taxonID	type	description	language	source
66798798FFACFFF1634CFB73FF4AFCB8.taxon	description	The selected tribe Cardamineae species were investigated for GSLs with the overall evolutionary history in mind. In particular, we searched for similarities to the deviating genus Barbarea and C. pratensis. The characteristic GSLs in Barbarea include the homoPhe-derived “ glucobarbarin ” (BAR, 40 S), which is (S) - 2 - hydroxy- 2 - phenylethylGSL, epiglucobarbarin (EBAR, 40 R), which is the epimer (R) - 2 - hydroxy- 2 - phenylethylGSL, the Trp-derived IM (43), and a number of derivatives. Among the derivatives are the disubstituted 1,4 moIM (138) (usually only detectable in roots), phenolic derivatives of both PE, BAR and EBAR, and isoferuloyl derivatives of the same and of IM (only known from seeds) (Supplementary Figure 1). Interestingly, the “ innovative ” C. pratensis also accumulates 1,4 moIM (Olsen et al., 2016). So far unique GSLs in C. pratensis are two hydroxylated GSLs with a 3 - methylpentyl skeleton, 2 h 3 mPe (149) and 3 hmPe (141), both predicted to be biosynthesized from 2 homoIle (Agerbirk et al., 2010; Olsen et al., 2016). One of them (149) was likewise confined to roots (Agerbirk et al., 2010). These recently discovered GSLs were found in some American and commercial accessions, while Danish accessions of C. pratensis contained a variety of other BCAA-derived GSLs and Phe or Tyr-derived GSLs (Agerbirk et al., 2010). A variety of organs were investigated whenever possible, in order to maximize the chance of detecting critical structures. In tables in this section, we consistently distinguished tentative from conclusive identifications based on an MS 2 dependent criterion: For each species, conclusive identification of each reported GSLs was concluded when two independent characteristics of a peak matched an authentic standard (Blaˇzevic´et al., 2020): (1) correct t R of the relevant m / z value was observed in extracted ion chromatograms, and (2) the correct MS 2 spectrum was observed. Correctness of t R and MS 2 were based on comparison with an authentic standard (Table 2). When conclusive identification was obtained from one organ, minor peaks from other organs were accepted as conclusive as well. In contrast, tentative identification (Blaˇzevic´et al., 2020) was concluded when a clearly distinguishable peak was observed but the MS 2 criterion was not fulfilled for any organ of the species, or when reasonable t R and MS 2 was observed but an authentic standard was not available. In either case, the tentative nature was indicated by labelling the data with an asterisk (*). It follows from these criteria that identification of GSLs for which authentic standards were not available were by definition tentative, even for members of a homologous series with expectable MS 2. In those cases, the GSL number in the entire table was labelled with an asterisks, essentially 3 mSp (95 *), 9 mSOn (68 *), 5 mSp (94 *) and 9 mSn ([89] *), S 2 hBuen (24 S *) and 2 RhaOBZ (109 *). For individual levels of GSLs of this small group, an additional asterisk at individual data points means that also MS 2 confirmation was lacking, while individual data points without an asterisk represent data validated by expectable MS 2 for the side chain type. Documentation for botanical identity and specification of origin followed recent recommendations (Zidorn, 2017) as far as possible (Section 4.2.).	en	Agerbirk, Niels, Hansen, Cecilie Cetti, Olsen, Carl Erik, Kiefer, Christiane, Hauser, Thure P., Christensen, Stina, Jensen, Karen R., Ørgaard, Marian, Pattison, David I., Lange, Conny Bruun Asmussen, Cipollini, Don, Koch, Marcus A. (2021): Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS / MS data of reference desulfoglucosinolates. Phytochemistry (112658) 185: 1-19, DOI: 10.1016/j.phytochem.2021.112658, URL: http://dx.doi.org/10.1016/j.phytochem.2021.112658
