2007 vol. 66, 197-203 DOI: 10.2478/v10032-007-0022-8 ______

INFLUENCE OF CULTIVAR AND STORAGE TIME ON THE CONTENT OF HIGHER FATTY ACIDS IN CUTICLE

Marjan SIMČIČ, Melita KADUNC, Janez HRIBAR, Rajko VIDRIH Biotechnical Faculty, Dep. of Food Science and Technology University of Ljubljana Jamnikarjeva 101, 1000 Ljubljana, Slovenia

Summary The main objective was to determine the fatty acid composition on the sur- face of apple peel and in seeds. 7 apple cultivars were used: , , Glo- ster, , , Pink Lady and . Apple seeds contain from 8.4% to 15% of water and from 17.1% to 28.0% (f.w.) of fat. The fatty acid content in seeds ranged as follows: linoleic (55.9-62.3), oleic (23.9- 30.7%), palmitic (7.1-9.3%), stearic (1.6-2.3%), arachidic (1.2-1.6%), linolenic (0.4-0.7%. Linolic acid is prevalent fatty acid on peel as well. Ratio of linolenic acid on peel is up to 30 fold higher than in seeds. Saturated palmitic and stearic fatty acid ratio is considerably higher in peel than in seeds while is the ratio of oleic acid nearly the same as in seeds. During storage of the ratio of pal- mitic and oleic acid increases, while ratio of linoleic acid considerably de- creases. key words: apples, higher fatty acids, peel, seeds

INTRODUCTION

As far as apples are concerned fatty acids are present in seeds and in much lower concentration also on the upper part of peel. Fatty acids in apple seeds are either esterified with glycerol or are present as free fatty acids (Lu & Foo 1997). Fruits in general are coated with layers of lipids that are involved in the protection of fruits toward atmosphere conditions. Protection layers serve to adapt the fruit to the extreme temperatures and other stress conditions which in fruit may lead to different storage disorders (Marangoni et al. 1996, Saquet et al. 2000). Fatty acids build up in developing and ripening apples especially during climacteric respiratory rise. After the respiratory rise, fatty acids begin to decline and are metabolised to various lipid fractions (Meigh & Hulme 1965). On the other hand fatty acids are known to act as precursors of aroma volatile production (Fellman et al. 2000, Harb et al. 2000).

Corresponding author e-mail: [email protected] © Copyright by RIVC 198 VEGETABLE CROPS RESEARCH BULLETIN 66 ______

Little is known about the role of higher fatty acids in scald appearance. Higher fatty acids are present on the skin either as free or in wax. Chemically is wax esterified with alcohol (Nelson & Cox 2000) and has a function to protect the fruit against environmental factors (Morice & Shortland 1973). Biosynthesis of waxes starts in plastids, the fatty acids are later elongated (Kunst & Samuels 2002). Typical characteristics of waxes depend on cultivar and climatological conditions like rainfall and temperature (Veraverbecke et al. 2001). Waxes not only protect the fruit in the outside, but play some protective role during fruit storage as well. During storage, waxes have a protective role against water loss while their composition is changing. Content of free fatty acids present on apple skin are changing during stor- age period (Morice & Shortland 1973). Total free fatty acids insoluble in petrol ether increased slightly during storage On the other hand, total free fatty acids soluble in petrol ether did not change significantly (Morice & Shortland 1973). Content of linolenic acid on the peel of cv. increased signifi- cantly during storage (Morice & Shortland 1973).

MATERIAL AND METHODS

Fatty acid composition was analysed in seeds and on the surface of seven apple cultivars (Elstar, Fuji, Gloster, Idared, Jonagold, Pink Lady and Golden Delicious) immediately after harvest or after 3 months of staorage at +1°C. Fatty acids in seeds were analysed according to the method described by Garces & Mancha (1993). Methyl esters of fatty acids (palmitic C 16 : 0; stearic C 18 : 0; oleic C 18 :1; linolic C 18 : 2; linolenic C 18 : 3 and arachidic C 20 : 0) were analysed on GC and peaks were recorded. Content of fatty acids on apple peel was analysed in hexane extract of apple surface. 5 apples were washed with 300 ml of hexane for 2 minutes. The volume of hexane was evaporated to 50 ml. Three ml of hexane extract were used to determine the fatty acid composi- tion on apple peel according to the method described by Garces & Mancha (1993). GC: Agilent technologies 6890N; Column: SUPELCO – SPB PUFA; 30 m x 0.25 mm x 0.2 μm; Detector: FID; Column temperature: 210°C; Detec- tor temperature: 260°C; Injector temperature: 250°C (split 1: 100); Carrier gas: He 1 mL·min-1; Injection volume: 1.0 μL.

