Influence of Fermentation Conditions on Glucosinolates, Ascorbigen, and Ascorbic Acid Content in White Cabbage (Brassica oleracea var. capitata cv. Taler) Cultivated in Different Seasons

Authors

  • C. Martinez-Villaluenga,

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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  • E. Peñas,

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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  • J. Frias,

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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  • E. Ciska,

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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  • J. Honke,

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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  • M.K. Piskula,

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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  • H. Kozlowska,

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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  • C. Vidal-Valverde

    1. Authors Martinez-Villaluenga, Peñas, Frias, and Vidal-Valverde are with Instituto de Fermentaciones (CSIC), Juan de la Cierva 3, 28006 Madrid, Spain, and authors Ciska, Honke, Piskula, and Kozlowska are with Division of Food Science, Institute of Animal Reproduction and Food Research of Polish Academy of Sciences, Tuwima 10, P.O. Box 55, 10-747 Olsztyn 5, Poland. Direct inquiries to author Vidal-Valverde (E-mail: ificv12@ifi.csic.es).
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Abstract

ABSTRACT:  The content of glucosinolates (GLS), ascorbigen, and ascorbic acid in white cabbage (Brassica oleracea var. capitata cv. Taler) cultivated in different seasons (summer and winter) was determined, before and after spontaneous and starter-induced fermentation. Different salt concentrations (0.5% NaCl or 1.5% NaCl) were used for sauerkraut production. Glucoiberin, sinigrin, and glucobrassicin were dominating in raw white cabbage cultivated either in winter or summer seasons. Ascorbigen precursor, glucobrassicin, was found higher in cabbage cultivated in winter (2.54 μmol/g dw) than those grown in summer (1.83 μmol/g dw). Cabbage fermented for 7 d was found to contain only traces of some GLS irrespective of the fermentation conditions used. Ascorbigen synthesis occurred during white cabbage fermentation. Brining cabbage at low salt concentration (0.5% NaCl) improved ascorbigen content in sauerkraut after 7 d of fermentation at 25 °C. The highest ascorbigen concentration was observed in low-sodium (0.5% NaCl) sauerkraut produced from cabbage cultivated in winter submitted to either natural (109.0 μmol/100 g dw) or starter-induced fermentation (108.3 and 104.6 μmol/100 g dw in cabbages fermented by L. plantarum and L. mesenteroides, respectively). Ascorbic acid content was found higher in cabbage cultivated in summer and fermentation process led to significant reductions. Therefore, the selection of cabbages with high glucobrassicin content and the production of low-sodium sauerkrauts may provide enhanced health benefits towards prevention of chronic diseases.

Introduction

Many epidemiological studies have shown an inverse relationship between Brassica vegetable consumption and cancer risk (Zhang and others 2006; Higdon and others 2007). The beneficial effects on human health of cruciferous vegetables have been attributed to the presence of high levels of phytochemicals termed glucosinolates (GLS) (Moreno and others 2006; Higdon and others 2007). GLS are sulfur-containing glucosides, which are classified into 3 different classes: aliphatic GLS, indole GLS, and aromatic GLS. The amount of GLS in plants is influenced by the growth season (Rosa and others 1996; Ciska and others 2000). When the plant tissue is damaged, either by handling, processing, or chewing, GLS are broken down by the endogenous enzyme myrosinase (β-thioglucoside glucohydrolase; EC 3.2.3.1.) to release a complex variety of breakdown products (Mithen and others 2000).

At neutral pH, indole GLS yield first an unstable isothiocianate that further degrades with loss of S to form indole-3-carbinol (I3C), ascorbigen, and related compounds, which are known to be anticarcinogens, and may play a protective role against cancer in the diet (Higdon and others 2007). The prevailing mechanism for the protective activity of these compounds has been considered to be the induction of phase 2 enzymes, which are involved in detoxification of xenobiotic compounds (Talalay and others 1995). Other health-promoting effects of isothiocyanates are related to their antioxidant properties (Fahey and Talalay 1999; Barillari and others 2005), their capacity for inducing apoptosis (programmed cell death) in cancer cells (Kuang and Chen 2004; Rose and others 2005).

