Masked mycotoxins: determination of a deoxynivalenol glucoside in artificially and naturally contaminated wheat by liquid chromatography-tandem mass spectrometry

Conjugated mycotoxins, in which the toxin is usually bound to a more polar substance like glucose, are referred to as masked mycotoxins, as these substances escape routine detection methods but can release their toxic precursors after hydrolysis. This is the first report on the natural occurrence of a glucoside of deoxynivalenol (DON) in Fusarium-infected wheat and maize. To obtain appropriate standards, we chemically synthesized deoxynivalenol-3-beta-D-glucopyranoside (DON-3-glucoside) and deoxynivalenol-15-beta-D-glucopyranoside (DON-15-glucoside). The synthesis products were characterized by liquid chromatography-tandem mass spectrometry. The DON-glucosides showed different collision-induced dissociation (CID) fragmentation behaviors and could therefore be distinguished. Wheat plants were either treated with DON (n = 52) or with Fusarium spp. (n = 4) at anthesis, and after harvest, wheat ears were analyzed for DON and DON-glucosides. All 56 treated wheat samples contained DON and a DON-glucoside with the same retention time, molecular mass, and CID fragmentation behavior as the synthetic DON-3-glucoside. Moreover, the DON-glucoside was also found in two out of three analyzed naturally DON-contaminated maize and in five out of five naturally contaminated wheat samples, in a range from 4 to 12% of the DON concentration. To further confirm the identity of the DON-glucoside, the compound was isolated from wheat extracts and characterized as DON-3-glucoside with NMR. The results of this study indicate the importance to consider both DON and DON-3-glucoside with regard to food and feed safety.     

Effects of milling and baking technologies on levels of deoxynivalenol and its masked form deoxynivalenol-3-glucoside

The co-occurrence of the major Fusarium mycotoxin deoxynivalenol (DON) and its conjugate deoxynivalenol-3-glucoside (DON-3-Glc) has been documented in infected wheat. This study reports on the fate of this masked DON within milling and baking technologies for the first time and compares its levels with those of the free parent toxin. The fractionation of DON-3-Glc and DON in milling fractions was similar, tested white flours contained only approximately 60% of their content in unprocessed wheat grains. No substantial changes of both target analytes occurred during the dough preparation process, i.e. kneading, fermentation, and proofing. However, when bakery improvers enzymes mixtures were employed as a dough ingredient, a distinct increase up to 145% of conjugated DON-3-Glc occurred in fermented dough. Some decrease of both DON-3-Glc and DON (10 and 13%, respectively, compared to fermented dough) took place during baking. Thermal degradation products of DON, namely norDON A, B, C, D, and DON-lactone were detected in roasted wheat samples and baked bread samples by means of UPLC-Orbitrap MS. Moreover, thermal degradation products derived from DON-3-Glc were detected and tentatively identified in heat-treated contaminated wheat and bread based on accurate mass measurement performed under the ultrahigh mass resolving power. These products, originating from DON-3-Glc through de-epoxidation and other structural changes in the seskviterpene cycle, were named norDON-3-Glc A, B, C, D, and DON-3-Glc-lactone analogically to DON degradation products. Most of these compounds were located in the crust of experimental breads.     

Ellagitannins,flavonoids, and other phenolics in red raspberries and their contribution to antioxidant capacity and vasorelaxation properties

Analysis of extracts of Glen Ample raspberries (Rubus idaeus L.) by gradient, reverse phase HPLC with diode array and tandem mass spectrometry identified eleven anthocyanins, including cyanidin-3-sophoroside, cyanidin-3-(2(G)-glucosylrutinoside), cyanidin-3-glucoside, cyanidin-3-rutinoside, pelargonidin-3-sophoroside, pelargonidin-3-(2(G)-glucosylrutinoside), and pelargonidin-3-glucoside. Significant quantities of an ellagitannin, sanguiin H-6, with an M(r) of 1870 were detected along with lower levels of a second ellagitannin, lambertianin C, which has an M(r) of 2804. Other phenolic compounds that were detected included trace levels of ellagic acid and its sugar conjugates along with one kaempferol- and four quercetin-based flavonol conjugates. Fractionation by preparative HPLC revealed that sanguiin H-6 was a major contributor to the antioxidant capacity of raspberries together with vitamin C and the anthocyanins. Vasodilation activity was restricted to fractions containing lambertianin C and sanguiin H-6.     