66798798FFAFFFF1601AFCCFFE56FA8E.taxon	description	Our previous investigations of Barbarea GSL profiles had focused on species from the Northern temperate zone (Agerbirk et al., 2003 a, 2015), while many other species of the genus appeared to be poorly known with respect to chemistry. For the purpose of investigating distantly related species, we turned to two endemic Australian species, Barbarea australis Hook f. and Barbarea grayi Hewson. Both species contained expectable homoPhe and Trp derived GSLs in leaves and roots (Table 3), and no surprising GSLs were observed. Rather, PE and the usual derivatives BAR and EBAR were dominating GSLs, along with expectable indole GSLs. Seed GSLs of B. australis had already been reported (Agerbirk and Olsen, 2011), and included most of the GSLs from leaves and roots (except substituted Trp-derived) as well as low levels of 6 ′ - isoferuloyl derivatives of the major seed GSLs PE and BAR. Seed GSLs of B. grayi were analyzed for this report and contained the expected isoferuloylated GSLs known from the genus, but at rather low levels (Table 3). A large number of putative aliphatic GSLs and derivatives of the major GSLs were searched for but not found (Table 3).	en	Agerbirk, Niels, Hansen, Cecilie Cetti, Olsen, Carl Erik, Kiefer, Christiane, Hauser, Thure P., Christensen, Stina, Jensen, Karen R., Ørgaard, Marian, Pattison, David I., Lange, Conny Bruun Asmussen, Cipollini, Don, Koch, Marcus A. (2021): Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS / MS data of reference desulfoglucosinolates. Phytochemistry (112658) 185: 1-19, DOI: 10.1016/j.phytochem.2021.112658, URL: http://dx.doi.org/10.1016/j.phytochem.2021.112658
66798798FFAFFFF0630AFF35FAECFAE2.taxon	description	The genus Planodes comprises two species only. Seeds of the small weedy plant P. virginica had been investigated previously using now historical methods (Gmelin et al., 1970). Those authors reported the presence of EBAR (40 R). 7 mSOh (66) and 8 mSOo (69), two of which were deduced for the first time in that paper, and one or more lipophilic GSLs that were not identified. We wished to repeat and expand this experiment, testing all major plant parts, with focus on searching for similarities with and contrast to Barbarea and Cardamine species. In particular, we wanted to search for oxidized derivatives of homoPhe derived GSLs, for distubstituted Trp-derived in roots, for acyl derivatives in seeds, and for GSLs derived from BCAAs, as many of these kinds of structures were potentially due to de novo evolution in one or more species in the tribe. Likewise, we wished to know the full complement of n-homoMet-derived GSLs in the species, as this profile was a candidate for an ancestral profile of the Barbarea species. The historical report on the dominant glucosinolates in P. virginica seeds was fully confirmed (Fig. 3 A). Of particular interest was the dominance of EBAR that also occurs in most Barbarea species (Table 4). In addition, minor levels of BAR and NAS were discovered. Of Trp-derived GSLs, only the parent structure IM was detected, in contrast to the prominence of monosubstituted indoles in roots of most other known crucifers. Clearly, this species did not accumulate the peculiar disubstituted indole, 1,4 moIM, known from roots of one Barbarea species. In addition to the previously known dominating methylsulfinyl alkyls with chain length 7 – 8, a wide range of chain lengths were found (Table 5), supported by coelution with authentic standards except in the case of 9 mSOn (68). For 5 mSOp (72), the only authentic standard available was A. thaliana seeds, which was considered reliable. The retention times of the homologs increased in an expectable way for a homologous series (Supplementary Fig. S 3 C). The characteristic MS 2 fragmentation (including loss of CH 3 SOH) was observed for all major peaks including the tentatively identified 9 mSOn (Supplementary Fig. S 3 D). From the latter fragmentation, isomeric hydroxylated methylthioalkyls could be ruled out. The ‘ bell-shaped’ level distribution of chain lengths (Table 5) was similar to distributions in some other species, as expected from the properties of MAM-enzymes that are responsible for the chain elongation (Textor et al., 2007; Kumar et al., 2019; Petersen et al., 2019). The more lipophilic methylthioalkyl glucosinolates were mainly detected in seeds (Table 5). Only two were identified with certainty, 8 mSo from comparison with an authentic standard, and 7 mSh from comparison with A. thaliana seeds, considered a reliable reference material. However, additional members of the entire homologous series from n = 5 to n = 9 were tentatively detected, supported by reasonable t R compared to that of 8 mSo, 7 mSh and of 4 mSb available as standards. Due to the wide range of GSLs confirmed in P. virginica seeds and their public availability at Nordgen, we imagined that they