The most common product among the fermented white cabbage (Brassica oleracea var. capitata) is sauerkraut, which results from lactic acid fermentation of shredded and salted white cabbage. During shredding, glucobrassicin (GB) is transformed into I3C by the action of myrosinase and during fermentation, as the pH decrease, this indole reacts nonenzymatically with L-ascorbic acid to yield ascorbigen (Aleksandrova and others 1992). Some reaserchers reported that ascorbigen might be the dominant end product of indole GLS in sauerkraut (Agerbirk and others 1998; Ciska and Pathak 2004). Several in vivo studies have reported the anticarcinogenic properties of ascorbigen against colon and mammary tumors in rodents (Mukanov 1984; Sepkovic and others 1994). Moreover, ascorbigen is thought to be responsible for the anticarcinogenic properties in humans with a high intake in white cabbage (Lysenkova and others 2001; Smiechowska and others 2008).

Sauerkraut natural fermentation is a process with a definite sequence of microorganisms. Starter cultures have been proposed for sauerkraut production to minimize the variability in food quality and potentially reduce the amount of salt required for fermentation (Viander and others 2003). Sauerkraut usually contains between 0.6% and 2% NaCl and the amount used depends on consumer demands and on traditional considerations in the producing countries (Holzapfel and others 2003). The general trend by consumers in industrialized countries is to reduce the salt content of foods. The ability to reduce the salt required for fermentation would have the advantage of reducing the liquid brine and, therefore, an increase in yield of sauerkraut, as it has been reported by Johanningsmeier and others (2007).

Ascorbigen concentrations in sauerkraut may depend on factors including GLS and ascorbic acid content in white cabbage. Moreover, there is a lack of information about the effect of different fermentation conditions on ascorbigen content in sauerkraut. Therefore, the present study shows the effect of fermentation conditions on GLS, ascorbigen, and ascorbic acid content in white cabbage (Brassica oleracea var. capitata cv. Taler) cultivated in winter and summer. Natural and induced fermentations using starter cultures were performed by adding 0.5% and 1.5% NaCl.

Materials and Methods

Material

Commercial white cabbages (Brassica oleracea L. var. capitata cv. Taler), cultivated in winter and summer, were purchased in a local supermarket in September and March, respectively. Fresh cabbages were stored for less than 2 d at 4 °C until further analysis.

Fermentation process

A random representative selection of the cabbages was chosen and stripped of their outer leaves and divided in 4 similar pieces; the central core was removed, and the pieces were shredded to a 2-mm width size using a domestic shredder. Raw shredded white cabbage was sampled and used as reference to unprocessed material. Different salt concentrations were added (0.5% or 1.5% NaCl) into shredded cabbage and mixed thoroughly. L. plantarum (CECT 748) and L. mesenteroides (CECT 219) strains were obtained from the Spanish Type Culture Collection (CECT, Valencia, Spain) and were grown in MRS broth (Difco Lab., Detroit, Mich., U.S.A.) for 18 h at 30 °C. The cells were washed twice in sterile saline solution (0.9% NaCl) and used as starter cultures. Shredded and salted cabbage was fermented spontaneously by the microorganisms present naturally on the cabbage surface (natural fermentation) or by using starter cultures inoculated at approximately 106 colony forming units (CFU)/g of cabbage (induced fermentation). Cabbage and brine were transferred to fermentation vessels (1.5 L) and pressed to remove air bubbles. Fermentation was carried out in 3 parallel batches at room temperature (22 to 25 °C) for 7 d. On the 3rd day of the fermentation process, cabbage was pricked to release fermentation gasses. Raw and fermented cabbage were freeze-dried, milled, and stored at −20 °C until further analysis.