Spermidine and flavonoid conjugates from peanut (Arachis hypogaea) flowers

A new spermidine triamide derivative has been isolated from peanut flowers and identified as N (1)-acetyl- N (5), N (10)-di- p-( EE)-coumaroylspermidine on the basis of detailed analysis of NMR, MS, and UV data. Two other spermidine conjugates, N (1), N (5), N (10)-tri- p-( EEE)-coumaroylspermidine and di- p-( EE)-coumaroylspermidine, as well as four flavonoid conjugates (quercetin-3-glucoside, quercetin-3-glucuronide, isorhamnetin-3-glucoside, and isorhamnetin-3-glucuronide) that have been previously reported in organs of other plants, have been found in this study in peanut ( Arachis hypogaea L.), a representative of the Leguminosae family, for the first time. The dynamics of photoisomerization in the spermidine conjugates have been investigated.     

Confirmation of reduced toxicity of deoxynivalenol in extrusion-processed corn grits by the MTT bioassay

The objective of this study was to determine the loss of toxicity of deoxynivalenol in extruded cereal-based products by the tetrazolium salt (MTT) bioassay using a sensitive Chinese hamster ovary (CHO-K1) cell line and to compare the results to chemical (high-performance liquid chromatography, HPLC) and biochemical (enzyme-linked immunosorbant assay, ELISA) methods of analysis. A split-split plot design was used for the extrusion process experiments at temperatures of 150, 175, and 200 degrees C and screw speeds of 70 and 140 rpm. The initial mean deoxynivalenol concentration in the corn grits artificially contaminated with Fusarium graminearum was found to be 23.5 mug/g as measured by HPLC. The percent reductions of deoxynivalenol in the contaminated corn grits upon extrusion processing ranged from 22 to 35%, from 21 to 34%, and from 21 to 37% as measured by HPLC, ELISA, and MTT bioassay, respectively. The MTT bioassay results were more closely correlated with HPLC (r = 0.90) results than with ELISA results (r = 0.78). The MTT bioassay, using a sensitive mammalian cell line, was demonstrated to be a useful method for quantification of deoxynivalenol as well as a potential toxicity screening method for contaminated extruded cereal-based products.     

Deoxynivalenol, deoxynivalenol-3-glucoside, and enniatins: the major mycotoxins found in cereal-based products on the Czech market

Fusarium toxins, Alternaria toxins, and ergot alkaloids represent common groups of mycotoxins that can be found in cereals grown under temperate climatic conditions. Because most of them are chemically and thermally stable, these toxic fungal secondary metabolites might be transferred from grains into the final products. To get information on the commensurate contamination of various cereal-based products collected from the Czech retail market in 2010, the occurrence of "traditional" mycotoxins such as groups of A and B trichothecenes and zearalenone, less routinely determined Alternaria toxins (alternariol, alternariol monomethyl ether and altenuene), ergot alkaloids (ergosine, ergocryptine, ergocristine, and ergocornine) and "emerging" mycotoxins (enniatins A, A1, B, and B1 and beauvericin) were monitored. In a total 116 samples derived from white flour and mixed flour, breakfast cereals, snacks, and flour, only trichothecenes A and B and enniatins were found. Deoxynivalenol was detected in 75% of samples with concentrations ranging from 13 to 594 μg/kg, but its masked form, deoxynivalenol-3-β-d-glucoside, has an even higher incidence of 80% of samples, and concentrations ranging between 5 and 72 μg/kg were detected. Nivalenol was found only in three samples at levels of 30 μg/kg. For enniatins, all of the samples investigated were contaminated with at least one of four target enniatins. Enniatin A was detected in 97% of samples (concentration range of 20-2532 μg/kg) followed by enniatin B with an incidence in 91% of the samples (concentration range of 13-941 μg/kg) and enniatin B1 with an incidence of 80% in the samples tested (concentration range of 8-785 μg/kg). Enniatin A1 was found only in 44% of samples at levels ranging between 8 and 851 μg/kg.     

