Article pubs.acs.org/JAFC
Allium Discoloration: Color Compounds Formed during Greening of Processed Garlic Roman Kubec,*,† Petr Curko,† Petra Urajová,† Josep Rubert,‡ and Jana HajšlovᇠDepartment of Applied Chemistry, University of South Bohemia, Branišovská 31, 370 05 Č eské Budějovice, Czech Republic Department of Food Analysis and Nutrition, Institute of Chemical Technology, Technická 5, 166 28 Prague 6, Czech Republic
† ‡
S Supporting Information *
ABSTRACT: Structures and formation pathways of compounds responsible for blue−green discoloration of processed garlic were studied in model systems. A procedure was developed for isolation of the color compounds and their tentative identification by high-performance liquid chromatography coupled to a diode array detector and tandem mass spectrometry. It was found that the pigment is a mixture of numerous pyrrole-based purple/blue and yellow species. Experiments with isotope-labeled precursors revealed that two molecules of an amino acid are involved in the formation of each color compound. In the purple/blue species (λmax = 565−600 nm), both amino acid molecules are incorporated into two 3,4-dimethylpyrrole-derived rings linked together by a propenylidine bridge. On the other hand, the yellow compounds (λmax = 420−450 nm) contain only one N-substituted 3,4dimethylpyrrole ring, to which the second amino acid is bound via a propenylidine side chain. KEYWORDS: discoloration, plant pigment, greening, garlic, Allium sativum, isoalliin, alliin, pyrrole, HPLC−DAD−MS/MS
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INTRODUCTION The mincing, crushing, slicing, pureeing, or other mechanical disruption of garlic, onion, and leek often lead to the formation of intensely colored compounds. Processed garlic tends to turn vividly green or blue, while onion and leek may acquire a pink or red discoloration (Figure 1). The formation of these
sulfoxide (isoalliin, 1) and other S-substituted cysteine derivatives (e.g., 2 and 3) that gives rise to a great number of various sulfur compounds. Some of them, namely, 1-propenylcontaining thiosulfinates and cepathiolanes (referred to as “color developers”), subsequently react with amino acids to yield N-substituted 3,4-dimethylpyrroles (called “pigment precursors”). These colorless pyrroles further react with various (thio)carbonyl species to produce the color compounds.1−13 It is known that these color compounds can also be formed under acidic conditions independent of the action of alliinase.14 For example, intensely green “Laba” garlic (a delicacy consumed during celebrations of the Chinese New Year) is prepared by soaking intact peeled cloves in vinegar for several weeks.15,16 It has been reported by several research groups that the green color of processed garlic is a consequence of parallel formation of purple/blue and yellow species.6,7 However, exact structures of these color compounds are still very poorly understood. In fact, only a single purple compound (PUR-1; λmax = 570 nm) related to garlic greening has thus far been fully characterized by spectroscopic methods [nuclear magnetic resonance (NMR) and mass spectrometry (MS)].4 On the other hand, a series of yellow species (λmax = 401−437 nm) generated in various model systems were identified only tentatively on the basis of MS data (Figure 2).17−19 This paper describes our investigations into the color compounds formed during greening of processed garlic. A better knowledge about these compounds may help to find effective ways to prevent this undesirable phenomenon. As a result of the overwhelming complexity of reactions taking place
Figure 1. Discoloration of homogenized garlic and onion stored at 23 °C.
compounds represents a serious issue for companies selling processed garlic/onion/leek products because consumers often tend to associate these color changes with spoilage. The green color of processed garlic develops much faster (typically within several hours) compared to the pink discoloration of onion/leek. Upon prolonged storage, the green color gradually fades, turning into brownish tones after several days. Despite the obvious visual difference in the colors formed, the greening of garlic and pinking of onion/leek were shown to be very closely related. Both processes are triggered by the alliinase-mediated cleavage of S-(1-propenyl)cysteine © 2017 American Chemical Society
Received: Revised: Accepted: Published: 10615
October 4, 2017 November 1, 2017 November 5, 2017 November 6, 2017 DOI: 10.1021/acs.jafc.7b04609 J. Agric. Food Chem. 2017, 65, 10615−10620
Article
Journal of Agricultural and Food Chemistry
Pigment Purification. The procedure developed in our previous paper13 was followed. Model solutions were cooled to room temperature, filtered, and loaded onto a C-18 solid-phase extraction (SPE) column (60 mL/10 g, Supelco). The column was washed successively with 50 mL of the following solvents: H2O, acetone, pentane, and CH2Cl2. These fractions (usually colorless) were discarded. Finally, the color compounds were eluted with CH3OH (50 mL). The solvent was removed at a reduced pressure (