Analysis of GLS

Individual GLS were analyzed as described previously by Ciska and others (2000). GLS were extracted from raw cabbage according to the method reported in the Official Journal European Communities (1990). Briefly, duplicate 200 mg of freeze-dried samples were extracted 3 times with boiling 70% methanol. Because all the cultivars examined lacked glucotropaeolin (Merck, Darmstadt, Germany), its known amount was added to each sample just before the 1st extraction as an internal standard for the high-performance liquid chromatography (HPLC) analysis. The isolation, desulfatation, and HPLC analysis of GLS were carried out according to a modified version of the method of Heaney and others (1986). Desulfo-GLS were separated in the HPLC system (Shimadzu, Kyoto, Japan) with an autoinjector (20 μL loop), a Spherisorb ODS-2 3 Micron column (150 × 4.6 mm), and 1.2 mL/min flow rate at 32 °C by eluting with a gradient of water (A) and 20% acetonitrile (B) as follows: isocratically 1% B for 1 min, linear gradient to 99% B for 30 min (curve 3), isocratically 99% B for 6 min, linear gradient to 1% B for 5 min, 1% B for 8 min. Desulfo-GLS were detected at λ= 229 nm. The sample content of GLS was quantified on the basis of the internal standard and relevant relative response factors (Official Journal European Communities 1990).

Analysis of ascorbigen

The content of ascorbigen in fermented cabbages was carried out as in Ciska and Pathak (2004) with some modifications. Briefly, 1 g of freeze-dried cabbage was homogenized with 15 mL of distilled water and 1 g of NaCl using and Ultra Turrax homogenizer T-25 digital (IKA® KE GmbH & Co.KG, Stanten, Germany). Further, homogenates were extracted twice with 15 mL acetone (Carlo Erba, Rodano, Italy). Acetone extracts were combined and, after filtration, concentrated to 3 mL total volume. The concentrate was extracted twice with 15 mL of ethyl acetate (Fluka, Buchs, Switzerland). The combined organic layers were dried over anhydrous sodium sulfate (Panreac, Barcelona, Spain), filtered, and evaporated under vacuum to dryness. The residue was dissolved in acetonitrile (LabScan, Gliwice, Poland) and made up to 2 mL with 0.1 M ammonium acetate, pH 5.7 as mobile phase composition.

Quantification of ascorbigen was performed by HPLC using an Alliance Separation Module 2695 (Waters, Milford, U.S.A.), a Photodiode Array detector 996 at 280 nm (Waters), and a personal computer running the Empower 2 for Microsoft Windows chromatographic software (Waters). The sample (20 μL) was injected into an ODS-2 column 150 × 4.6 mm i.d., 5 μm size column (Waters) at 30 °C. The chromatogram was developed at a flow rate of 1.2 mL/min by eluting in a gradient of mobile phase A (0.1 M ammonium acetate, pH 5.7 containing 10% acetonitrile) and mobile phase B (0.1 M ammonium acetate, pH 5.7 containing 80% acetonitrile) as follows: linear gradient of 100% A – 100% B for 25 min, isocratic 100% B for 5 min, linear gradient of 100% B – 100% A for 5 min, equilibrate for 5 min.

For identification and quantification ascorbigen standard was used. Ascorbigen standard was synthesized following the procedure described by Kiss and Neukom (1966). Reactions were run at room temperature, pH 4, and a substrate molar ratio of 1: 1. After duplicate extraction with diethyl ether, the reaction mixture was re-extracted with ethyl acetate. The ethyl acetate extract was dried over anhydrous sodium sulfate and evaporated under vacuum. Ascorbigen purity determined by HPLC reached 99.6%.

Analysis of ascorbic acid

Ascorbic acid content in raw and fermented cabbage was determined by capillary electrophoresis (CE) using a P/ACE system 2050 (Beckman Instruments, Fullerton, Calif., U.S.A.) and UV detection at 254 nm (Frias and others 2005). Briefly, 1 g of freeze-dried cabbage was extracted with 20 mL metaphosphoric acid (Sigma Aldrich, Steinheim, Germany) and 100 μL isoascorbic acid 0.6 mg/mL were added as internal standard (Fluka, Steinheim, Germany). An aliquot of extract was treated with D,L-dithiothreitol (Sigma Aldrich) to convert any dehydroascorbic acid present in the sample to ascorbic acid (Sapers and others 1990). Extractions were carried out in triplicate. Ascorbic acid was quantified from a calibration curve built with pure ascorbic acid standard (Merck, Darmstadt, Germany), and with the response factor relative to internal standard.

