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Jean-hugues Renault - One of the best experts on this subject based on the ideXlab platform.
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pilot scale ion exchange centrifugal partition chromatography purification of Sinalbin from white mustard seeds
IEEE Journal of Solid-state Circuits, 2009Co-Authors: Alix Toribio, Benoit Pinel, François De La Poype, Jean-marc Nuzillard, Leslie Boudesocque, Michel Lafosse, Jean-hugues RenaultAbstract:The purification of p-hydroxybenzylglucosinolate (Sinalbin) on a multigram scale from a crude aqueous extract of white mustard seeds (Sinapis alba var. concerta) was successfully achieved by scaling up a strong ion-exchange centrifugal partition chromatography (SIXCPC) laboratory procedure. Thus, the one-step Sinalbin purification was performed with 2.35 g of crude extract in approximately 170 min (830 mg/h) up to 70.3 g in approximately 160 min (26.3 g/h) by switching from a 200 mL laboratory scale column to a 5.7 L pilot-scale column. The required biphasic solvent system contained ethyl acetate, n-butanol, and water in 3:2:5 v/v/v proportions, Aliquat 336 (trioctylmethyl ammonium chloride) was added to the organic stationary phase (80 mM) and acted as ion-exchanger. Potassium iodide in the aqueous mobile phase (80 mM) was used as Sinalbin displacer. The 28.5 mass scale factor arose from the increase in mobile phase flow-rate (from 2 to 50 mL/min), from the higher mass of injected white mustard seed extract (from 12 to 350 g), and from the calculated productivity (from 830 mg to 26.3 g). These results demonstrate that industry scale production of glucosinolates is easily performed by SIXCPC, thus providing pure reference standards for pharmacology studies.
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strong ion exchange centrifugal partition chromatography as an efficient method for the large scale purification of glucosinolates
Journal of Chromatography A, 2007Co-Authors: Alix Toribio, Jean-marc Nuzillard, Jean-hugues RenaultAbstract:The glucosinolates Sinalbin and glucoraphanin were purified by strong ion-exchange displacement centrifugal partition chromatography (SIXCPC). The optimized conditions involved the biphasic solvent system ethyl acetate/n-butanol/water (3:2:5, v/v), the lipophilic anion-exchanger Aliquat 336 (trioctylmethylammonium chloride, 160 and 408 mM) and a sodium iodide solution (80 and 272 mM) as displacer. Amounts as high as 2.4 g of Sinalbin and 2.6g of glucoraphanin were obtained in one step in 2.5 and 3.5h respectively, starting from 12 and 25 g of mustard and broccoli seed aqueous extracts, using a laboratory scale CPC column (200 mL inner volume).
Alix Toribio - One of the best experts on this subject based on the ideXlab platform.
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pilot scale ion exchange centrifugal partition chromatography purification of Sinalbin from white mustard seeds
IEEE Journal of Solid-state Circuits, 2009Co-Authors: Alix Toribio, Benoit Pinel, François De La Poype, Jean-marc Nuzillard, Leslie Boudesocque, Michel Lafosse, Jean-hugues RenaultAbstract:The purification of p-hydroxybenzylglucosinolate (Sinalbin) on a multigram scale from a crude aqueous extract of white mustard seeds (Sinapis alba var. concerta) was successfully achieved by scaling up a strong ion-exchange centrifugal partition chromatography (SIXCPC) laboratory procedure. Thus, the one-step Sinalbin purification was performed with 2.35 g of crude extract in approximately 170 min (830 mg/h) up to 70.3 g in approximately 160 min (26.3 g/h) by switching from a 200 mL laboratory scale column to a 5.7 L pilot-scale column. The required biphasic solvent system contained ethyl acetate, n-butanol, and water in 3:2:5 v/v/v proportions, Aliquat 336 (trioctylmethyl ammonium chloride) was added to the organic stationary phase (80 mM) and acted as ion-exchanger. Potassium iodide in the aqueous mobile phase (80 mM) was used as Sinalbin displacer. The 28.5 mass scale factor arose from the increase in mobile phase flow-rate (from 2 to 50 mL/min), from the higher mass of injected white mustard seed extract (from 12 to 350 g), and from the calculated productivity (from 830 mg to 26.3 g). These results demonstrate that industry scale production of glucosinolates is easily performed by SIXCPC, thus providing pure reference standards for pharmacology studies.