RESULTS

Content of total fats in apple seeds varies from 17 to 28%, Golden Deli- cious has the highest content and Jonagold the lowest (Table 1). Compared to other fruit seeds, apple seeds have lower content of total fats. Linolic acid is the prevalent higher fatty acid in apple seeds of all cultivars investigated. Apple seeds contain from 55 to 62% of linolic acid. Its content is cultivar dependent, the highest ratio was found in Jonagold, the lowest in Glo- ster (Table 2). M.SIMCIC et al. – INFLUENCE OF CULTIVAR AND STORAGE ... 199 ______

Table 1. Fruit firmness (kg) at harvest and after storage and content of total fats in apple seeds at harvest (% in dry matter)

Firmness (kg) cultivar At harvest After storage Total fats (%) in dry matter Elstar 6.7 3.8 20.4 Jonagold 7.6 4.9 17.1 Golden Delicious 7.5 4.3 28.0 Gloster 8.2 5.8 26.3 Idared 7.0 4.7 22.9 Fuji 7.5 6.7 25.8 Pink Lady 8.9 8.4 23.7

Table 2. Fatty acid composition of apple seeds (as % of total fatty acids)

Cultivar C 16 : 0 C 18 : 0 C 18 :1 C 18 : 2 C 18 : 3 C 20 : 0 Elstar 7.47 2.24 30.75 57.37 0.61 1.56 Jonagold 9.32 1.63 24.48 62.27 0.70 1.16 Golden 7.53 2.26 30.00 58.25 0.44 1.51 Gloster 7.37 1.84 32.93 55.94 0.64 1.27 Idared 7.85 1.77 29.65 58.90 0.52 1.31 Fuji 7.15 1.85 30.63 58.46 0.51 1.40 Pink Lady 7.35 2.15 28.17 60.31 0.52 1.50

Next to linolic fatty acid is oleic, its ratio varies from 24 to 30%. Apple seeds contain below 1% of linolenic acid. Among saturated fatty acids apple seeds contain 7% of palmitic acid, around 2% of stearic acid and 1.5% of ara- chidic acid. The sum of linolic and oleic acid is very similar for all cultivars; cultivars that contain more linolic acid contain less oleic acid and vice versa. Linolic acid is prevalent fatty acid in seeds as well as on the surface of ap- ple peel (Table 3). The concentration of total fatty acids on apple peel is rather low ca. 0.02 mg·cm-2 of apple peel. At harvest cultivars Jonagold and Golden Delicious have the highest amount of total fatty acids on peel, cultivar Elstar and Fuji the lowest (Table 3). Content of arachidic acid was below threshold level. Compared to seeds, much higher ratio of linolenic, palmitic and stearic acid was determined on apple peel (Table 4). After storage period of three months the content of total fatty acids on ap- ple peel decreased at all cultivars except Idared (Table 3). Meigh & Hulme (1965) reported a decline of fatty acids after the respiratory climacteric. Brack- mann et al. (1995) found the fatty acid content to be dependent on the storage atmosphere. Low oxygen concentration inhibited the synthesis of fatty acids and aroma volatiles. Song & Bangerth (2003) reported of an increase of unsaturated linolic and oleic acid during on tree maturation of ‘Golden Delicious apples. 200 VEGETABLE CROPS RESEARCH BULLETIN 66 ______

Table 3. Fatty acid composition of apple peel (mg·100 mL-1 hexane extract) at harvest and during storage

SUM of Cultivar C16 : 0 C18 : 0 C18 : 1 C18 : 2 C18 : 3 fatty acids Elstar at harvest 1.36 0.78 1.05 1.38 0.99 5.56 after 37 days 1.28 1.16 1.77 1.94 1.17 7.32 after 68 days 1.03 0.50 0.97 1.05 0.68 4.23 after 102 days 1.43 0.50 1.09 0.88 1.06 4.96 Jonagold at harvest 6.01 3.68 8.67 17.82 2.09 39.52 after 37 days 2.14 1.43 3.93 7.28 0.96 15.74 after 68 days 1.50 0.84 1.74 2.8 0.64 7.63 after 102 days 1.04 0 0.83 0.75 0 2.62 Golden Delicious at harvest 5.19 3.26 9.60 18.25 2.79 39.09 after 37 days 4.64 2.77 9.12 15.07 2.94 35.54 after 68 days 2.19 1.06 3.43 4.23 1.23 12.14 after 102 days 2.23 0.93 3.23 3.34 0.82 10.55 Gloster at harvest 5.33 1.56 3.74 8.73 1.77 21.13 after 37 days 4.32 1.54 3.34 6.31 1.45 16.96 after 68 days 3.16 1.04 2.03 2.86 0 9.09 after 102 days 2.97 0.97 2.56 3.09 0 9.59 Idared at harvest 2.66 1.04 3.1 5.88 1.00 13.72 after 37 days 1.52 0.65 1.66 1.87 0.00 5.70 after 68 days 3.4 1.07 5.64 6.29 1.09 17.49 Fuji at harvest 1.20 1.00 2.40 3.38 0.62 8.60 after 37 days 1.81 1.44 4.80 5.34 0.81 14.20 after 68 days 1.29 0.89 3.06 2.95 0.00 8.19 Pink Lady at harvest 5.64 2.84 7.32 13.87 2.75 33.35 after 37 days 4.50 1.95 6.62 8.98 2.44 24.49 after 68 days 4.74 1.53 6.42 7.38 2.20 22.27