HPLC-DAD-MS/MS ESI characterization of unusual highly glycosylated acylated flavonoids from cauliflower (Brassica oleracea L. var. botrytis) agroindustrial byproducts

In a recent program to investigate the possible use of vegetable handling and commercialization wastes and residues as a source of potentially bioactive extracts, we have studied cauliflower byproducts. A total of twenty-eight compounds were characterized by HPLC-DAD-MS, of these, twenty-two are produced naturally by the plant. The main compounds found were kaempferol 3-diglucoside-7-glucoside and its combinations with different hydroxycinnamic acids. Moreover, some flavonoids with an unusual high grade of glycosylation have been isolated and tentatively identified for the first time. To our knowledge, the characterization of flavonoids with more than four sugars has not been previously reported. The new products isolated were tentatively identified as kaempferol 3-diglucoside-7-diglucoside, kaempferol 3-triglucoside-7-diglucoside, kaempferol 3-feruloyldiglucoside-7-diglucoside, kaempferol 3-sinapoyltriglucoside-7-glucoside, kaempferol 3-disinapoyltriglucoside-7-glucoside, kaempferol 3-sinapoyltriglucoside-7-diglucoside and kaempferol 3-disinapoyltriglucoside-7-diglucoside.     

Antiradical properties of red wine portisins

The free radical chemistry of two polyphenolic pigments from red wine, belonging to the family of portisins, has been investigated after reaction with O(2)(?-) radicals using electron paramagnetic resonance (EPR) spectroscopy. Two portisins derived from malvidin-3-glucoside and cyanidin-3-glucoside were used for this study. Stable free radicals were detected with both portisins and correspond to unpaired electrons localized on the B as well as F rings of the portisins. Interpretations were confirmed by comparison with the spectra of free radicals formed by oxidation of the model compounds cyanidin-3-glucoside, malvidin-3-glucoside, and catechin. These results concur with previous work reporting the higher antiradical properties of these pigments compared to their anthocyanin precursors.     

Effect of postharvest ultraviolet irradiation on resveratrol and other phenolics of cv. Napoleon table grapes

In the skin of cv. Napoleon table grapes, the anthocyanins malvidin 3-glucoside (and its acetyl and p-coumaroyl derivatives), cyanidin 3-glucoside, peonidin 3-glucoside, cyanidin 3-glucoside, petunidin 3-glucoside, and delphinidin 3-glucoside were identified by HPLC-DAD-MS. In addition, quercetin 3-glucoside and 3-glucuronide, caffeoyltartaric, piceid, and resveratrol were also detected. The content of most phenolics remained quite constant during postharvest refrigerated storage (10 days at 0 degrees C) while the resveratrol derivatives increased 2-fold. Postharvest treatments of grapes with UVC and UVB light induced a large increase in resveratrol derivatives (3- and 2-fold, respectively). This means that a serving of mature Napoleon grapes (200 g) provides approximately 1 mg of resveratrol, which is in the range of the amount supplied by a glass of red wine. This can be increased to 2 or 3 mg of resveratrol per serving in grapes that have been irradiated with UVB or UVC, respectively. These results show that refrigerated storage and UV irradiation of table grapes can be beneficial in terms of increasing the content of potentially health-promoting phenolics.     

Separation, identification, quantification, and method validation of anthocyanins in botanical supplement raw materials by HPLC and HPLC-MS

A method has been established and validated for identification and quantification of individual, as well as total, anthocyanins by HPLC and LC/ES-MS in botanical raw materials used in the herbal supplement industry. The anthocyanins were separated and identified on the basis of their respective M(+) (cation) using LC/ES-MS. Separated anthocyanins were individually calculated against one commercially available anthocyanin external standard (cyanidin-3-glucoside chloride) and expressed as its equivalents. Amounts of each anthocyanin calculated as external standard equivalent were then multiplied by a molecular-weight correction factor to afford their specific quantities. Experimental procedures and use of a molecular-weight correction factors are substantiated and validated using Balaton tart cherry and elderberry as templates. Cyanidin-3-glucoside chloride has been widely used in the botanical industry to calculate total anthocyanins. In our studies on tart cherry and elderberry, its use as external standard followed by use of molecular-weight correction factors should provide relatively accurate results for total anthocyanins, because of the presence of cyanidin as their major anthocyanidin backbone. The method proposed here is simple and has a direct sample preparation procedure without any solid-phase extraction. It enables selection and use of commercially available anthocyanins as external standards for quantification of specific anthocyanins in the sample matrix irrespective of their commercial availability as analytical standards. It can be used as a template and applied for similar quantification in several anthocyanin-containing raw materials for routine quality control procedures, thus providing consistency in analytical testing of botanical raw materials used for manufacturing efficacious and true-to-the-label nutritional supplements.     

