Eur Food Res Technol (2000) 212 : 70–74
Q Springer-Verlag 2000
ORIGINAL PAPER
Cristina Inocencio 7 Diego Rivera 7 Francisco Alcaraz Francisco A. Tomás-Barberán
Flavonoid content of commercial capers (Capparis spinosa, C. sicula and C. orientalis) produced in mediterranean countries
Received: 3 March 2000
Abstract The flavonol content of commercial capers (pickled flower buds of Capparis spinosa L., C. sicula Veill. in Duham. and C. orientalis Veill. in Duham.) produced in different Mediterranean countries (Spain, Italy, Greece, Turkey and Morocco) has been analysed. The content of quercetin 3-rutinoside, kaempferol 3-rutinoside; and kaempferol 3-rhamnosyl-rutinoside as well as the aglycones (quercetin and kaempferol) were evaluated by HPLC coupled with a diode-array detector. The free aglycones were not detected in the original flower buds, indicating that they were produced during the brining process. In addition, brining extracted most of the kaempferol 3-rhamnosyl rutinoside present in the original buds, this being a minor constituent in pickled capers. The average content of flavonoid glycosides in commercial capers was 5.18 mg/g fresh weight. When this value is converted to the corresponding aglycones (quercetin and kaempferol), capers provide an average of 3.86 mg/g fresh weight. A serving of capers (10 g) will provide 65 mg of flavonoid glycosides or the equivalent 40 mg of quercetin as aglycone. Keywords Capparis spinosa 7 Capparis sicula 7 capers 7 flavonols
Introduction Capparis sicula Veill. in Duham, C. orientalis Veill. in Duham and C. spinosa L. are small shrubs widely distributed in Mediterranean countries. The genus Capparis
C. Inocencio 7 D. Rivera 7 F. Alcaraz Department of Botany, Biology, University of Murcia, Espinardo 30100, Spain F.A. Tomás-Barberán (Y) Phytochemical Laboratory, Department of Food Science and Technology, CEBAS (CSIC), P.O. Box 4195, Murcia 30080, Spain e-mail:
[email protected]
is included in the Capparidaceae family, which is closely related to the mustard family (Cruciferae). Their healing properties, including antirheumatic, tonic or expectorant activities, have been known since antiquity from numerous tribes of different countries around the Mediterranean sea, and caper plants have been used in folk medicinal remedies for many years [1–3]. Recent studies have shown the antihepatotoxic activity of caper extracts [4]. The flower buds, the fruits and occasionally the shoots of C. spinosa and C. sicula, have been used as a condiment in Greece since ancient times, being greatly appreciated for their pungent and bitter flavour. The pickled flower buds, known commercially as capers, are the most appreciated product of this plant. They are harvested when they are still tightly closed, brined, and packed with vinegar [5]. They are selected by size, the smaller ones being the most appreciated in the market due to their concentrated flavour and to the difficulties of their collection in the wild. The caper berries are also commercialised after pickling [6]. Capers are usually commercialised in some Mediterranean countries such as Greece, Italy, Turkey, Morocco and Spain, and they are exported mainly to central European countries, the USA and the UK as a delicatessen product. However, they are consumed, primarily, in their production areas, where they are integrated in the culinary culture, normally being consumed fresh as ingredients in salad seasoning, although they are also used in diverse plates of pasta, fish or meat. The usual commercial processing for the manufacture of pickled capers begins with a pre-treatment in high brine (ca 16% NaCl w/v, at equilibrium, increasing to 20% after changing the first brine). At the end of the pre-treatment stage, capers are desalted and packed into a finished product containing about 6% salt and 1% acetic acid (Peregrin, personal communication). From a dietary point of view, the flavonoid content gives a special value to this product. The occurrence of the flavonol rutin in extracts of different parts of this plant has been known for a long time [7]. In more re-
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cent studies, based on spectrophotometric and chromatographic analyses, the flavonols kaempferol and quercetin were identified as well as certain glycosylated derivatives (quercetin, kaempferol 3-rutinosides and kaempferol 3-O-rhamnosyl-rutinoside) [8, 9]. Commercial pickled capers show characteristic yellowish spots, and it was demonstrated that these are, in fact, precipitates of rutin and other flavonols [11]. Flavonoids, and especially quercetin derivatives, have received special attention as dietary constituents during the last few years, as epidemiological studies point out that they might have a role in preventing cardiovascular diseases and cancer [12–16]. This healthpromoting activity seems to be related to the antioxidant (free-radical scavenging) activity of these compounds [17]. Therefore, there has been an interest in the identification and quantification of the dietary sources of flavonoids, and in the correlation of flavonoid content with health-promotion potential of specific plant food products. The purpose of the present work was the study of the effect of processing (brining) on flavonoids from flower buds of Capparis spinosa, C. sicula and C. orientalis, and the quantification of flavonoids in commercial capers harvested and produced in different Mediterranean countries.