could be useful as reference material and compiled various illustrative chromatograms (Supplementary Fig. S 3). Pioneers in the study of P. virginica also confirmed the R - configuration of the chiral sulfur in the side-chain methylsulfinyl groups (Gmelin et al., 1970), which adds further reliability to this reference material. Since P. virginica shared high levels of EBAR with Barbarea, some other characteristic Barbarea GSLs were searched for (Table 4). ThioGlcacylated GSLs were searched for but not found. Similarly, hydroxyl and methoxyl derivatives of the major EBAR were searched for. A possible trace of the 4 - hydroxy derivative 4 hEBAR (139 R) was detected in many samples at very low levels but not confirmed by MS 2 using ion trap HPLC-MS. As 4 - hydroxylation is seasonally regulated in Barbarea (Agerbirk and Olsen, 2015), a new set of samples were collected earlier in the next year (accession 2), in an attempt to confirm or reject the presence of 4 hEBAR. The putative trace of 4 hEBAR was also observed in this second accession, but of the same very low level as in the first accession and still MS 2 confirmation was not obtained by ion trap HPLC-MS. If 4 hBAR or 4 hEBAR were present, it would be the first occurrence outside the genera Barbarea and Arabis. In order to settle the uncertainty, we carried out more sensitive UHPLC-QToF MS / MS with higher mass- and chromatographic resolution. At these conditions, authentic desulfo 4 hBAR and 4 hEBAR mainly resulted in proton adducts after loss of water; [M-H 2 O + H] +. Re-analysis of three P. virginica leaf samples of accession 2 and seeds of accession 1 demonstrated that 4 hEBAR was present at very low levels, supported by matching t R and MS 2. The high-resolution mass matched the expected (found: 358.0953 (mean, N = 4), calculated for C 15 H 20 O 7 NS + ([M-H 2 O + H] +): 358.0955). Critical MS 2 features supporting the identification was loss of anhydroGlc and a characteristic major loss of m / z 220, plus a combined loss of anhydroGlc and hydroxylamine (Supplementary Fig. S 4 A-C). In the seed sample, which had the highest level, loss of hydroxylamine from the [M – H 2 O – ahGlc + H] + ion was also confirmed (Supplementary Fig. S 4 E). We concluded that 4 hEBAR had been detected conclusively in two accessions of P. virginica, but at rather low levels (Table 4). We did not detect any other known derivatives of EBAR (the 3 - hydroxy derivative 3 hEBAR (142 R), the 4 - methoxy derivative 4 moEBAR (50 R) or the 3 / 4 methoxy-hydroxy derivative “ x 1 ”) in any sample. As P. virginica shared chain elongated Met derived GSLs with A. thaliana, some rare and characteristic GSLs from this species were searched for but not found (hydroxyalkyls and their benzoyl esters). This negative result was likewise obtained for two more species discussed below (Table 5). We also searched for GSLs derived from BCAAs, because their distribution in the tribe was poorly known. No GSL derived from non-chain elongated BCAAs was detected (Table 5). However, a peak corresponding to an isomeric hydroxyhexylGSL was detectable at minute levels, with a further isomer at close to trace levels (Supplementary Figure 4 H, panels H 1 – H 3). The sum formula was supported by QToF-MS (found 340.1424, calculated for [C 13 H 26 O 7 NS] +: 340.1425) (panel H 1). Two such isomers are known, 141 and 149. The t R of the unknown dGSLs were far from that of d 141 but the major was close to d 149. However, a reproducible slight (0.1 min) mismatch with d 149 was observed using UHPLC, confirmed by spiking that resulted in a splitpeak (results not shown). A quite characteristic MS 2 ion trap spectrum, with six major fragment ions, was confirmed in several samples (Supplementary Fig. S 3 G). The ion trap MS 2 included an e-fragment (Table 1), which is indicative of a β- hydroxylated dGSL (Olsen et al., 2016). We conclude the presence of a hitherto unknown isomeric β- hydroxyhexyl GSL at levels around 0.01 μmol / g dry wt. in all tested plant parts of both P. virginica accessions. Possible identities could be a straight chain isomer or “ 2 h 4 mPe ” derived from 2 - homoLeu, not yet known to science. Several structurally related known GSLs were searched for but not found, including the parent GSL 3 mPe, the δ- hydroxylated isomer 3 hmPe and lower homologs (Table 5).	en	Agerbirk, Niels, Hansen, Cecilie Cetti, Olsen, Carl Erik, Kiefer, Christiane, Hauser, Thure P., Christensen, Stina, Jensen, Karen R., Ørgaard, Marian, Pattison, David I., Lange, Conny Bruun Asmussen, Cipollini, Don, Koch, Marcus A. (2021): Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS / MS data of reference desulfoglucosinolates. Phytochemistry (112658) 185: 1-19, DOI: 10.1016/j.phytochem.2021.112658, URL: http://dx.doi.org/10.1016/j.phytochem.2021.112658