Statistical analysis

Data were expressed as mean ± standard deviation and subjected to one-way analysis of variance (ANOVA) using the Fischer LSD test with the Statgraphic 5.0 Program (Statistical Graphic, Rockville, Md., U.S.A.) for Windows.

Results and Discussion

Influence of growth season and fermentation conditions on GLS content of white cabbage

Individual and total GLS content in raw white cabbages cultivated in winter and summer are presented in Table 1. Aliphatic GLS were found in the highest absolute values comprising 72% and 87% in white cabbage cultivated in winter and summer, respectively. Our results show that sinigrin was the most abundant GLS in the white cabbage cultivated in winter, constituting 31% of total GLS content. However, glucoiberin was the major GLS in the white cabbage cultivated in summer constituting 48% of total GLS content. Glucobrassicin (GB) (3-indolylmethylglucosinolate) was the most abundant indole GLS, constituting 12% and 24% of total GLS for summer and winter cabbage, respectively. Concentrations of glucoraphanin, glucoibervirin, glucobrassicanapin, glucoerucin, glucorasnutin, 4-OH-glucobrassicin, 4-MeO-glucobrassicin, and neoglucobrassicin were never higher than 1 μmol/g DM either in white cabbage cultivated in winter or summer. Cabbage cultivated in summer was characterized with higher total GLS content than cabbage cultivated in winter.

Table 1—.  Glucosinolate content in Brassica olereacea var. capitata cv. Taler cultivated in different seasons.A
SeasonsAliphaticIndoleTotal GLS
IBaPROGbSINcGRAPdNAPeIBERfBNAPgERUhNASRi4-OH-GBjGBk4-met-GBlneo-GBm
  1. AMean values are expressed in μmol/g dw; standard deviations are indicated in brackets. ND = non detected; trace ≤ 0.05 μmol/g.

  2. aGlucoiberin, bProgoitrin, cSinigrin, dGlucoraphanin, eGluconapin, flucoibervirin, gGlucobrassicanapin, hGlucoerucin, iGluconasrutin, j4-Hydoxyglucobrassicin, kGlucobrassicin, l4-Methoxy-glucobrassicin, and mNeo-glucobrassicin.

Winter2.621.133.350.100.33trace0.06trace0.08trace2.540.450.0810.74
(0.08)(0.01)(0.00)(0.01)(0.00) (0.00) (0.00) (0.02)(0.02)(0.00) 
Summer7.570.125.720.140.05tracetracetraceNDtrace1.830.170.0515.66
(0.07)(0.00)(004)(0.00)(0.00)     (0.00)(0.01)(0.00) 

The observed GLS profile is in agreement with previous results (Wold 2004; Rungapamestry and others 2006; Volden and others 2008). Unlike results shown in the present study, sinigrin has been reported as the major glucosinolate in edible cabbage (Kushad and others 1999; Kushad and others 2004; Charron and others 2005), with values ranging from 6 μmol/g DM (Kushad and others 1999) to 125 μmol/g DM (Tookey and others 1980) depending upon the cultivar and the tissue assessed. Yet, cabbage cultivars reported as having progoitrin as the dominating glucosinolate are also frequent (Kushad and others 2004; Tian and others 2005). Differences in GLS profiles may occur between species, cultivars of a single species, and between tissues of a single plant (Bellostas and others 2007). The total GLS levels (average 13.15 μmol/g DM, equivalent to 131.5/100 g FW) determined in white cabbage correspond well with the findings by others (Ciska and Kozłowska 2001) who found 73.3 and 112 μmol/100 g of FW, respectively. However, Rosa and Heaney (1993) found 335 μmol/100 g of FW of total GLS, Volden and others (2008) reported 340 μmol/100 g FW of total GLS, and Wold (2004) determined the average levels of total GLS in the identical cultivar (“Bartolo”) during 2 y on average to be 388 μmol/100 g of FW. Variations in cultivars, growing conditions, and environmental factors are likely to cause the observed deviations as reported previously (Ciska and others 2000; Vallejo and others 2003a). Moreover, Pereira and others (2002) evaluated the effect of temperature on the GLS content of broccoli sprouts and found that high temperatures lead to higher GLS levels than low temperatures, results consistent with the higher content of GLS in summer cabbage than in winter cabbage found in the present study.

