5. Hoffmeister, John E. 1974. Land from the Sea. Univ. of Miami 11. Popenoe, Wilson. 1920. Manual of Tropical and Subtropical Fruits. Press, Miami, FL. 143p. Hafner Press. 474p. 6. Ingram, Martha H. 1976. Crysophylum cainito—Star apple. The 12. Ruehle, G.D. and P. J. Westgate. 1946. Annual Rept., Fla. Agr. Propagation of Tropical Fruit Trees, (ed.) R. J. Garner et. al. Exp. Sta., Homestead, FL. Hort. Rev. No. 4, Commonwealth Agr. Bureaux, Farnham Royal, 13. Small, John K. 1933. Manual of Southeastern Florida. Hafner Pub. England, p. 314-320. Co., New York. 1554p. 7. Lendine, R. Bruce. 1952. The Naranjilla (Solanum quitoense 14. Stover, L. H. 1960. Progress in the Development of Grape Varieties Lam.) Proc. Fla. State Hort. Soc. p. 187-190. for Florida. Proc. Fla. State Hort. Soc. 73:320-323. 8. Long, Robert W. 1974. Origin of the vascular flora of South 15. Tomlinson, P. B. and F. C. Craighead, Sr. Growth-ring studies on Florida. In: Environments of South Florida (present and past), the native trees of sub-tropical Florida, p. 39-51. Research Trends in (ed.) Patrick J. Gleason, Miami Geological Soc, Miami, FL. p. 28- Plant Anatomy, K. A. Chowdhury Commemoration Volume 1972. 36. (eds.) A. K. M. Ghouse and Mohd. Yunus, New Delhi, India. 9. and O. Lakela. 1972. A Flora of Tropical Florida. Univ. 16. Winters, H. F. and Robert J. Knight, Jr. 1975. Selecting and breed of Miami Press, Miami, FL. 962p. ing hardy passion flowers. Am. Hort. 54(5):22-27. 10. Menzel, Margaret Young and F. O. Wilson. 1963. Allododecaploid 17. Zimmerman, G. A. 1941. Hybrids of the American pawpaw. J. of hybrid of Hibiscus diversifolius and some related J?1 Hybrids. /. of Hered. 32(3):83-91. Hered. 54(2):55-60.

Proc. Fla. State Hort. Soc. 92:298-300. 1979.

