reported for powdered oven-dried bulk samples of each accession and each location by important quality attributes (capsaicinoids, flavonoids, tocopherols, antioxidant capacity, total polyphenols, extractable color (ASTA 20.1) and surface color). Multivariate data evaluation by principle component analysis and partial least square discriminant analysis did not show any underlying structure. Moreover, a high influence of the environment on the analyzed traits could be demonstrated by analysis of variance. Significant differences (p ≤ 0.001) between the accessions and all locations were observed for all traits. Besides, significant interaction between accessions and locations indicated that the accessions responded differently to changes of the locations. The calculation of an environmental impact factor allowed differing between chili peppers provided consistent phytochemical levels widely independent of the location or those that provided exceptional high levels for a specific trait at one of the locations