J. Agric. Food Chem. 2006, 54, 457−462
457
Cutin Composition of Five Finnish Berries HEIKKI KALLIO,* RIIKKA NIEMINEN, SASKA TUOMASJUKKA,
AND
MARI HAKALA
Department of Biochemistry and Food Chemistry, University of Turku, FI-20014 Turku, Finland
The raw cutin (i.e., extractive-free isolated cuticular membrane) fraction from Finnish berries, sea buckthorn (Hippophae¨ rhamnoides), black currant (Ribes nigrum), cranberry (Vaccinium oxycoccos), lingonberry (Vaccinium vitis-idaea), and bilberry (Vaccinium myrtillus), was depolymerized by NaOMecatalyzed methanolysis. The composition of cutin monomers was determined by GC-(EI)MS analysis either as methyl esters or as TMSi esters, with OH groups derivatized to TMSi ethers. There was a notable difference in the degree of depolymerization, ranging from 6 to 47%. The extractive-free berry cuticle, that is, raw cutin, thus contains 50% of the raw cutin in plants. Cutin is known to represent ∼5060% of the weight of the isolated apple fruit membranes obtained with transesterification (15). We have obtained from apple peel yields of up to 80% (unpublished results). Over 70% of membrane material from tomato peels is released as cutin monomers, the remaining part being mostly carbohydrates (16). The typical yield in alkaline hydrolysis of lime fruit cutin is 60-65% and in transesterification ∼50% (17). This resistance to hydrolytic breakdown of lime fruit raw cutin may be attributed to non-ester cross-links within the polymeric structure. The unreacted residue has been found to consist almost exclusively of rigid, solid-like functional groupings, determined with solidstate 13C NMR (17). This could be the case for berry cutins as well, but the residual material may also be a mixture of polymers, for example, polysaccharide and non-ester cutin, or
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Cutin in Berries
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Figure 1. Total ion chromatogram obtained from derivatized sea buckthorn (A), black currant (B), and bilberry (C) methanolysates. Table 1. Cutin Monomer Yields as Percentage of the Extractive-free Raw Cutin by Methanolysis sample
cutin monomer yielda (%)
sea buckthorn (Hippophae¨ rhamnoides) black currant (Ribes nigrum) cranberry (Vaccinium oxycoccos) lingonberry (Vaccinium vitis-idaea) bilberry (Vaccinium myrtillus)
46 8 27 30 6
a Determined gravimetrically of CHCl -soluble material from NaOMe methanolysis, 3 SD ) 1−3%.
cutan. Because the composition of this residual material is not well understood and structural studies have not been carried
out in our experiments, we have not been able to make any conclusions about the amount and composition of cutan in the berry cuticles. Apparently, berry cuticle is quite resistant to esterbreaking reactions, which may be due to the high amount of cutan present. Cuticles from some berries seem to almost totally lack the ester-linked cutin. This may have a strong influence on the various functions of cuticle in berries and also as dietary fiber. Cutin Monomer Compositions. The compositions of the hydroxy acid monomers released from the raw cutin fraction of the berries by alkaline transesterification were identified by GC retention times and EI mass spectra. The analyses were carried out as TMSi ethers of methyl or TMSi esters. Only a few reference compounds were available. The retention behavior
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Kallio et al.
Table 2. Important Ions in Mass Spectra of Cutin Monomers and Calculated Kovats Indices important ions in mass spectra (m/z) ω-hydroxy
acida
16-hydroxyhexadecanoic 16-hydroxy-10-oxohexadecanoic (main isomer) 18-hydroxyoctadec-9-enoic