66798798FFAEFFF0630AFAE5FA05F8E9.taxon	description	The genus had been investigated in detail earlier (Olsen et al., 2016), except seeds of R. sylvestris. Here, the seeds were investigated with the aim to search for seed-specific acylated GSL (Table 5). Green parts and flowers of the sampled individual matched the previously reported profiles (results not shown). The GSL profile of the seeds was dominated by 6 homoMet-derived 8 mSOo (69) and the corresponding methylthio precursor 8 mSo (92), i. e. the “ C 8 ” chain length group corresponding to six rounds of chain elongation in the early biosynthesis, in agreement with the previously reported leaf and root profiles. From visual comparison of chain-length distributions in the common table (Table 5), the distribution in R. sylvestris was obviously more narrow than seen for Nasturtium and Planodes, suggesting a more stringent, processive chain elongation system (Textor et al., 2007; Kumar et al., 2019; Petersen et al., 2019). Acylated GSLs, BCAA-derived, Trp-derived or Phe-derived GSLs were not detected (Table 4, Table 5) but Trp-derived GSLs are confirmed in other organs of the same species (Olsen et al., 2016).	en	Agerbirk, Niels, Hansen, Cecilie Cetti, Olsen, Carl Erik, Kiefer, Christiane, Hauser, Thure P., Christensen, Stina, Jensen, Karen R., Ørgaard, Marian, Pattison, David I., Lange, Conny Bruun Asmussen, Cipollini, Don, Koch, Marcus A. (2021): Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS / MS data of reference desulfoglucosinolates. Phytochemistry (112658) 185: 1-19, DOI: 10.1016/j.phytochem.2021.112658, URL: http://dx.doi.org/10.1016/j.phytochem.2021.112658
66798798FFA3FFFB634CF96BFA8EF8F0.taxon	description	As observed for the Australian Barbarea species (Section 2.2.1.), aliphatic GSLs generally appear to be absent from the genus, except for a single historical report of isothiocyanate (ITC) hydrolysis products of aliphatic GSLs, prop- 2 - enyl ITC, isopropyl ITC and 3 - (methylthio) propyl ITC, from Barbarea spp. (Cole, 1976). However, identification of some ITCs in that report was of questionable reliability (Olsen et al., 2016). Indeed, that historical paper also (unexpectedly) reported prop- 2 - enyl ITC from Plantago major L., (Plantaginaceae: Lamiales] although competent researchers were later unable to detect any GSL in that species (Larsen et al., 1983). However, more recently data also suggested the possibility of aliphatic GSLs in Barbarea. Three observations were particularly suggestive: CYP 79 F 6 and other putative GSL biosynthetic enzymes were reported to be induced by larvae of the moth Plutella xylostella (L.) (Liu et al., 2016), B. vulgaris amino acid chain elongating enzymes were reported to chain elongate Met and Leu in addition to Phe (Wang et al., in press), and putative GSL products were induced by spraying leaves with 10 mM CuCl 2 (aq.) (Pedras et al., 2015). Hence, we decided to search for proposed aliphatic GSLs after various kinds of induction and from normal growth conditions. Specifically, we re-investigated both ecotypes of B. vulgaris for 2 - propenylGSL (Pren, 107) and related aliphatic GSLs by ion trap HPLC-MS inspection for trace peaks, experimental testing of recovery of added GSLs, and investigation of plants challenged by herbivory and CuCl 2. We also investigated the morphologically deviating B. vulgaris ssp. vulgaris. The GSL precursor of 2 - propenyl ITC, Pren, was not detectable in B. vulgaris (Fig. 6 B), in agreement with previous investigations of a range of P-type and G-type accessions from a large geographical range or harvested at different seasons (Agerbirk et al., 2015 and references cited therein). Experimental addition of (intact) Pren to crude extracts followed by desulfation confirmed linear detection of this GSL down to trace levels (Fig. 6 C – E). The limit of detection was estimated as 0.1 μmol / g dry wt. at normal sample concentration (Fig. 6 E) and 10 - fold lower when using concentrated samples or high injection