The ratio of fatty acids on apple peel is quite different from ratio in seeds. In both cases linolic acid prevails although its ratio in apple peel is substantially lower than in seeds. The ratio of saturated palmitic and stearic acid is much higher on apple peel than in seeds while is the ratio of oleic acid nearly the same as in seeds. Ratio of saturated/unsaturated fatty acids on apple peel in- creased during storage with the exception of cultivar Idared. Ratio of linolenic acid on the peel is up to 30 fold higher than in seeds. The ratio of fatty acids is changing during storage. Ratio of palmitic and oleic was increasing during storage (Table 4). Morice & Shortland (1973) find a decrease of palmitic and oleic acid ratio during storage of cultivars Dougherty, M.SIMCIC et al. – INFLUENCE OF CULTIVAR AND STORAGE ... 201 ______

Granny Smith and Sturmer. Ratio of stearic acid remained more or less stable during storage period which differs with the observation by Morice & Shortland (1973). They found stearic acid ratio to increase. Linolenic acid ratio remained more or less stable during storage at all cultivars with the exception of Fuji and Gloster where we did not detect it at all after storage. Considerable changes oc- curred with linolic acid. Ratio of linolic acid decreased considerably at all culti- vars with the exception of Fuji. In the observation made by Morice & Shortland (1973) the linolic acid ratio of cultivars Dougherty, Granny Smith and Sturmer increased during storage.

Table 4. Fatty acid composition of apple peel (as % of total fatty acids) at harvest and after storage

Fatty acid C 16 : 0 C 18 : 0 C 18 :1 C 18 : 2 C 18 : 3 at after at after at after at after at after Cultivar harvest storage harvest storage harvest storage harvest storage harvest storage Elstar 24.46 28.83 14.03 10.08 18.89 21.97 24.82 17.74 17.8 21.37 Jonagold 15.21 19.66 9.31 11.01 21.94 22.8 45.09 36.7 5.29 9.79 Golden 13.28 21.14 8.34 8.82 24.56 30.62 46.69 31.66 7.14 7.77 Gloster 25.22 30.97 7.38 10.11 17.7 26.70 41.32 32.22 8.38 0 Idared 19.39 19.44 7.58 6.12 22.59 32.25 42.86 35.96 7.29 6.23 Fuji 13.95 15.75 11.63 10.87 27.91 37.36 39.3 36.02 7.21 0 Pink Lady 16.91 21.28 8.52 6.87 21.95 28.83 41.59 33.14 11.04 9.88

DISCUSSION

Apple seeds contain 20% of fat expressed on dry matter. Fats from apple seeds belong into the group of linolic acid. The main fatty acid is linolic acid with the ratio 55-60%. Oleic acid represents from 23 to 30% of total fatty acid. Among unsaturated fatty acids, palmitic acid represents 8% and stearic acid 2% of total fatty acids. Arachidic acid represents 1.5% of total fatty acids present in apple seeds. Linolic acid is prevalent fatty acid on apple peel. Ratio of linolenic acid on peel is up to 30 fold higher than in seeds. Ratio of oleic acid is on the peel nearly the same as in seeds. Saturated fatty acids like palmitic and stearic is higher than in seeds. During storage of apples ratio of fatty acids is a matter of change. We have found the ratio of palmitic and oleic acid to increase during storage. That is not in concordance with the observation of Morice & Shortland (1973). They found a decrease of both fatty acids during storage of cultivars Dougherty, Granny Smith and Sturmer. Considerable changes occurred with linolic acid, which decreased sub- stantially during storage. Morice & Shortland (1973) found an increase of lino- lic acid during storage on above mentioned cultivars. Ratio of linolenic and stearic acid remained stable during storage what is not in accordance with Morice & Shortland (1973), who found stearic acid to increase and linolenic 202 VEGETABLE CROPS RESEARCH BULLETIN 66 ______acid to be stable during storage. Interestingly the ratio of saturated/unsaturated fatty acids on apple peel increased during storage at all cultivars except Idared.