Material and methods
Table 1 Percentage of flavonoid glycosides in caper buds from herbarium material Sample
Locality
% K3-RR
% Q3-R
% K3-R
C. C. C. C. C. C. C. C. C. C.
Mallorca Murcia Alicante Mallorca Málaga Mallorca Valencia Valencia Mallorca Alicante
21.29 27.60 19.15 17.43 16.27 15.07 12.55 8.00 6.59 10.25
43.37 25.49 33.57 45.47 25.35 28.55 37.69 70.00 68.41 49.55
35.35 46.90 47.29 37.10 58.39 56.39 49.76 22.00 25.00 40.20
sicula sicula sicula sicula sicula orientalis orientalis orientalis spinosa spinosa
Values are percentage of the total absorbance of the HPLC chromatograms at 340 nm (K3-RR) Kaempferol 3-rhamnosyl-rutinoside; (Q3-R) Quercetin 3-rutinoside; (K3-R) Kaempferol 3-rutinoside
90% B at 45 min. The flavonoids were detected with a diodearray detector (Merck-Hitachi L-3000) to obtain the UV spectra of the different phenolic compounds, and the chromatograms were recorded at 340 and 290 nm. Flavonoid quantitation Wherever possible, the different flavonoids were identified by chromatographic comparisons with authentic compounds (commercial or previously isolated and identified from capers) [8–11] and by matching their UV spectra with those of the standards. Flavonoids were quantified at 340 nm as rutin (quercetin 3-rutinoside).
Capers Seventeen commercial samples processed into brines were studied. They were produced in different Mediterranean countries. Samples from Spain (5; 1 from Iberian Peninsula and 4 from Balearic Islands), Turkey (3), Morocco (7), Italy (1) and Greece (1) were analysed (Table 1). All samples were preserved into brines, with the exception of the sample from Italy that was preserved with salt. Capparis flower buds from different species and localities were obtained from the herbarium of the Department of Botany, Murcia University, where voucher specimens have been deposited.
Flavonoid extraction For each sample three replicates were analysed, using about 2 g of capers in each one (ca 0.5 g dried flower buds from herbarium material). The buds were homogenised with 20 mL of a solution containing MeOH, H2O, and DMSO (2 : 2 : 1). The homogenised samples were filtered and then extracted again with 20 mL of extraction solution.
HPLC analysis All HPLC analyses were carried out using a Merck-Hitachi Liquid Chromatograph L-6200 with a diode array detector MerckHitachi L-3000 and an autosampler Merck-Hitachi AS-2000A. Data were stored and processed with the DAD-Manager software (Merck). The column used was a LichroCART RP-18 (Merck) (12.5!0.4 cm, 5 mm particle size) eluted with water-formic acid (19 : 1, v:v) (solvent A) and methanol (solvent B) at 1 mL/min. The following gradient was used: 10% B in A increasing to reach 50% B at 25 min, 80% B at 35 min, 80% methanol at 40 min, and
Results and discussion Qualitative HPLC analysis of flower buds flavonoids The flavonoid profile of flower buds from different Capparis species was evaluated using herbarium material deposited in the Department of Botany at Murcia University. The extracts were analysed by HPLC and three main flavonoids were detected in all the analysed samples (Fig. 1). These flavonoids were identified as kaempferol 3-rhamnosyl-rutinoside (1), quercetin 3-rutinoside (2) and kaempferol 3-rutinoside (3), by chromatographic comparison with authentic standards previously isolated from capers and identified in the Phytochemical Laboratory (CEBAS-CSIC, Murcia) [8–10]. Different species, subspecies, and hybrids growing wild in different Spanish localities were analysed (Table 1). All the analysed samples showed a common flavonoid profile, but some differences in the relative amounts of the individual flavonoids were detected. For instance, in the sample of C. orientalis collected in Valencia 70% of the absorbance corresponding to flavonoids is related to quercetin 3-rutinoside, while in the sample of C. sicula from Málaga, the corresponding value is only 25% (Table 1). This indicates that there is a wide variation in the relative amounts of the different flavonoids in caper flower buds, and that environmental and phys-
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Fig. 1 Caper HPLC chromatograms of the extracts obtained from herbarium flower buds of Capparis spp.. (A) Capparis orientalis; (B) Capparis sicula. (1) Kaempferol 3-rhamnosyl-rutinoside; (2) Quercetin 3-rutinoside; (3) Kaempferol 3-rutinoside
Fig. 2 HPLC chromatograms of commercial pickled capers. (A) Sample Morocco 7 (pickled); (B) Sample from Italy (in salt). (1) Kaempferol 3-rhamnosyl-rutinoside; (2) Quercetin 3-rutinoside; (3) Kaempferol 3-rutinoside; (4) Quercetin; (5) Kaempferol
iological factors can have important effects on the flavonoid profile. It is interesting to mention that kaempferol 3-rhamnosyl-rutinoside is present in most of the herbarium samples in percentages above 10%, reaching in some cases values above 20%.