dihydroxyhexadecanoic (major 10,16-diOH, minors 9,16 and 8,16-diOH) 20-hydroxyeicosanoic
9,10-epoxy-18-hydroxyoctadec12-enoic 9,10-epoxy-18-hydroxyoctadecanoic 9,10,18-trihydroxyoctadec12-enoic 9,10,18-trihydroxyoctadecanoic 22-hydroxydocosanoic
methyl ester TMSi
etherb
IDc
Ikd
TMSi ester TMSi etherb
343 [M − 327 [M − 311 [M − 59 and 146,* 103, 89, 75, 73, 55 357 [M − 15]+, 341 [M − 31]+, 325 [M − 47]+, 214, 126, 111, 83, 75, 73, 55 384 [M]+, 369 [M − 15]+, 353 [M − 31]+, 337 [M − 47]+, 159 and 146,* 129, 109, 95, 75, 73, 55 431 [M − 15]+, 415 [M − 31]+, 399 [M − 47]+, 317, 303, 289, 275, 273, 259, 245, 159 and 146,* 147, 129, 103, 95, 75, 73, 55 399 [M − 15]+, 383 [M − 31]+, 367 [M − 47]+, 159 and 146,* 103, 89, 75, 73, 55
S, R
2268
D
2425
R, D
2435
R, D
2460
D
2665
471 [M − 31]+, 383, 303, 271, 259, 243, 213, 191, 155, 129, 121, 109, 103, 89, 75, 73, 55 489 [M − 15]+, 473 [M − 31]+, 457 [M − 47]+, 385, 303, 274,* 259, 201, 155, 129, 121, 109, 103, 81, 75, 73, 55 545 [M − 15]+, 361, 317, 271, 259, 155, 147, 129, 109, 103, 75, 73 547 [M − 15]+, 332,* 303, 259, 243, 212, 155, 147, 129, 109, 103, 81, 75, 73 442 [M]+, 427 [M − 15]+, 411 [M − 31]+, 395 [M − 47]+, 159 and 146,* 129, 103, 83, 75, 73, 55
R, D
2700
R, D
2731
R, D
2743
R, D
2771
R, D
2863
15]+,
31]+,
47]+,
IDc
Ikd
refs
401 [M − 385 [M − 311 [M − 217 and 204,* 147, 117, 103, 75, 73, 55 415 [M − 15]+, 272, 251, 215, 201, 126, 111, 83, 75, 73, 55 442 [M]+, 427 [M − 15]+, 411 [M − 31]+, 383, 337 [M − 105]+, 217 and 204,* 147, 135, 129, 117, 109, 95, 75, 73, 55 489 [M − 15]+, 399 [M − 90 − 15]+, 383 [M − 105]+, 331, 317, 303, 289, 275, 217 and 204,* 147, 129, 103, 95, 75, 73, 55 457 [M − 15]+, 441 [M − 31]+, 367 [M − 105]+, 217 and 204,* 147, 129, 117, 103, 75, 73, 55 545 [M − 15]+, 441, 423, 361, 329, 317, 301, 271, 259, 191, 149, 129, 121, 109, 103. 75, 73, 67 547 [M − 15]+, 443, 425, 332, 317, 303, 243, 147, 129, 121, 109, 103, 75, 73, 67
S, D
2390
11, 12
D
2548
10
D
2555
11
D
2575
11, 13
D
2783
14
D
2808
7
D
2840
7
513 [M − 105]+, 419, 329, 317, 301, 217 and 204,* 191, 147, 129, 109, 103, 75, 73 515 [M − 105]+, 390, 317, 303, 217 and 204,* 147, 129, 109, 103, 81, 75, 73 485 [M − 15]+, 469 [M − 31]+, 395 [M − 105]+, 217 and 204,* 147, 129, 117, 103, 75, 73, 55
D
2848
12
D
2878
11, 12
15]+,
31]+,
105]+,
R, D
14
a Analyzed as methyl ester TMSi ethers and TMSi ester TMSi ethers. b Asterisks indicate typical rearrangement ions. c S, identified by comparison to standard compound; R, identified by comparison to published spectrum; D, deduced according to the mass spectrum in the present study. d Kovats index.
Table 3. Approximate Composition (Percent) of Cutin Monomers Released from Five Berries by Methanolysis sea buckthorn
lingonberry
black currant
peaka
ω-hydroxy acidb
1 2 3 4 5 6 7 8 9 10
16-hydroxyhexadecanoic 16-hydroxy-10-oxohexadecanoic (main isomer) 18-hydroxyoctadec-9-enoicd dihydroxyhexadecanoic (major, 10,16-diOH; minor, 9,16 and 8,16-diOH) 20-hydroxyeicosanoic 9,10-epoxy-18-hydroxyoctadec-12-enoicc,d 9,10-epoxy-18-hydroxyoctadecanoicd 9,10,18-trihydroxyoctadec-12-enoicd 9,10,18-trihydroxyoctadecanoic 22-hydroxydocosanoic
1
1
1
1
2 15 1 30 41 2 3 tr
6 20
6 9
11 28 6 23 tr
4 56 1 20 1
9 13 tre 11 15 8 18 1
3 12 13 30 4 3 8 1 2 1
total hydroxy acids
94
95
97
77
76
1 1 5 100
1 1 3 100
tr tr 3 100
6 9 9 100
4 9 11 100
cranberry
bilberry
Other Compound Groupsb aromatic compounds (comprises also hydroxy acids) long-chain aliphatic acids, diacids, and alcohols unidentified total
d
a Peak number in Figure 1. b Compounds determined as methyl ester TMSi ether and/or TMSi ether TMSi ester. c Determined as corresponding methoxyhydrin compounds. Position of double bond not confirmed by chemical methods. e tr,