Patterns of trichothecene production, genetic variability, and virulence to wheat of Fusarium graminearum from smallholder farms in Nepal

Fusarium graminearum causes wheat head blight and contaminates grain with the trichothecenes 4-deoxynivalenol and nivalenol. Sequence analysis of trichothecene genes indicates that nivalenol production is the ancestral trait; however, deoxynivalenol producers occur worldwide and predominate in North and South America and in Europe. Analysis of a large field population (>500 strains) from Nepal identified three groups that were both genetically distinct and polymorphic for trichothecene production: SCAR1 comprising 95% deoxynivalenol producers, SCAR2 comprising 94% nivalenol producers, and SCAR3/5 comprising 34% deoxynivalenol producers/63% nivalenol producers. The ability to cause wheat head blight differed between SCAR groups and trichothecene chemotypes: deoxynivalenol producers were more virulent than nivalenol producers across all three SCAR groups and within the SCAR3/5 genetic background. These data support the hypothesis that production of deoxynivalenol rather than nivalenol confers a selective advantage to this important wheat pathogen.     

Investigations on anthocyanins in wines from Vitis vinifera cv. pinotage: factors influencing the formation of pinotin A and its correlation with wine age

Pinotage red wines were found to contain a reaction product of malvidin 3-glucoside and caffeic acid, the so-called pinotin A. A total of 50 Pinotage wines from the vintages 1996-2002 were analyzed for the content of pinotin A, malvidin 3-glucoside, caffeic acid, and caftaric acid. Statistical analyses were performed to reveal variations in the content of these compounds and to determine the factors that influence pinotin A formation during wine aging. An exponential increase of the concentration of this aging product was observed with prolonged storage time. The most rapid synthesis of pinotin A was observed in 2.5-4 year old wines, although at this age malvidin 3-glucoside is already degraded to a large extent. This phenomenon is explained by the increased ratio of caffeic acid/malvidin 3-glucoside, which strongly favors the formation of pinotin A and makes side reactions less likely. Pinotin A formation proceeds as long as a certain level of malvidin 3-glucoside is maintained in the wines. In wines >5-6 years old degradation or polymerization of pinotin A finally exceeds the rate of its de novo synthesis.     

Ingested delphinidin-3-rutinoside is primarily excreted to urine as the intact form and to bile as the methylated form in rats

Many reports have described the bioavailability of anthocyanins; however, most of these reports investigated only the amount of anthocyanins excreted in urine. In the present study, we calculated the pharmacokinetic bioavailability of anthocyanins in rats by measuring the plasma concentration of delphinidin-3-rutinoside that had been administered orally or intravenously. Delphinidin-3-rutinoside was primarily absorbed in the blood and excreted into urine as unmetabolized forms with a T(max) of 26.3 min and a C(max) of 0.285 +/- 0.071 micromol/L. We detected small amounts of the metabolite 4'-O-methyl-delphinidin-3-rutinoside in the plasma, but we detected neither anthocyanidin (aglycone) nor glucuro- or sulfoconjugates. For the 8 h period after intake, delphinidin-3-rutinoside and 4'-O-methyl-delphinidin-3-rutinoside were excreted to urine at 795 +/- 375 and 12.3 +/- 2.91 nmol, respectively. Relative to intravenous injection, oral administration of delphinidin-3-rutinoside resulted in complete bioavailability (0.49 +/- 0.06%). Analysis of delphinidin-3-rutinoside plasma concentrations in bile cannulated rats revealed that, for the 8-h period after intake, the intact delphinidin-3-rutinoside excretion ratio in bile was 11% of the excretion ratio of 4'-O-methyl-delphinidin-3-rutinoside, 1.91 +/- 0.35 nmol versus 17.4 +/- 8.67 nmol, respectively. Setting the bile duct cannulation in a Bollman-type cage, however, significantly increased the bioavailability of orally administered delphinidin-3-rutinoside (18.14 +/- 6.24%). This effect appears to stem immobilization stress by reducing gastrointestinal motility. The cumulative excretion of delphinidin-3-rutinoside and 4'-O-methyl-delphinidin-3-rutinoside in urine and bile was 2.67 +/- 1.24% (w/w) of the dose ingested. Studies report that several metabolites are formed after oral ingestion of anthocyanins. Examples include glucuronyl from cyanidin-3-glucoside and both glucuronyl and sulfate conjugates from pelargonidin-3-glucoside. Our results indicate that delphinidin-3-rutinoside might be metabolized differently from cyanidin-3-glucoside and pelargonidin-3-glucoside.     