LiPID MARKERS IN CHEMOTAXONOMY OF TROPICAL FRUITS: PRELIMINARY STUDIES WITH CARAMBOLA AND LOQUAT H. E. Nordby and N. T. Hall gotic seedlings. Because these lipid markers have not been U.S. Citrus and Subtropical Products Laboratory,,1 investigated to any extent in subtropical and tropical fruits P. O. Box 1909, other than Citrus, we undertook this preliminary investiga Winter Haven, Florida 33880 tion on the lipid composition of two fruits, the carambola (Averrhoa carambola L.) and the loquat (Eriobotrya Abstract. Long-chain hydrocarbons, desmethyl sterols and japonica Lindl.). A study was conducted (12) previously on fatty acids have previously been demonstrated to be suit the organic acids, total soluble solids, flavor and texture able markers for the chemotaxonomy of citrus. We have of 18 carambola cultivars. extended these studies to include two tropical fruits: caram- bola (Averrhoa carambola L), 5 cultivars, and loquat (Erio- Materials and Methods botrya japonica Lindl.), 2 cultivars. The profiles ofthese Samples. Fruits from 5 cultivars of carambola and 2 three lipids in the 7 cultivars were determined by gas- cultivars of loquat were obtained from the USDA Sub liquid chromatography. C21 to C31 long-chain hydrocarbons tropical Horticulture Research Unit (Miami, Florida), were present in both species but C22 to C29 monoenes were frozen and stored at 0°F until extracted (within 2 weeks). present only in carambola. Major sterols for both fruits were Lipid Extraction. Four fruits of each carambola cultivar /3-sitosterol, campesterol, and isofucosterol in order of their were thawed and then individually cut, separated from their prevalence. Both fruits contained primarily the four major seeds and weighed (total fruit minus seeds). Fruits from each plant fatty acids—palmitic, oleic, linoleic and linolenic; how cultivar of loquat were pooled, separated from their seeds, ever, the ratios of these and 3 other acids were quite differ divided into four replicates and weighed (40 to 60 g). The ent in the two fruits. lipid of each sample was extracted with Folch reagent as reported by Nordby and Nagy for citrus lipid (6), weighed Chemotaxonomy is used in the delimitation of closely and stored at —40°F in benzene/ethanol, 4:1. related species, cultivars, selections, etc., of plants, fungi Fatty Acid Analyses. An aliquot of each lipid sample and other organisms. The markers generally used in these was evaporated to dryness on a rotoevaporator and studies include flavanoids, coumarins, phenolics, terpenes, transesterified with MeOH-NaOH-BF3; the resultant methyl long-chain hydrocarbons, sterols and fatty acids. The last esters were purified by thin-layer chromatography (TLC) 3 types of compounds are generally classified as lipids since and then analyzed by gas liquid chromatography (GLC) these compounds are soluble in organic solvents. Other than on 3% SP-1000 (11) and on 10% Apolar 10C at 160°C. reports of the fatty acid composition of seeds (2), very little Desmethyl Sterol Analyses. An aliquot of each lipid work has been done on the chemotaxonomy of tropical and sample was evaporated to dryness as above, saponified with subtropical fruits. 6% KOH and extracted into hexane (5). The desmethyl We have extensively studied the chemotaxonomy of the sterols were separated from the monomethyl sterols, di genus Citrus with these lipid markers (3, 7-11), and observed methyl sterols, hydrocarbons and other nonsaponifiables by trends in the lipid compositions of the various cultivars. silica gel TLC with chloroform as the developing solvent Each citrus species had intrinsic profiles, and hybrids (5). Underivatized sterols were analyzed by GLC on a 1% generally had profiles characteristic of both parents. Pro SP-1000 column at 220°C (5). files of nucellar seedlings were different from those of zy- Hydrocarbon Analyses. The hydrocarbons eluted from the TLC platings of the nonsaponifiables, were analyzed by iSouthern Region, U. S. Department of Agriculture, Science and programmed GLC in which monounsaturates (monoenes) Education Administration. were resolved from saturates(alkanes) (10). For verification Mention of a trademark or proprietary product is for identification of the monoene fractions, the total hydrocarbon fraction only and does not constitute a guarantee or warranty of the product by the U. S. Department of Agriculture and does not imply its ap was subjected to silver nitrate TLC (10), the isolated proval to the exclusion of others which may also be suitable. monoenes were hydrogenated (4), and the two saturated

298 Proc. Fla. State Hort. Soc. 92: 1979. fractions (original alkanes and the hydrogenated alkenes) Table 4. Relative percentages of alkanes in loquat fruit. were analyzed by GLC (10). Values in Tables 1-4 are the means of the four replicate Cultivar analyses for fatty acids, desmethyl sterols and hydrocarbons, Alkane M-18553 20 respectively.