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strong ion exchange centrifugal partition chromatography as an efficient method for the large scale purification of glucosinolates
Journal of Chromatography A, 2007Co-Authors: Alix Toribio, Jean-marc Nuzillard, Jean-hugues RenaultAbstract:The glucosinolates Sinalbin and glucoraphanin were purified by strong ion-exchange displacement centrifugal partition chromatography (SIXCPC). The optimized conditions involved the biphasic solvent system ethyl acetate/n-butanol/water (3:2:5, v/v), the lipophilic anion-exchanger Aliquat 336 (trioctylmethylammonium chloride, 160 and 408 mM) and a sodium iodide solution (80 and 272 mM) as displacer. Amounts as high as 2.4 g of Sinalbin and 2.6g of glucoraphanin were obtained in one step in 2.5 and 3.5h respectively, starting from 12 and 25 g of mustard and broccoli seed aqueous extracts, using a laboratory scale CPC column (200 mL inner volume).
Niels Agerbirk - One of the best experts on this subject based on the ideXlab platform.
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Hydroxyl and Methoxyl Derivatives of Benzylglucosinolate in Lepidium densiflorum with Hydrolysis to Isothiocyanates and non-Isothiocyanate Products: Substitution Governs Product Type and Mass Spectral Fragmentation
2017Co-Authors: Eleonora Pagnotta, Niels Agerbirk, Carl E. Olsen, Luisa Ugolini, Susanna Cinti, Luca LazzeriAbstract:A system of benzylic glucosinolates was found and characterized in common pepperweed, Lepidium densiflorum Schrad. The major glucosinolate was the novel 4-hydroxy-3,5-dimethoxybenzylglucosinolate (3,5-dimethoxySinalbin), present at high levels in seeds, leaves, and roots. Medium-level glucosinolates were 3,4-dimethoxybenzylglucosinolate and 3,4,5-trimethoxybenzylglucosinolate. Minor glucosinolates included benzylglucosinolate, 3-hydroxy- and 3-methoxybenzylglucosinolate, 4-hydroxybenzylglucosinolate (Sinalbin), the novel 4-hydroxy-3-methoxybenzylglucosinolate (3-methoxySinalbin), and indole-type glucosinolates. A biosynthetic connection is suggested. NMR, UV, and ion trap MS/MS spectral data are reported, showing contrasting MS fragmentation of p-hydroxyls and p-methoxyls. Additional investigations by GC-MS focused on glucosinolate hydrolysis products. Whereas glucosinolates generally yielded isothiocyanates, the dominating 3,5-dimethoxySinalbin with a free p-hydroxyl group produced the corresponding alcohol and syringaldehyde (4-hydroxy-3,5-dimethoxybenzaldehyde). After thermal deactivation of the endogenous myrosinase enzyme, massive accumulation of the corresponding nitrile was detected. This case study points out how non-isothiocyanate glucosinolate hydrolysis products are prevalent in nature and of interest in both plant–pathogen interactions and human health
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variable glucosinolate profiles of cardamine pratensis brassicaceae with equal chromosome numbers
Journal of Agricultural and Food Chemistry, 2010Co-Authors: Niels Agerbirk, Carl Erik Olsen, Frances S Chew, Marian OrgaardAbstract:A novel glucosinolate, 3-(hydroxymethyl)pentylglucosinolate, was present at high levels in Cardamine pratensis L. from eastern North America and in commercially obtained seeds, but not in C. pratensis plants from southern Scandinavia. Glucosinolates in a number of accessions of C. pratensis included glucosinolates with the side chains 1-methylethyl, 1-(hydroxymethyl)ethyl, 1-methylpropyl, 1-(hydroxymethyl)propyl, 3-methylpentyl, 3-(hydroxymethyl)pentyl, benzyl, 4-hydroxybenzyl, 4-methoxybenzyl, indol-3-ylmethyl (as well as its 1-methoxy, 4-hydroxy, and 4-methoxy derivatives) and the rare side chain 1,4-dimethoxyindol-3-ylmethyl. Substantial variation was observed for four biosynthetic characters: (i) extent of chain elongation of Ile-derived glucosinolates; (ii) biosynthesis of Phe/Tyr-derived glucosinolates in general; (iii) hydroxylation of branched-chain glucosinolates; and (iv) O-methylation of 4-hydroxybenzylglucosinolate (Sinalbin). Cytological analysis of pollen mother cells and root tip cells in meiosis and mitosis established the chromosome number to be 2n = 30 for all accessions, irrespective of glucosinolate profile.