Total GLS concentration was sharply decreased and only traces of individual GLS were detected after fermentation for 7 d irrespective of the conditions used for sauerkraut production (data not shown). Our results are in agreement with previous studies showing a gradual decrease of GLS to zero levels after fermentation of white cabbage (Taipale and others 2000; Tolonen and others 2002). Tolonen and others (2002) concluded that GLS are decomposed during cabbage fermentation process to form several potentially beneficial breakdown products such as Isothiocyanates, I3C, and sulforaphane. Ascorbigen was not determined although some researchers suggested that is the dominant end product of indol GLS (Agerbirk and others 1998; Ciska and Pathak 2004).

Influence of growth season and fermentation conditions on ascorbigen content of cabbage

Table 2 shows the effect of fermentation conditions on ascorbigen content in white cabbage (Brassica oleracea var. capitata cv. Taler) cultivated in winter and summer. Ascorbigen content in shredded cabbage was found at low concentrations reaching 6 and 8 μmol/100 g dw when cabbage was cultivated in winter and summer, respectively. However, ascorbigen concentration increased up to 20-fold in sauerkraut. Growth season of cabbage and fermentation conditions used for sauerkraut production had an impact on ascorbigen content. The highest ascorbigen concentration (ranging from 104.6 to 109.0 μmol/100 g dw) was observed in sauerkraut produced from cabbage cultivated in winter when 0.5% NaCl was added during sauerkraut manufacture and no statistical differences (P ≤ 0.05) were found in either natural or induced fermentations. Sauerkraut obtained from cabbage cultivated in summer, with 0.5% NaCl added to it, exhibited a significantly (P ≤ 0.05) lower ascorbigen content than sauerkraut obtained from cabbage cultivated in winter; moreover, controlled fermentation using L. plantarum gave rise to a significantly lower ascorbigen concentration (P ≤ 0.05) compared to natural and induced fermentation by L. mesenteroides. The addition of higher salt concentrations (1.5% NaCl) in sauerkraut manufacturing gave rise to significantly (P ≤ 0.05) lower ascorbigen levels in sauerkrauts produced by natural or induced fermentation from cabbages cultivated either in winter (ranging from 75.39 to 83.22 μmol/100 g dw) or summer (ranging from 65.76 to 80.96 μmol/100 g dw).

Table 2—.  Effect of fermentation conditions on ascorbigen content of Brassica oleracea var. capitata cv. Taler cultivated in different seasons.
Fermentation conditionsCabbage cultivated in winterCabbage cultivated in summer
Ascorbigen μmol/100g dwDry matter (%)Ascorbigen μmol/100g dwDry matter (%)
  1. Mean values ± standard deviation of 3 determinations. Different superscripts in the same column indicate significant differences (P ≤ 0.05).

Raw 5.71 ± 0.10a9.20 ± 0.00 8.12 ± 0.38a10.03 ± 0.00 
Natural fermentation
 NaCl 0.5%108.97 ± 6.55h10.51 ± 0.03 97.06 ± 4.49g8.88 ± 0.02
 NaCl 1.5%83.22 ± 5.51e10.60 ± 0.01 65.76 ± 2.20b9.87 ± 0.46
Fermentation by L. plantarum
 NaCl 0.5%108.34 ± 5.92h9.65 ± 0.0291.98 ± 3.50f9.58 ± 0.34
 NaCl 1.5% 76.80 ± 5.35cd10.06 ± 0.03  79.17 ± 5.07cd9.18 ± 0.31
Fermentation by L. mesenteroides
 NaCl 0.5%104.56 ± 3.93h9.10 ± 0.1296.67 ± 2.44g8.73 ± 0.07
 NaCl 1.5%75.39 ± 2.63c9.70 ± 0.03 80.96 ± 4.02de10.62 ± 0.01 

Ascorbigen formation is initiated by plant tissue disruption after which GB and myrosinase come into contact. Thus, GB and ascorbic acid in intact plant tissues are precursors in ascorbigen formation. Hrncirik and others (2001) reported that as ascorbic acid is in an appreciable excess in fresh Brassica vegetables, therefore, the limiting factor of ascorbigen yield is only GB. This fact justifies higher ascorbigen content found in sauerkrauts obtained from winter cabbage (2.54 μmol GB/g dw) compared to summer cabbage (1.83 μmol GB/g dw) with 0.5% NaCl added to it.