Table 1. Relative percentages of major fatty acids in carambola and 21 0.5 1.8 loquat fruits. 22 0.4 0.5 23 0.7 0.6 24 0.3 0.4 Fatty acidsz 25 0.7 0.6 Cultivar 14 16 16:1 18 18:1 18:2 18:3 26 0.1 0.4 27 12.1 9.8 28 2.0 2.0 Carambola 29 76.9 77.8 37 4.1 17.2 2.1 2.1 51.3 9.0y 14.2 30 1.3 0.9 44 6.1 16.7 2.1 1.8 51.8 9.2y 12.3 31 5.1 5.2 42 4.9 15.7 2.4 2.0 54.7 8.0y 12.3 Tean Ma 2.3 15.5 2.1 1.9 56.0 8.5y 13.7 17 7.1 15.4 2.1 1.6 50.9 10.4y 12.5 species are difficult to separate into edible (flesh) and non- edible portions, we used entire fruit minus its seeds in Loquat these taxonomic studies. No attempt was made to determine M-18553 0.9 22.5 0.8 7.0 20.0 36.9 11.9 20 0.9 24.4 0.5 8.7 13.7 38.5 13.3 the location of specific lipids. Generally, hydrocarbons are derived from the wax coating of fruits, sterols from the structural membrane and fatty acids from storage tissue ^Number of carbons in chain: number of double bonds. yMixture of linoleate and another 18:2 acid of unknown structure. as a component of triglycerides. Programmed GLG analyses revealed fatty acids from Table 2. Relative percentages of desmethyl sterols in carambola and C12 to C26 in both fruits; however, only the seven C14 to loquat fruits. C18:3 fatty acids listed in Table 1 were present at relative percentages greater than one. There were only minor

Sterol differences between the fatty acid profiles of the five caram- Cultivar Cholesterol Campestero]I /?-Sitosterol 1 sofucosterol bolas, the differences being greatest for G14 and oleic acid (C18:1). Tean Ma and cultivar 42 had from 3.8 to 5.1 per centage points more oleic acid than the other three culti Carambola vars, whereas Tean Ma and cultivar 17 had the lowest and 37 0.5 18.3 75.8 5.4 44 0.5 14.8 80.1 4.6 highest values for C14 of the five carambolas. In Citrus, fatty 42 0.5 21.6 73.1 4.8 acid profiles of specific lipids in the fruit were more useful Tean Ma 0.5 16.2 79.0 4.3 for taxonomic studies than were profiles of other lipids (7).. 17 0.4 13.1 80.4 6.1 The same may be true for carambola, but further studies Loquat are required to determine whether the oleic acid triglyceride M-18553 1.5 3.1 90.4 5.0 levels in Tean Ma and cultivar 42 were significantly different 20 1.0 1.9 93.5 3.6 than those in the other three cultivars. In all five carambola cultivars the values for C18:2 in clude two fatty acids that could not be resolved on the Table 3. Relative percentages of saturated (alkanes) and monoun- saturated (alkenes) hydrocarbons in carambola fruit. Apolar-lOC GLG phase but were partially resolved on SP- 1000. The ratio of the common C18:2 fatty acid (linoleic acid) to the unknown C18:2 acid was approximately 3:2. The Alkanes structure of this acid was not determined; however, its Cultivar 23 24 25 27 29 31 Other z 21 22 presence in carambola flesh and absence in its seeds (1) would make this acid a useful marker in further breeding 37 3.5 3.5 29.9 4.6 28.2 6.8 18.6 2.7 2.2 and chemotaxonomic studies. 44 3.8 4.6 31.8 5.3 30.4 7.1 13.5 1.2 2.3 The fatty acid profiles of the loquat cultivars also 42 2.0 4.0 34.3 6.1 33.5 7.9 9.1 1.2 1.9 Tean Ma 1.4 6.1 38.0 5.0 29.2 6.6 8.9 3.2 1.6 were similar except for their percentages of oleic acid. 17 2.1 3.4 33.0 7.7 31.9 6.3 10.9 3.3 1.4 Desmethyl sterols in most higher plants consist mainly