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host plant dependent metabolism of 4 hydroxybenzylglucosinolate in pieris rapae substrate specificity and effects of genetic modification and plant nitrile hydratase
Insect Biochemistry and Molecular Biology, 2007Co-Authors: Niels Agerbirk, Carl Erik Olsen, Henrik Bak Topbjerg, Jens Christian SorensenAbstract:After ingestion of transgenic Arabidopsis thaliana CYP79A1 containing Sinalbin (4-hydroxybenzylglucosinolate) due to genetic modification, only one major Sinalbin-derived sulphate ester (the sulphate ester of 4-hydroxyphenylacetonitrile) was excreted by Pieris rapae caterpillars (corresponding to 69 mol% of ingested Sinalbin). An additional sulphate ester (the sulphate ester of 4-hydroxyphenylacetamide) was excreted when the caterpillars were reared on two plant species (Sinapis alba and Sinapis arvensis) that contained Sinalbin naturally. Artificial addition of Sinalbin to S. arvensis leaves resulted in increased levels of the sulphated amide, and an enzymatic activity (nitrile hydratase) explaining the formation of the sulphated amide from Sinalbin was detected in both Sinapis species, but not in A. thaliana. In agreement with the suggested minor metabolic pathway, the caterpillars were able to sulphate 4-hydroxyphenylacetamide offered as part of an artificial diet. In fact, phenol and seven para-substituted phenol derivatives with substituents of moderate size were sulphated and excreted, but all tested phenols devoid of a nitrile functional group were less efficiently sulphated than the primary Sinalbin detoxification product, 4-hydroxyphenylacetonitrile. This suggests that the specificity of the sulphation step involved in Sinalbin metabolism may be adapted to nitriles formed as metabolites of phenolic glucosinolates. On the contrary, there was no specificity for products (4-hydroxybenzylascorbigen and 4-hydroxybenzylalcohol) derived from the semistable isothiocyanate produced from Sinalbin in the absence of nitrile specifier protein.
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lack of sequestration of host plant glucosinolates in pieris rapae and p grarricae
Chemoecology, 2003Co-Authors: Caroline Müller, Niels Agerbirk, Carl Erik OlsenAbstract:Sequestration of plant toxins in herbivores is often correlated with aposematic coloration and gregarious behaviour. Larvae of Pieris brassicae show these conspicuous morphological and behavioural characteristics and were thus suggested to sequester glucosinolates that are characteristic secondary metabolites of their host plants. P. rapaeare camouflaged and solitary, and are thus not expected to sequester. To test this hypothesis and to check the repeatabi-lity of a study that did report the presence of the glucosinolate sinigrin in P. brassicae, larvae were reared on three species of Brassicaceae (Sinapis alba, Brassica nigra and Barbarea stricta), and different leaf and insect samples were taken for glucosinolate analysis. The major host plant glucosinolates could only be found in traces or not at all in larval haemolymph, bled or starved larvae, faeces or pupae of both species or P. brassicae regurgitant. Haemolymph of both Pieris spp. was not rejected by the ant Myrmica rubra in dual-choice assays; the regurgitant of P. brassicae was rejected. This suggests the presence of compounds other than glucosinolates that might be sequestered in or produced by P. brassicae only. In faeces of both Pieris spp. a compound which yielded 4-hydroxybenzylcyanide (HBC) upon incubation with sulfatase was detected in high concentrations when larvae had been reared on S. alba. This compound may be derived from hydrolysis of Sinalbin, the main glucosinolate of that plant. The unidentified HBC progenitor was apparently not sequestered in the two Pieris spp., and was not detected in faeces of larvae reared on B. nigra or B. stricta.