volumes. In the latter case, however, linearity of dominating GSLs suffered. The morphologically deviating ssp. vulgaris (in this report represented by accession B 59), characterized by erect (vertical) rather than arcuate siliques, had a GSL profile indistinguishable from that of G-type plants (results not shown), in agreement with a previous report including ssp. vulgaris (Agerbirk et al., 2003). We subjected P and G-type B. vulgaris to herbivory by larvae of the butterfly Pieris brassicae L. (3 days or 7 days), or by P. xylostella larvae (4 days), to spraying with 10 mM CuCl 2 (harvest after 4 days), and a parallel control treatment. These treatments were followed by GSL analysis using HPLC-UV for observing the general GSL profile and by ion trap HPLC-MS with injection of quite concentrated samples for a search for induced GSLs. The general leaf GSL profile was as expected for the type in both control and challenged plants, and we observed no candidate induced GSL peak. Some rather drastic treatments resulted in lower mean GSL levels, such as herbivory by the large P. brassicae for 7 days, after which time much of the leaves had been eaten (Fig. 7). The lower GSL levels in remaining leaves may well be due to preferential ingestion of GSL rich plant parts of these GSL-stimulated larvae. We carefully searched for but did not detect Pren in any HPLC-MS sample in extracted ion chromatograms. Likewise, we systematically searched for but did not detect 1 mEt, 1 mPr, 2 mPr, 2 mBu, 2 h 2 mBu, 3 mPe, 2 h 3 mPe, 3 hmPe, 4 mSb, 4 mSOb, Buen, 8 mSOo or BZ, selected from prominent GSLs in other tribe Cardamineae members and from GSLs produced by combination of A. thaliana and B. vulgaris biosynthetic enzymes after heterologous expression in tobacco (Wang et al., in press). Neither did visual inspection reveal any other new GSL for the species. However, we did detect trace levels of the known minor constituents 4 mIM and PE, included in each search as a positive control. During the final editing of the present work, a claim of ITCs corresponding to 3 mSOp (73) and 4 mSObuen (63) in P. xylostella - induced B. vulgaris was published (Hussain et al., 2020). The claim, lacking actual evidense for the identifications as well as levels calculated on a plant weight basis, was based on analysis of G-type B. vulgaris of the Hedeland accession also used in our induction experiments, and no other isothiocyanate (such as the expected phenethyl ITC corresponding to PE) was reported (Hussain et al., 2020). Hence, we searched three desulfoGSL analysis files of P. xylostella - induced B. vulgaris, including both the Hedeland and Suserup accessions, for any signs of d 63 or d 73. We also searched for d 95 from the expected biosynthetic precursor 95. No trace of any of these dGSLs was detected, although we are able to detect them in general (Table 2). We also searched non-induced control samples with the same negative result. We notice that one of the claimed ITCs (from 73) would be isobaric with the expected ITC from PE (105), and that the corresponding dGSL (d 105) was detected as a minor constituent in several of our P. xylostella induced and control samples. In conclusion, there are no reliable reports of aliphatic GSLs in the genus Barbarea. In contrast, extensive scrutiny has failed to reveal aliphatic GSLs above an estimated limit of detection of 0.1 – 0.01 μmol / g dry wt.	en	Agerbirk, Niels, Hansen, Cecilie Cetti, Olsen, Carl Erik, Kiefer, Christiane, Hauser, Thure P., Christensen, Stina, Jensen, Karen R., Ørgaard, Marian, Pattison, David I., Lange, Conny Bruun Asmussen, Cipollini, Don, Koch, Marcus A. (2021): Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS / MS data of reference desulfoglucosinolates. Phytochemistry (112658) 185: 1-19, DOI: 10.1016/j.phytochem.2021.112658, URL: http://dx.doi.org/10.1016/j.phytochem.2021.112658