Alkenes of C28 and C29 sterols and traces of C27 sterols. Garambola and loquat fruit followed this general trend, with the 22 23 24 25 27 29 C27 sterol cholesterol making up less than 2% of the total 37 0.6 51.6 2.8 34.9 6.6 3.5 sterols (Table 2). Stigmasterol, a C29 sterol present in most 44 2.0 48.8 3.6 35.1 7.5 3.0 42 1.0 49.6 2.4 35.0 8.6 3.4 higher plants, was not observed in either carambola or Tean Ma 0.6 46.2 0.8 38.9 10.2 3.3 loquat fruit. However, isofucosterol, another C29 sterol was 17 1.2 48.4 3.6 33.4 9.2 4.1 detected in both species at a relative percentage that is commonly observed for stigmasterol in higher plants. Iso ^Includes less than 1% each of C , C and C . fucosterol differs from stigmasterol only in the position of one of its two double bonds. Campesterol in carambola Results and Discussion showed the greatest range of values (8.5 percentage points). Cultivars with high /3-sitosterol values had lower campes The lipids extracted from carambola fruit (as per terol values than cultivars with low /3-sitosterol values; very centage of the fresh fruit) varied very little between the little difference was observed in the values for the other two five cultivars (0.12 ±: .01). The lipid in loquat (0.24 ± sterols. The two loquat cultivars had unusually high values .01) was twice the amount found in carambola; however, for /3-sitosterol, with cholesterol, campesterol and isofuco the two species had similar ratios of nonsaponifiable ma sterol levels each less than 5%. terial, fatty acid and residue. Because the fruits of both The alkane profiles of the carambola cultivars followed

Proc. Fla. State Hort. Soc. 92: 1979. 299 the general pattern for alkanes in higher plants: 87 to 90% may be useful as a marker in crosses of this species. 4) Iso- of the C21 to C31 alkanes had carbon skeletons with odd fucosterol was present and stigmasterol absent in both fruit. numbers of carbon atoms (Table 3). Branched alkanes, 5) The campesterol//3-sitosterol ratio was different for each which are prevalent in citrus (3, 10) were not observed in of the 7 cultivars. 6) The ratios of total alkenes to total carambola or loquat fruit. Of the even numbered alkanes alkanes are different in the five carambola cultivars while only C22 and C24 were observed at levels higher than 1%. in loquat alkenes were not detected. 7) Percentages of C29 The greatest differences in relative percentages between culti- and to a lesser degree C23, C25 alkanes were different for vars occurred with C29, and to a lesser degree with G23 and the five carambola cultivars. C25 alkanes. In most cases, a low level of one of these alkanes The data in this report and previous studies with citrus was compensated for by high levels in one or both of the (8-11) indicate that of the three lipids, hydrocarbons show others. the most potential as markers in chemotaxonomy of tropical Carambola fruit contained monounsaturated hydro fruits. Hydrocarbons are easily isolated, very stable and very carbons (alkenes) in addition to their alkanes (Table 3). adaptable to analyses by GLC. Alkenes accounted for 38% (cultivar 37), 40% (cultivars 17 and 44), 46% (cultivar 42) and 57% (Tean Ma) of the total Literature Cited hydrocarbons in a fruit. The majority (94 to 99%) of these 1. Berry, S. K. 1978. The composition of the oil of starfruit (Averrhoa were odd numbered with C23:1 and C25:1 accounting for carambola, Linn.) seeds. /. Am. Oil Chemists Soc. 55:340-341. 82 to 87% of the alkenes. Differences in alkene profiles for 2. Hilditch. T. P. 1947. The component acids of vegetable fats. Ch. the five cultivars were small. Differences occurred however, 4 in T. P. Hilditch, ed. The Chemical Constitution of Natural Fats. between the ratios of C23:1 and C25:1 alkenes to their respec John Wiley and Sons, New York. 3. Nagy, S. and H. E. Nordby. 1971. Distribution of free and conju tive C23 and C25 alkanes. These differences accounted, to a gated sterols in orange and tangor juice sacs. Lipids 6:826-830. large extent, for the total hydrocarbon percentages of the 4. and . 1973. Saturated and monounsaturated higher alkenes in cultivar 42 and Tean Ma. longchain hydrocarbon profiles of sweet oranges. Phytochemistry Alkane profiles of loquat fruit were quite unique with 12:801-805. 5. , and L. Telek. 1978. Lipid distributions in three odd numbered (C27, C29, C31) making up 92 to 94% green leaf protein concentrates from four tropical leaves. /. Agric. and each of the other alkanes less than 2% of the total Food Chem. 26:701-706. alkanes (Table 4). Differences between the two cultivars 6. Nordby, H. E. and S. Nagy. 1969. Fatty acid profiles of citrus juice were minimal. and seed lipids. Phytochemistry 8:2027-2038. 7. and . 1971. Comparative citrus fatty acid No attempt was made to correlate the differences in profiles of triglycerides, monogalactosyl diglycerides, steryl esters these three markers with the differences in oxalic acid, and esterified steryl glucosides. Lipids 6:554-561. ascorbic acid, and total acid contents reported for four 8. and . 1974. The relationship of longchain of the carambola cultivars (12). In this preliminary chemo- hydrocarbons to the chemotaxonomy of citrus. Proc. Fla. State Hort. Soc. 87:70-74. taxonomic study with carambola and loquat fruit, various 9. and . 1977. Hydrocarbons from epicuticular differences in lipid profiles were observed which may be waxes of citrus peels. Phytochemistry 16:1393-1397. applicable to other tropical fruits as well. 1) The caram 10. , and J. M. Smoot. 1979. Selected leaf wax bola and loquat fruit had fatty acid, sterol and hydrocarbon alkanes in chemotaxonomy of citrus. /. Amer. Soc. Hort. Sci. 104: 3-8. profiles intrinsic to their respective species. Thus, hybrids 11. , and . 1979. Relationship of root- would most likely have their own intrinsic profiles. 2) Oleic stock to leaf and juice lipids in citrus. /. Amer. Soc. Hort. Sci. acid showed the greatest differences between cultivars for 104:280-282. both fruit. 3) A C18:2 fatty acid with its double bonds in 12. Wagner, C. J., Jr., W. L. Bryan, R. E. Berry and R. J. Knight, Jr. 1975. Carambola selection for commercial production. Proc. Fla. positions other than 9, 12 is present in carambola fruit and State Hort. Soc. 88:466-469.