CONCLUSION

Fat from apple seeds belong to the group of fats with prevailing linolic acid with ratio of 60%. Linolic acid is also prevalent on apple peel although with lower ratio. Apple peel contains up to 30 fold higher ratio of linolenic acid which is difference with the fatty acid composition in seeds. During stor- age of apples fatty acid composition is changing. Palmitic and oleic acid ratio increases, while that of linolic considerable decrease. As seen from other re- search papers fatty acids are known to play an important role in aroma volatiles production, while is their role in scald appearance not well elaborated.

REFERENCES

Brackmann A., Streif J., Bangerth F. 1995. Einfluss von CA-bzw. ULO- Lagerbedingungen auf Fruchtqualitat und Reife bei praklimakterisch ung klimak- terisch geerenteten Apfeln II: Auswirkung auf Ethylen-, CO2-, Aroma- und Fettsaurenproduktion. Gartenbauwissenschaft, 60, 1: 1-6. [in German with English summary] Fellman J.K., Miller T.W., Mattinson D:S., Mattheis J.P. 2000. Factors that influence biosynthesis of volatile flavour compounds in apple fruits. HortScience 35: 1026- 1033. Harb J., Streif J., Bangert F. 2000. Response of controlled atmosphere (CA) stored ‘Golden Delicious’ apples to the treatments with alcohols and aldehydes as aroma precursors. Gartenbauwissenschaft 65: 154-161. Kunst L., Samuels A.L. 2002. Biosynthesis and secretion of plant cuticular wax. Prog- ress in Lipid Research 42, 1: 51-80. Lu Y., Foo L.Y. 1997. Constitution of some chemical components of apple seeds. Food Chem. 61: 29-33. Garces M., Mancha M. 1993. One step lipid extraction and fatty acid methyl esters preparation from fresh plant tissues. Anal. Biochem. 211: 139-143. Marangoni A.G., Palma T., Stanley D.W. 1996. Membrane effects in postharvest physi- ology. Postharvest Biol. Technol. 7: 193-217. Meigh G.H., Hulme A.C. 1965. Fatty acid metabolism in the apple fruit during the res- piration climacteric. Phytochemistry 4: 863-871. Morice I.M., Shortland F.B. 1973. Composition of the surface waxes of apple fruits and changes during storage. J. Sci. Food & Agric. 24, 11: 1331-1339. Nelson D.L., Cox M.M. 2000. Lehninger principle of biochemistry. 3rd ed. New York, Mona group: 1152 p. Saquet A.A., Streif J., Bangert F. 2000. Changes in ATP, ADP and pyridine nucleotide level related to the incidence of physiological disorders in ‘Conference’ pears and ‘Jonagold’ apples. J. Hort. Sci. Biotechnol. 75: 243-249. Song J., Bangerth F. 2003. Fatty acids as precursors for aroma volatile biosynthesis in pre-climacteric and climacteric apple fruit. Postharvest Biol. & Technol. 30, 2: 113-121. M.SIMCIC et al. – INFLUENCE OF CULTIVAR AND STORAGE ... 203 ______

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WPŁYW ODMIANY I DŁUGOŚCI OKRESU PRZECHOWANIA NA ZAWARTOŚĆ WYŻSZYCH KWASÓW TŁUSZCZOWYCH W EPIDERMIE JABŁKA

Streszczenie Celem badań było określenie zawartości kwasów tłuszczowych w skórce i nasio- nach jabłka. Testy wykonywano na 7 odmianach jabłek: Elstar, Fuji, Gloster, Idared, Jonagold, Pink Lady i Golden Delicious. Nasiona jabłek zawierały od 8,4 do 15% wody i od 17,1 do 28,0% tłuszczów (w świeżej masie). Stwierdzono następujące zawartości poszczególnych kwasów tłuszczowych w nasionach: kwas linolowy (55,9-62,3%), kwas oleinowy (23,9-30,7%), kwas palmitynowy (7,1-9,3%), kwas stearynowy (1,6-2,3%), kwas arachidowy (1,2-1,6%), kwas linolenowy (0,4-0,7%). Kwas linolowy jest domi- nującym kwasem tłuszczowym również w skórce. Zawartość kwasu linolenowego jest do 30 razy wyższa w skórce niż w nasionach. Zawartość nasyconych kwasów tłuszczo- wych: palmitynowego i stearynowego, jest znacznie wyższa w skórce niż w nasionach, podczas gdy zawartość kwasu oleinowego jest zbliżona w skórce i nasionach. W czasie przechowywania jabłek, zawartość kwasu palmitynowego i oleinowego wzrasta nato- miast kwasu linolowego znacznie obniża się.