variable amounts. While sample 6 from Morocco contained more than 1 mg/g of quercetin and close to 2 mg/ g of free kaempferol, other samples like the one from Italy, only contained traces of the free aglycones (Fig. 2). The total glycoside contents averaged 5.18 mg/ g f.w. with a maximum value of 15.29 mg/g for sample Morocco 1 and a minimum value of 0.8 mg/g for sample Morocco 6. The content of free aglycones was also very variable with free quercetin in the range 0.03–1.45 mg/g and free kaempferol in the range 0.1–2.8 mg/g (Table 2). The occurrence of free flavonoid aglycones in pickled capers means that during the brining process some hydrolysis has occurred releasing quercetin and, mainly, kaempferol from the original glycosides, because the free aglycones were not detected in the chromatograms of flower buds. Supporting this hydrolysis due to the brining process is the fact that the commercial sample from Italy, which is preserved in salt and not pickled, contains only traces of the corresponding aglycones. The large variability in flavonoid aglycone content of the available commercial samples indicates that this can be related to differences in the conditions of the brining process (time of brining and possible microbial growth and fermentation). This can lead to the hydrolysis of the main part of caper bud flavonoids,
HPLC analysis of commercial caper flavonoids The extraction system used extracted 95% of the flavonoid present in capers. A qualitative analysis of the extracts revealed that, in most samples, rutin (quercetin 3-rutinoside) was the main constituent, and kaempferol 3-rutinoside was the second compound. Kaempferol 3rhamnosyl-rutinoside was only detected in some samples and then in rather small amounts. This is different to the flavonoid patterns found in the flower buds, in which kaempferol 3-rhamnosyl-rutinoside is present in all samples in significant amounts. This shows that, during the brining process, this flavonoid which is the most soluble is preferentially extracted from the buds and, therefore, its content in the pickled capers is generally very small (Table 2). In most commercial samples the aglycones kaempferol and quercetin are also detected although in very
73 Table 2 Flavonoid composition of commercial Mediterranean capers. Values are mg/g f.w. Samples
K3-RR
Spain (Iberian Peninsula) Spain (Balearic Islands 1) Spain (Balearic Islands 2) Spain (Balearic Islands 3) Spain (Balearic Islands 4) Turkey 1 Turkey 2 Turkey 3 Morocco 1 Morocco 2 Morocco 3 Morocco 4 Morocco 5 Morocco 6 Morocco 7 Italy Greece Mean
0.19 0.00 0.00 0.25 0.00 0.00 0.00 0.09 0.17 0.51 0.34 0.50 0.18 0.00 0.32 0.15 0.71
Q3-R 2.01 1.31 1.14 3.11 1.30 5.07 5.41 0.76 10.47 3.03 4.00 4.96 1.81 0.80 1.19 7.83 2.29
K3-R
Free Kaempferol
Free Quercetin
Total glycosides
Total Flavonoids
0.75 0.16 0.18 1.09 0.16 2.63 2.76 0.15 4.65 1.62 3.24 3.50 1.51 0.00 1.45 2.72 1.61
0.42 2.80 1.09 1.46 1.41 1.70 1.10 1.20 0.50 0.66 0.39 0.35 0.43 1.82 1.43 0.10 0.87
0.21 0.50 0.45 0.32 0.52 0.46 0.32 0.15 0.03 0.03 0.08 0.03 0.03 1.45 0.43 0.03 0.54