Proc. Fla. State Hort. Soc. 92:300-303. 1979.

PROMISING MEXICAN GUAVA SELECTIONS RICH IN VITAMIN C1 S. Lakshminarayana Abstract. The study reports the occurrence of several Departamento de Biotecnologia, variations in the fruit of Mexican guava with regard to size, Universidad Autonoma Metropolitana—Iztapulapa, color, pulp-seed ratio, vitamin C content and various other Apartado Postal 55-535, Mexico—13 D.F. morphological and chemical parameters. From amongst several trees producing apparently different types of fruits M. A. Moreno Rivera in 2 principle production centers 10 tentative selections were Departamento de Industrias Agricolas, made based on the vitamin C content of fruits and their Escuela Nacional de Agricultura (U.A.Ch), Chapingo, botanical and physicochemical characteristics. Fruits from 9 Estado de Mexico, Mexico of these selections had a vitamin C content of 500 mg or more per 100 g of pulp and 1 had more than 1000 mg. Additional index words. Psidium guajava L., types, selec Vegetative propagation of these is suggested to study their tion, cultivars, fruit weight, pulp-seed ratio, ascorbic acid, behaviour in future generations for a possible recommenda chemical and minerals constituents. tion as commercial cultivars.

iThe Authors wish to thank Ing. S. Sanchez Colin, Director General Guava (Psidium guajava L.) is one of the most important of CONAFRUT/S.A.G., Mexico (1971-76) for his keen interest and encouragement and Ings. I. Ortiz \ Robles and L. Sarmiento L6pez fruit crops of Mexico with a production of 161,115 MT for the technical assistance. distributed over an area of 12,148 ha valued ca. 250 million

Proc. Fla. State Hort. Soc. 92: 1979.