66798798FFA3FFFB634CF96BFA8EF8F0.taxon	discussion	2.3. Phylogenetic analysis of Barbarea vulgaris accessions In order to better understand whether the tested types and subspecies of B. vulgaris were representative for the entire species, we sequenced the nuclear ribosomal DNA internal transcribed spacer (ITS) region of a range of our accessions. For the P-type, we included our Danish “ typeaccession ” (B 4) as well as three accessions representing major parts of northern Russia (Supplementary Table S 1), and contrasted with two different morphological types of plants with glabrous leaves: the G-type (Danish “ type accession ” B 44) of ssp. arcuata (characterized by arcuate siliques) and a newly collected accession (B 59) with erect (vertical) siliques traditionally named ssp. vulgaris by Danish botanists (Hartvig et al., 2015). The sequence variation of our accessions supported distinction of P- and G-types, with only two sequence types detected in the six sequenced accessions. The G-type as well as accession B 59 of the morphological type traditionally known as ssp. vulgaris shared the same ITS sequence, while all four sequenced P-type accession differed from the G-type sequence at eight nucleotide positions (Table 8) but where identical to each other. This ITS sequence variation was subsequently compared with all B. vulgaris ITS sequences available from GenBank by mid- 2019, most of them from Lange et al. (in review), resulting in a slightly more complex pattern. In this and the following discussion, we accepted the original sequence-contributors’ choice of intraspecific nomenclature. Currently, no intraspecific taxa are generally accepted and the intraspecific nomenclature of B. vulgaris is in need of revision (Lange et al., in review). The names represented with the GenBank material were: G-type, P-type and two traditionally recognized subspecies with more or less erect siliques; ssp. vulgaris and ssp. rivularis (Hegi, 1958). In total, seven ITS sequence variants were represented in GenBank (Suppl. Table S 1, Table 8). The majority of these sequences likewise grouped with either our G-type or P-types in agreement with their respective type identities. One group (Group 4) was intermediary of the P-type and G-type sequences, and was tentatively interpreted as a hybrid of the G- and P-types. Indeed, one of them had been deposited as a hybrid sequence. Another group (group 7) was dominated by sequences deposited as ssp. rivularis and ssp. vulgaris and was more similar to the Gtype than to the P-type (Table 8), for simplicity this group is named the rivularis group here, although the name may be temporary. A few GenBank sequences did not agree with the group designations used here. Three sequences attributed to PxG hybrids clustered with the G-types in sequence group 1, as did two sequences attributed to ssp. vulgaris, including our accession B 59. Likewise, one sequence attributed to G-type and two sequences attributed to hybrids grouped with the Ptypes in sequence group 6. Three other sequence groups (Groups 2, 3 and 5) represented sequences deposited as G-type, GxP hybrid or undetermined. These groups were represented by few deposited sequences. The parsimony network analysis (Fig. 8) suggested two main clusters of B. vulgaris, the one including the P-type and the small group 5 and the other including most of the remaining groups (G-type, rivularis and the small sequence groups 2 and 3). Group 4, tentatively interpreted as hybrid of P- and G-types, was intermediate. A separation of B. vulgaris into two gene pools, correlated with G-type and P-type, has also been shown earlier using different genetic marker systems (Christensen et al., 2014; Toneatto et al., 2012). In conclusion, the investigated G-type and P-type of B. vulgaris are rather representative for the known genetic variation of the species, which has mainly been investigated in the European part of its main original distribution area (Christensen et al., 2014; Toneatto et al., 2010, 2012). Hence, lack of aliphatic GSLs may be a general property of the species (and genus). However, other sequence groups were found, with the “ rivularis ” group most deviating. These groups may represent other chemotypes as well. Hence, lack of aliphatic GSLs in the genus (Section 2.2.7.) can of course only be strictly concluded for the accessions investigated, not in general for the species and genus.	en	Agerbirk, Niels, Hansen, Cecilie Cetti, Olsen, Carl Erik, Kiefer, Christiane, Hauser, Thure P., Christensen, Stina, Jensen, Karen R., Ørgaard, Marian, Pattison, David I., Lange, Conny Bruun Asmussen, Cipollini, Don, Koch, Marcus A. (2021): Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS / MS data of reference desulfoglucosinolates. Phytochemistry (112658) 185: 1-19, DOI: 10.1016/j.phytochem.2021.112658, URL: http://dx.doi.org/10.1016/j.phytochem.2021.112658