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sequestration of host plant glucosinolates in the defensive hemolymph of the sawfly athalia rosae
Journal of Chemical Ecology, 2001Co-Authors: Caroline Müller, Carl Erik Olsen, Niels Agerbirk, Jeanluc Boeve, Urs Schaffner, Paul M BrakefieldAbstract:Interactions between insects and glucosinolate-containing plant species have been investigated for a long time. Although the glucosinolate–myrosinase system is believed to act as a defense mechanism against generalist herbivores and fungi, several specialist insects use these secondary metabolites for host plant finding and acceptance and can handle them physiologically. However, sequestration of glucosinolates in specialist herbivores has been less well studied. Larvae of the turnip sawfly Athalia rosae feed on several glucosinolate-containing plant species. When larvae are disturbed by antagonists, they release one or more small droplets of hemolymph from their integument. This “reflex bleeding” is used as a defense mechanism. Specific glucosinolate analysis, by conversion to desulfoglucosinolates and analysis of these by high-performance liquid chromatography coupled to diode array UV spectroscopy and mass spectrometry, revealed that larvae incorporate and concentrate the plant's characteristic glucosinolates from their hosts. Extracts of larvae that were reared on Sinapis alba contained Sinalbin, even when the larvae were first starved for 22 hr and, thus, had empty guts. Hemolymph was analyzed from larvae that were reared on either S. alba, Brassica nigra, or Barbarea stricta. Leaves were analyzed from the same plants the larvae had fed on. Sinalbin (from S. alba), sinigrin (B. nigra), or glucobarbarin and glucobrassicin (B. stricta) were present in leaves in concentrations less than 1 μmol/g fresh weight, while the same glucosinolates could be detected in the larvae's hemolymph in concentrations between 10 and 31 μmol/g fresh weight, except that glucobrassicin was present only as a trace. In larval feces, only trace amounts of glucosinolates (Sinalbin and sinigrin) could be detected. The glucosinolates were likewise found in freshly emerged adults, showing that the sequestered phytochemicals were transferred through the pupal stage.
Matthew J. Morra - One of the best experts on this subject based on the ideXlab platform.
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simultaneous quantification of sinigrin Sinalbin and anionic glucosinolate hydrolysis products in brassica juncea and sinapis alba seed extracts using ion chromatography
Journal of Agricultural and Food Chemistry, 2014Co-Authors: Inna E. Popova, Matthew J. MorraAbstract:Although mustards such as Sinapis alba and Brassica juncea contain glucosinolates (Sinalbin and sinigrin, respectively) that hydrolyze to form biopesticidal products, routine quality control methods to measure active ingredients in seed and seed meals are lacking. We present a simple and fast ion chromatography method for the simultaneous quantification of sinigrin, Sinalbin, and anionic hydrolysis products in mustard seed to assess biological potency. Optimum conditions include isocratic elution with 100 mM NaOH at a flow rate of 0.9 mL/min on a 4 × 210 mm hydroxide-selective anion-exchange column. All anion analytes including sinigrin, Sinalbin, SO42–, and SCN– yielded recoveries ranging from 83 to 102% and limits of detection ≤0.04 mM, with samples displaying little interference from plant matrix components. Sample preparation is minimized and analysis times are shortened to <90 min as compared with previous methods that took days and multiple instruments.