In fermented cabbage, Ciska and Pathak (2004) reported ascorbigen values of 140 μmol/100 g DW. Lower ascorbigen content observed in sauerkraut in the present study may be due to lower GB levels in white cabbage cv. Taler. GB content in winter and summer cabbage were 23.3 and 18.3 μmol/100 g FW, respectively, while Ciska and Pathak (2004) found a GB concentration of 31.95 μmol/100 g FW. The latter researchers also concluded that the content of GLS breakdown products in fermented cabbage does not depend only on the content of native GLS in raw cabbage, but may substantially depend on factors such as physicochemical properties as volatility, stability, and reactivity in an acidic environment and microbiological stability as well.

As mentioned previously, reaction conditions, including pH value, are other important factors, considerably affecting ascorbigen formation (Hrncirik and others 2001). GB is transformed in myrosinase-catalyzed reactions to their highly unstable isothiocyanates, which releases the stable thiocyanate ion and the indol-3-ylmethyl carbonium ions. These carbonium ions, or their isothiocyanate precursors, react easily with nucleophilic reagents such as water or ascorbic acid. It is found that ascorbic acid, at weakly acid to neutral conditions, is a much better nucleophile than water, thus, favoring ascorbigen formation (Buskov and others 2000). In reaction media with too low pH, for example, below 4, the reactivity of ascorbic acid, with a pKa of around 4.2, is reduced, and the formation of nitriles in myrosinase-catalyzed reaction is increased at the expense of isothiocyanate formation (Agerbirk and others 1998). Johanningsmeier and others (2005) reported that brining cabbage at 0.5% NaCl resulted in delayed acid production, and therefore a slower initiation of pH decrease, than in dry-salted (2% NaCl) treatments. Higher ascorbigen levels in 0.5 NaCl sauerkraut could be explained to a slower initiation on pH decrease, which consequently would lead to a higher yield in the isothiocyanates precursors of ascorbigen.

Influence of growth season and fermentation conditions on ascorbic acid content of cabbage

Table 3 shows the effect of fermentation conditions on ascorbic acid content in white cabagge (Brassica oleracea var. capitata cv. Taler) cultivated in winter and summer. The vitamin C content measured as ascorbic acid in raw white cabbage was higher in cabbage cultivated in summer (373.33 mg/100 g dw, equivalent to 37.3 mg/100 g fw) compared to cabbage cultivated in winter (302.96 mg/100 g dw equivalent to 27.9 mg/100 g fw). Cabbage fermentation led to a decrease in ascorbic acid content (average 48% and 34% in winter and summer cabbages, respectively).

Table 3—.  Effect of fermentation conditions on ascorbic acid of Brassica oleracea var. capitata cv. Taler cultivated in different seasons.
CabbageCabbage cultivated in winterCabbage cultivated in summer
Ascorbic acid mg/100g dwAscorbic acid mg/100g dw
  1. Mean values ± standard deviation of 3 determinations. Different superscripts in the same column indicate significant differences (P ≤ 0.05).