Enzymatic synthesis of [3'-O-methyl-(3)H]malvidin-3-glucoside from petunidin-3-glucoside

Malvidin-3-glucoside has been labeled by enzymatic synthesis in a single-step experiment. Catechol-O-methyl transferase catalyzed the B-ring methylation of petunidin-3-glucoside, and S-Adenosyl-L-[methyl-(3)H] methionine was the methyl donor. Solid phase extraction and preparative high-performance liquid chromatography were necessary to separate [3'-O-methyl-(3)H]malvidin-3-glucoside from an isomer and the starting material. The specific activity was 2.2 Ci mmol(-1), and the yield of incorporation was 1.1%. A possible application of the labeled material is the study of anthocyanin reactions in complex mixtures such as red wine where products are difficult to isolate and analyze.     

Isolation and determination of anthocyanins in seed coats of black soybean (Glycine max (L.) Merr.).

Anthocyanin pigments in seed coats of black soybean (Glycine max (L.) Merr.) were extracted with 1% HCl-CH(3)OH, and the crude anthocyanin extract was purified by Shepadex LH-20 and Lichroprep RP-18 open-column chromatography. Three major anthocyanins were isolated, and their chemical structures were identified by spectroscopic methods (UV-visible, FABMS, (1)H and (13)C NMR, and by TLC). The complete structures of these anthocyanins were elucidated as delphinidin-3-glucoside, cyanidin-3-glucoside, and petunidin-3-glucoside. Among them, petunidin-3-glucoside was identified as a new anthocyanin in black soybeans. On the basis of RP-HPLC with a UV-vis detector, the contents of delphinidin-3-glucoside, cyanidin-3-glucoside, petunidin-3-glucoside, and total anthocyanins in seed coats of 10 black soybeans were found in the ranges of 0-3.71, 0.94-15.98, 0-1.41, and 1.58-20.18 mg/g, respectively. The results obtained in this study imply that the seed coats of black soybean can be used as a good source for cyanidin-3-glucoside and delphinidin-3-glucoside.     

Black rice (Oryza sativa L. indica) pigmented fraction suppresses both reactive oxygen species and nitric oxide in chemical and biological model systems

Anthocyanins, present in fruits and vegetables as natural colorants, have been well characterized to possess bioactive properties. Anthocyanin components extracted from black rice (Oryza sativa L. indica) separated by gel filtration and identified using LC-MS were cyanidin 3-glucoside and peonidin 3-glucoside. A standardized extract of black rice pigmented fraction (BRE) containing known proportions of cyanidin 3-glucoside and peonidin 3-glucoside exhibited marked antioxidant activities and free radical scavenging capacities in a battery of in vitro model systems. Significant (p < 0.05) prevention of supercoiled DNA strand scission induced by reactive oxygen species (specifically, peroxyl radical and hydroxyl radicals) and suppression of the oxidative modification of human low-density lipoprotein was obtained with BRE. In addition, BRE reduced (p < 0.05) the formation of nitric oxide by suppressing inducible nitric oxide synthase expression in murine macrophage RAW264.7 cells, without introducing cell toxicity. The results of this study show that black rice contains anthocyanin pigments with notable antioxidant and anti-inflammatory properties for potential use in nutraceutical or functional food formulations.     

Determination of anthocyanins from camu-camu (Myrciaria dubia) by HPLC-PDA, HPLC-MS, and NMR

Camu-camu [Myrciaria dubia (HBK) McVaugh] is a small fruit native to the Amazonian rain forest. Its anthocyanin profile has now been investigated for the first time. Fruits from two different regions of the S?o Paulo state, Brazil, were analyzed. The major anthocyanins were isolated by high-speed countercurrent chromatography. HPLC-PDA, HPLC-MS/MS, and 1H NMR were used to confirm the identity of the main anthocyanins of camu-camu. Cyanidin-3-glucoside was identified as the major pigment in the fruits from both regions, representing 89.5% in the fruits produced in Iguape and 88.0% in those from Mirandópolis, followed by the delphinidin-3-glucoside, ranging between 4.2 and 5.1%, respectively. Higher total anthocyanin contents were detected in the fruits from Iguape (54.0 +/- 25.9 mg/100 g) compared to those from Mirandópolis (30.3 +/- 6.8 mg/100 g), most likely because of the lower temperatures in the Iguape region.     