2.95 1.47 1.32 4.45 1.46 7.71 8.17 1.00 15.29 5.15 7.58 8.96 3.50 0.80 2.97 10.70 4.61 5.18B3.29
3.58 4.77 2.87 6.23 3.39 9.86 9.59 2.36 15.82 5.84 8.04 9.34 3.96 4.08 4.83 10.83 6.02 6.55B3.57
K3-RRpkaempferol 3-rhamnosyl-rutinoside; K3-Rpkaempferol 3-rutinoside; Q3-RpQuercetin 3-rutinoside. Total flavonoidsp Total glycosidescfree aglycones Table 3 Quercetin and kaempferol equivalents provided by the different glycosides. Commercial Mediterranean capers. Values are mg/g f.w. Samples
Free Total Total Total Kaempferol Quercetin Kaempferol Free Kaempferol Quercetin Kaempferol Quercetin Flavonoids from from from as aglycones K3-R Q3-R K3-RR
Spain (Iberian Peninsula) Spain (Balearic Islands 1) Spain (Balearic Islands 2) Spain (Balearic Islands 3) Spain (Balearic Islands 4) Turkey 1 Turkey 2 Turkey 3 Morocco 1 Morocco 2 Morocco 3 Morocco 4 Morocco 5 Morocco 6 Morocco 7 Italy Greece Mean
0.07 0.00 0.00 0.10 0.00 0.00 0.00 0.03 0.00 0.19 0.13 0.19 0.07 0.00 0.12 0.06 0.27
0.98 0.64 0.56 1.52 0.63 2.48 2.64 0.37 5.11 1.48 1.96 2.43 0.88 0.39 0.58 3.83 1.12
0.36 0.07 0.09 O.52 0.08 1.25 1.31 0.07 2.21 0.77 1.54 1.66 0.72 0.00 0.69 1.29 0.77
0.42 2.80 1.09 1.46 1.41 1.70 1.10 1.20 0.50 0.66 0.39 0.35 0.43 1.82 1.43 0.10 0.87
0.21 0.50 0.45 0.32 0.52 0.46 0.32 0.15 0.03 0.03 0.08 0.03 0.03 1.45 0.43 0.03 0.54
0.85 2.87 1.18 2.08 1.49 2.95 2.41 1.30 2.71 1.62 2.05 2.20 1.22 1.82 2.24 1.45 1.90
1.19 1.14 1.01 1.84 1.15 2.94 2.96 0.52 5.14 1.51 2.04 2.46 0.91 1.84 1.02 3.85 1.66
2.04 4.01 2.19 3.92 2.64 5.89 5.38 1.82 7.85 3.13 4.09 4.66 2.13 3.66 3.26 5.30 3.56 3.86B1.61
K3-RRpKaempferol 3-rhamnosyl-rutinoside; K3-RpKaempferol 3-rutinoside; Q3-RpQuercetin 3-rutinoside)
and the release of aglycones, which are much more susceptible to oxidative degradation than the corresponding glycosides, and are thus easily degraded. These results show that capers are a very rich source of the flavonols quercetin and kaempferol. In Table 3, quercetin and kaempferol equivalents provided by the different flavonol glycosides present in the analysed caper samples are shown. Total flavonoids, expressed as aglycones, range between 1.82 and 7.85 mg/g with an average of 3.86 mg/g. These values are very relevant. If we compare this flavonol content with those reported for the edible part of onions (one of the foods richer in quercetin glycosides) [18], we find that capers are a
very rich source of flavonoids. The values reported for onions are as follows: raw onion 39–42 mg/100 g edible portion, boiled onion 21–29 mg/100 g, microwaved onion 22–27 mg/100 g, and fried onion 22–29 mg/100 g f.w. If we consider the possible daily intake of capers in a salad serving (20 capers, 10 grams), this will provide 20–80 mg quercetin (30–160 mg flavonol glycosides). These values mean that a serving of capers provides at least the same amount of quercetin as 100 g of onions. Acknowledgements This work has been financially supported by the Spanish CICYT (AGF-96–1040).
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