Raw cabbage 302.96 ± 13.60b373.33 ± 14.11e
Natural fermentation
 NaCl 0.5%156.72 ± 8.90a256.31 ± 16.66cd
 NaCl 1.5%158.00 ± 9.56a182.83 ± 17.23a
Fermentation with L. plantarum
 NaCl 0.5%162.63 ± 8.20a238.00 ± 31.73bc
 NaCl 1.5%161.47 ± 6.50a266.25 ± 3.96d  
Fermentation with L. mesenteroides
 NaCl 0.5%159.85 ± 5.36a229.43 ± 5.06b  
 NaCl 1.5%160.68 ± 7.10a239.33 ± 23.36bc

Sauerkraut produced from cabbage cultivated in summer exhibited significantly (P ≤ 0.05) higher levels of ascorbic acid regardless of the fermentation conditions used. In general, salt concentration and microorganisms responsible for acid lactic fermentation seemed to have no impact on ascorbic acid levels in sauerkraut produced from cabbage cultivated in winter and summer. However, naturally fermented cabbage, cultivated in summer and with 1.5% NaCl added to it, exhibited significantly (P ≤ 0.05) lower ascorbic acid content compared to sauerkrauts from summer cabbage produced under different fermentation conditions.

The ascorbic acid content of white cabbage cv. Taler was higher compared to 18 cabbage cultivars reported by Singh and others (2007), in which the vitamin C content on fresh weight basis ranged from 5.7 to 23.5 mg/100 g. Vitamin C content of white cabbage cv. Taler was higher also in comparison with other Brassica vegetables such as cauliflower, brussels sprouts, and chinese cabbage, but lower compared to broccoli (Singh and others 2007). Summer-cultivated cabbage exhibited higher content in vitamin C. This may be due to climatic conditions, including light and average temperature during growth and development of plant tissues, have a strong influence on the ascorbic acid content of horticultural crops (Lee and Kader 2000). In general, the lower the light intensity during growth, the lower ascorbic acid content of plant tissues. Regarding temperature, ascorbic acid content depends on factors such as total available heat and the extent of low and high temperatures. Vallejo and others (2003b) concluded that those broccoli cultivars grown in the spring season showed higher vitamin C contents than those grown in the winter one.

Loss of vitamin C due to cabbage fermentation may be explained partially to be a result of ascorbic acid involvement in ascorbigen formation. Hrncirik and others (2001) reported that the decrease of ascorbic content as a consequence of its binding into ascorbigen will not achieve more than 10% of its total amount. The higher losses of ascorbic acid observed currently during sauerkraut manufacture probably include other mechanisms such as food processing. Ascorbic acid is very susceptible to chemical and enzymatic oxidation by ascorbic oxidase during processing (Lee and Kader 2000). Losses in vitamin C occur when vegetables are severely cut or shredded, as in the case of cabbage (Mozafar 1994). Excessive trimming of leafy vegetables results in loss of outer leaves that contain more vitamins than inner leaves. Trimming of outer leaves and of the core and associated inner leaves of Chinese cabbage had a greater effect on reduction of vitamin C content than storage at 4 °C such (Klieber and Franklin 2000) as enzymatic breakdown by ascorbate oxidase, autoxidation, and so on.

Conclusions

Cabbage cultivated in summer exhibited higher total GLS content than cabbage cultivated in winter. However, the ascorbigen precursor, GB, was found higher in cabbage cultivated in winter. Glucoiberin, sinigrin, and GB were dominating in raw white cabbage cultivated either in winter or summer seasons. Ascorbigen, acknowledged as having a beneficial impact on human health, is formed during white cabbage (Brassica oleracea var. capitata cv. Taler) fermentation. Brining cabbage at low salt concentration (0.5% NaCl) improved the ascorbigen content in sauerkraut after 7 d of fermentation at 25 °C. The highest ascorbigen concentration was observed in low-sodium (0.5% NaCl) sauerkraut produced from cabbage cultivated in winter submitted to either natural or started-induced fermentation. Higher ascorbigen content found in low-sodium sauerkrauts is in concordance with nowadays consumer preferences of lower their sodium intake. Ascorbic acid content was higher in cabbage cultivated in summer and fermentation process led to significant reduction. Therefore, the selection of cabbages with high GB content and the production of low-sodium sauerkrauts may provide enhanced health benefits towards prevention of chronic diseases.

Acknowledgments

This study was performed under the common Polish–Spanish (PAN–CSIC) collaboration 2006PL-0006 and it was funded by the Spanish Commission of Science and Technology AGL2007-62044, and statutory funds of the Dept. of Food Technology of the Inst. of Animal Reproduction and Food Research of Polish Academy of Sciences.

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