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sinigrin and Sinalbin quantification in mustard seed using high performance liquid chromatography time of flight mass spectrometry
Journal of Food Composition and Analysis, 2014Co-Authors: Inna E. Popova, Matthew J. MorraAbstract:Abstract Mustard crops such as Brassica juncea L. and Sinapis alba L. contain high concentrations of glucosinolates, that hydrolyze to form biologically active compounds that can impart flavors to foods, function as anticarcinogens in human nutrition, or can be used as broad spectrum antimicrobials to extend the shelf life of various food products. Most of the currently used methods for glucosinolate analysis in mustard prior to its utilization are time consuming and involve complicated sample preparation. We have developed and optimized a new method for glucosinolate quantification in mustard seed (B. juncea and S. alba) using HPLC with high-resolution time-of-flight mass spectrometry (TOF-MS) and compared it to two existing methods (HPLC–ultraviolet (UV) and ion chromatography (IC)). We also demonstrated the possibility of Sinalbin quantification using sinigrin calibration standards. The proposed HPLC–TOF-MS method is more sensitive than HPLC–UV and IC methods. Effects of the mustard seed matrix were negligible and virtually no sample preparation of the mustard seed extract was needed prior to analysis. For the developed HPLC–TOF–MS method, the overall analysis time including sample preparation was less than 1.5 h. The proposed HPLC–TOF-MS method is suitable for fast and simple real-time quantification of two major glucosinolates, Sinalbin and sinigrin, in mustard seed.
Carl Erik Olsen - One of the best experts on this subject based on the ideXlab platform.
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variable glucosinolate profiles of cardamine pratensis brassicaceae with equal chromosome numbers
Journal of Agricultural and Food Chemistry, 2010Co-Authors: Niels Agerbirk, Carl Erik Olsen, Frances S Chew, Marian OrgaardAbstract:A novel glucosinolate, 3-(hydroxymethyl)pentylglucosinolate, was present at high levels in Cardamine pratensis L. from eastern North America and in commercially obtained seeds, but not in C. pratensis plants from southern Scandinavia. Glucosinolates in a number of accessions of C. pratensis included glucosinolates with the side chains 1-methylethyl, 1-(hydroxymethyl)ethyl, 1-methylpropyl, 1-(hydroxymethyl)propyl, 3-methylpentyl, 3-(hydroxymethyl)pentyl, benzyl, 4-hydroxybenzyl, 4-methoxybenzyl, indol-3-ylmethyl (as well as its 1-methoxy, 4-hydroxy, and 4-methoxy derivatives) and the rare side chain 1,4-dimethoxyindol-3-ylmethyl. Substantial variation was observed for four biosynthetic characters: (i) extent of chain elongation of Ile-derived glucosinolates; (ii) biosynthesis of Phe/Tyr-derived glucosinolates in general; (iii) hydroxylation of branched-chain glucosinolates; and (iv) O-methylation of 4-hydroxybenzylglucosinolate (Sinalbin). Cytological analysis of pollen mother cells and root tip cells in meiosis and mitosis established the chromosome number to be 2n = 30 for all accessions, irrespective of glucosinolate profile.
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host plant dependent metabolism of 4 hydroxybenzylglucosinolate in pieris rapae substrate specificity and effects of genetic modification and plant nitrile hydratase
Insect Biochemistry and Molecular Biology, 2007Co-Authors: Niels Agerbirk, Carl Erik Olsen, Henrik Bak Topbjerg, Jens Christian SorensenAbstract:After ingestion of transgenic Arabidopsis thaliana CYP79A1 containing Sinalbin (4-hydroxybenzylglucosinolate) due to genetic modification, only one major Sinalbin-derived sulphate ester (the sulphate ester of 4-hydroxyphenylacetonitrile) was excreted by Pieris rapae caterpillars (corresponding to 69 mol% of ingested Sinalbin). An additional sulphate ester (the sulphate ester of 4-hydroxyphenylacetamide) was excreted when the caterpillars were reared on two plant species (Sinapis alba and Sinapis arvensis) that contained Sinalbin naturally. Artificial addition of Sinalbin to S. arvensis leaves resulted in increased levels of the sulphated amide, and an enzymatic activity (nitrile hydratase) explaining the formation of the sulphated amide from Sinalbin was detected in both Sinapis species, but not in A. thaliana. In agreement with the suggested minor metabolic pathway, the caterpillars were able to sulphate 4-hydroxyphenylacetamide offered as part of an artificial diet. In fact, phenol and seven para-substituted phenol derivatives with substituents of moderate size were sulphated and excreted, but all tested phenols devoid of a nitrile functional group were less efficiently sulphated than the primary Sinalbin detoxification product, 4-hydroxyphenylacetonitrile. This suggests that the specificity of the sulphation step involved in Sinalbin metabolism may be adapted to nitriles formed as metabolites of phenolic glucosinolates. On the contrary, there was no specificity for products (4-hydroxybenzylascorbigen and 4-hydroxybenzylalcohol) derived from the semistable isothiocyanate produced from Sinalbin in the absence of nitrile specifier protein.