Anthocyanin profile of Korean cultivated kidney bean (Phaseolus vulgaris L.)

This investigation was conducted to determine the structures and amounts of anthocyanins obtained from seed coats of kidney bean (Phaseolus vulgaris L.) cultivated in Korea. Anthocyanins in the seed coat of kidney bean were extracted with 1% HCl/20% CH(3)OH, and the crude anthocyanin extracts were purified by semipreparative HPLC. Five major anthocyanins were isolated, and their chemical structures were identified by spectroscopic methods (UV-vis, LC/ES-MS, and 1H and 13C NMR). The structures of these five anthocyanins were elucidated as cyanidin 3,5-diglucoside, delphinidin 3-glucoside, cyanidin 3-glucoside, petunidin 3-glucoside, and pelargonidin 3-glucoside. Using RP-HPLC with photodiode array detection, each of the five anthocyanins was separated within 12 min by using a gradient elution. It was proved that the application of RP-HPLC could be an excellent method for determining the composition and contents of anthocyanins in kidney bean. The preponderance of pelargonidin 3-glucoside and delphinidin 3-glucoside are observed in red and black kidney beans, respectively. However, in this study, it is reported for the first time that the contents and composition of anthocyanins in speckled seed depend on the classes of speckle color. The contents of cyanidin 3,5-diglucoside, delphinidin 3-glucoside, cyanidin 3-glucoside, petunidin 3-glucoside, pelargonidin 3-glucoside, and total anthocyanins in seed coats of 16 kidney beans cultivated in Korea were in the ranges of 0-0.04, 0-2.61, 0-0.12, 0-0.17, 0-0.59 and 0-2.78 mg/g of dried seed coats, respectively.     

Anthocyanin composition in black, blue, pink, purple, and red cereal grains

Anthocyanin pigments from a wide variety of edible and ornamental black, blue, pink, purple, red, and white wheat, barley, corn, rice, and wild rice were identified and quantified to evaluate their potential as natural colorants or functional food ingredients. The total anthocyanin contents varied significantly and exhibited a range of 7-3276 microg/g. Some grains, such as red rice and black rice, contained a limited number of pigments, whereas others, such as blue, pink, purple, and red corns, had complex anthocyanin profiles. Of the 42 anthocyanin compounds observed, 9 were characterized by comparison of the spectroscopic and chromatographic properties with those of authentic standards. The remaining compounds were tentatively identified on the basis of spectroscopic properties and electrospray ionization mass spectra. The most abundant anthocyanins were cyanidin 3-glucoside in black and red rices and in blue, purple, and red corns, pelargonidin 3-glucoside in pink corn, and delphinidin 3-glucoside in blue wheat.     

Phenolic compounds, carotenoids, anthocyanins, and antioxidant capacity of colored maize (Zea mays L.) kernels

In this study, the contents of total phenolics, flavonoids, anthocyanins, β-carotene, and lutein as well as free, conjugated, and insoluble bound phenolic acids were determined in whole kernels of 10 different colored maize genotypes. In addition, the antioxidant activity was evaluated as radical scavenging activity with ABTS (2,2-azino-bis/3-ethil-benothiazoline-6-sulfonic acid) and DPPH (2,2-diphenyl-1-picrylhydrazyl) reagents. Generally, considerable differences in phytochemical contents and antioxidant capacity were observed between the genotypes. The β-carotene and lutein contents ranged from 0 to 2.42 mg/kg d.m. and from 0 to 13.89 mg/kg d.m., respectively, whereas the total anthocyanin contents of anthocyanin-rich colored maize genotypes ranged from 2.50 to 696.07 mg CGE/kg d.m. (cyanidin 3-glucoside equivalent) with cyanidin 3-glucoside (Cy-3-Glu) as the most dominant form. The light blue ZPP-2 selfed maize genotype has a higher content of total phenolics, flavonoids, and ferulic acid as compared to other tested maize and the highest ABTS radical scavenging activity.