66798798FFA5FFF9634CF89DFE66FECF.taxon	description	To provide a perspective of the diversity in the tribe Cardamineae, the species Reseda luteola (family: Resedaceae) was investigated because the literature suggested the GSL profile to remarkably resemble that of B. vulgaris. The presence of BAR in R. luteola is well-established (Kjaer and Gmelin, 1958; Bennett et al., 2004), despite the great evolutionary distance to Barbarea. We revisited this species in order to check for traces of the epimer EBAR and related GSLs. The major peaks were confirmed to be BAR, PE and IM, in seeds (Fig. 9 A – C) and in leaves. A minor peak was conclusively identified as EBAR (Fig. 9 D). However, hydroxy or methoxy substituted glucobarbarins were not detected (Fig. 9 E – F). As these are available as references from Barbarea and Arabis spp. and known to be detectable and base line separated from the R. luteola peaks at our conditions (Table 2; Olsen et al., 2016), their absence (above the limit of detection) could be concluded with certainty. The level of EBAR in R. luteola seeds corresponded to ca. 5 % of the sum of EBAR and BAR as estimated from peak intensities, but was relatively lower in leaves, ca. 1 % of the sum of EBAR and BAR. The varying proportion of the epimeric glucobarbarins in seeds and leaves of R. luteola suggests that each epimer is due to a distinct genetic locus and biosynthetic enzyme, as is the case in B. vulgaris (Liu et al., 2019 a). Putative constituents like BZ, 1 mEt, 1 mPr / 2 mPr and isomers of mBu and mPe were searched for in both leaves and seeds of R. luteola but not detected (results not shown). However, an apparent isomer of hydroxybutylGSL was detected (Fig. 9 G), suggesting that use of amino acid precursors for seed GSL biosynthesis in R. luteola is not quite as restricted, to Trp and homoPhe, as in B. vulgaris, but include an aliphatic precursor. We further investigated the apparent hydroxybutylGSL in R. luteola. Only trace-levels were found in seeds, but analysis of leaves revealed appreciable levels of the apparent hydroxybutylGSL (Fig. 9 G), which eluted at 1.3 min and hence was not 1 hmPr (30) but could be either Met-derived 4 hBu ([26]) (not previously known from basal families) or Leuderived 2 h 2 mPr (31), not available in our reference library at that time, or several yet unknown isomers. Ion trap MS 2 was inconspicious and identical to both the MS 2 of d [26] and d 30. We re-analyzed the sample using UHPLC-QToF MS / MS and found the high-resolution mass to be in agreement with a desulfo hydroxybutylGSL (found: 312.1114 (mean, N = 2), calc. for C 11 H 22 O 7 NS + ([M + H] +): 312.1112). This result was in agreement with a previous report of an unspecified hydroxybutyl GSL in leaves of R. luteola (Griffiths et al., 2001). Neither BAR nor EBAR were detected in Reseda odorata seeds in the present study, in contrast to a previous report (Bennett et al., 2004). The dominating dGSL peak at m / z = 514 was presumably due to the well-established 2 RhaOBZ (109) (Pagnotta et al., 2020). The dGSL d 109 showed a distinct MS 2 compared to that from the isomer d 110 (Supplementary Fig. S 1 C) and confirmed our ability to detect a wide range of structural types including glycosides (Fig. 9 H). A minor peak of desulfo IM almost coeluted with desulfo 2 RhaOBZ. A range of aliphatic GSLs including butyls and hydroxybutyls were searched for but not detected in seeds. While both Reseda species accumulated IM, neither of the usual IM derivatives for tribe Cardamineae (Supplementary Fig. S 1 A) were detected, although they were specifically searched for. No previous author has reported IM derivatives from the genus, even when roots were examined (Pagnotta et al., 2020). However, roots should be investigated critically before concluding on presence or absence of substituted indole GSLs in this genus.	en	Agerbirk, Niels, Hansen, Cecilie Cetti, Olsen, Carl Erik, Kiefer, Christiane, Hauser, Thure P., Christensen, Stina, Jensen, Karen R., Ørgaard, Marian, Pattison, David I., Lange, Conny Bruun Asmussen, Cipollini, Don, Koch, Marcus A. (2021): Glucosinolate profiles and phylogeny in Barbarea compared to other tribe Cardamineae (Brassicaceae) and Reseda (Resedaceae), based on a library of ion trap HPLC-MS / MS data of reference desulfoglucosinolates. Phytochemistry (112658) 185: 1-19, DOI: 10.1016/j.phytochem.2021.112658, URL: http://dx.doi.org/10.1016/j.phytochem.2021.112658