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lack of sequestration of host plant glucosinolates in pieris rapae and p grarricae
Chemoecology, 2003Co-Authors: Caroline Müller, Niels Agerbirk, Carl Erik OlsenAbstract:Sequestration of plant toxins in herbivores is often correlated with aposematic coloration and gregarious behaviour. Larvae of Pieris brassicae show these conspicuous morphological and behavioural characteristics and were thus suggested to sequester glucosinolates that are characteristic secondary metabolites of their host plants. P. rapaeare camouflaged and solitary, and are thus not expected to sequester. To test this hypothesis and to check the repeatabi-lity of a study that did report the presence of the glucosinolate sinigrin in P. brassicae, larvae were reared on three species of Brassicaceae (Sinapis alba, Brassica nigra and Barbarea stricta), and different leaf and insect samples were taken for glucosinolate analysis. The major host plant glucosinolates could only be found in traces or not at all in larval haemolymph, bled or starved larvae, faeces or pupae of both species or P. brassicae regurgitant. Haemolymph of both Pieris spp. was not rejected by the ant Myrmica rubra in dual-choice assays; the regurgitant of P. brassicae was rejected. This suggests the presence of compounds other than glucosinolates that might be sequestered in or produced by P. brassicae only. In faeces of both Pieris spp. a compound which yielded 4-hydroxybenzylcyanide (HBC) upon incubation with sulfatase was detected in high concentrations when larvae had been reared on S. alba. This compound may be derived from hydrolysis of Sinalbin, the main glucosinolate of that plant. The unidentified HBC progenitor was apparently not sequestered in the two Pieris spp., and was not detected in faeces of larvae reared on B. nigra or B. stricta.
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sequestration of host plant glucosinolates in the defensive hemolymph of the sawfly athalia rosae
Journal of Chemical Ecology, 2001Co-Authors: Caroline Müller, Carl Erik Olsen, Niels Agerbirk, Jeanluc Boeve, Urs Schaffner, Paul M BrakefieldAbstract:Interactions between insects and glucosinolate-containing plant species have been investigated for a long time. Although the glucosinolate–myrosinase system is believed to act as a defense mechanism against generalist herbivores and fungi, several specialist insects use these secondary metabolites for host plant finding and acceptance and can handle them physiologically. However, sequestration of glucosinolates in specialist herbivores has been less well studied. Larvae of the turnip sawfly Athalia rosae feed on several glucosinolate-containing plant species. When larvae are disturbed by antagonists, they release one or more small droplets of hemolymph from their integument. This “reflex bleeding” is used as a defense mechanism. Specific glucosinolate analysis, by conversion to desulfoglucosinolates and analysis of these by high-performance liquid chromatography coupled to diode array UV spectroscopy and mass spectrometry, revealed that larvae incorporate and concentrate the plant's characteristic glucosinolates from their hosts. Extracts of larvae that were reared on Sinapis alba contained Sinalbin, even when the larvae were first starved for 22 hr and, thus, had empty guts. Hemolymph was analyzed from larvae that were reared on either S. alba, Brassica nigra, or Barbarea stricta. Leaves were analyzed from the same plants the larvae had fed on. Sinalbin (from S. alba), sinigrin (B. nigra), or glucobarbarin and glucobrassicin (B. stricta) were present in leaves in concentrations less than 1 μmol/g fresh weight, while the same glucosinolates could be detected in the larvae's hemolymph in concentrations between 10 and 31 μmol/g fresh weight, except that glucobrassicin was present only as a trace. In larval feces, only trace amounts of glucosinolates (Sinalbin and sinigrin) could be detected. The glucosinolates were likewise found in freshly emerged adults, showing that the sequestered phytochemicals were transferred through the pupal stage.