Walton B. Sinclair

Ether-Soluble Organic Acids and Buffer Properties of Citrus Peels

1. The concentrations of ether-soluble organic acids are much lower in peels of citrus fruits (oranges, grapefruit, and lemons) than in juice samples of corresponding pulps. The high pH values of the peel saps indicate that most of the organic acids are present in the salt form and not in the free state. The amounts of citric and malic acids in the expressed saps agreed satisfactorily with the total organic acids titrated between pH 7.8 and 2.6. The organic acids precipitated from the peel sap with lead acetate agreed with the total organic acids titrated between pH 7.8 and 2.6 on a water solution of the ether extract. The citric and malic acids values of the ether extract agreed substantially with similar values of the lead acetate precipitate. The ether liquid extraction method is satisfactory for the isolation of organic acids from citrus-peel saps. 2. The total nonvolatile organic acids were extracted from dried peel of citrus fruits with absolute ether at pH 1. The organic acid content consisted chiefly of citric, malic, and oxalic acids. The citric acid content is very low in comparison with the concentrations of malic and oxalic acids. The oxalic acid is present in the peel as insoluble calcium oxalate, with the exception of a trace occurring in the peel sap. 3. The titration curves of peel saps of citrus fruits (navel and Valencia oranges, grapefruit, and lemons) show that these systems contain comparatively small amounts of organic acids and their salts. In the absence of these constituents, the pH values of the peels are high, and these systems consequently have only a slight buffer capacity.

Carbohydrate Fractions of Grapefruit Peel

1. Whole peel of grapefruit was extracted with 80% ethyl alcohol and separated into two distinct fractions, the alcohol-soluble and alcohol-insoluble solids. The carbohydrate constituents of these two fractions were investigated, and certain of their physical and chemical properties were determined. All analyses were made on fruit samples obtained from trees that had been fumigated with hydrocyanic acid or sprayed with oil for pest control during the growing season. 2. The alcohol-soluble fraction of the peel contained substances which liberated a small amount of carbon dioxide on hydrolysis with 12% hydrochloric acid. The total solids of this fraction composed 66.31% and 67.35% of the dry matter of the peel of fumigated and oil-sprayed fruit, respectively. 3. The soluble sugars in the peel averaged 40.29% and 41.28% of the dry weight of the fumigated and oil-sprayed fruit, respectively. These amounts of soluble sugars accounted for 60.76% and 61.28% (mean values) of the total alcoholic extractives. Approximately 40% of the dry weight of the alcohol-soluble fraction was composed of substances other than sugars, namely, essential oil, waxes, organic acids, naringin, and various concentrations of undetermined constituents. 4. The alcohol-insoluble fraction of the peel contains the cell-wall constituents-cellulose, hemicellulose, and pectin. This fraction comprised 33.69% and 32.65% of the dry weight of the peel. Lignin and starch were not present in sufficient quantities to be determined by the prevailing chemical methods. 5. Since the alcohol-insoluble fraction contains the pectin, most of the carbon dioxide which can be liberated by hydrolysis with 12% HCl and all the methoxyl groups occur in this fraction. The ratio of methoxyl to carbon dioxide was much lower in grapefruit peel than in lemon peel. 6. The carbon dioxide equivalent to the sum of the esterified and non-esterified carboxyl groups was equal to the total carbon dioxide of the alcohol-insoluble fraction obtained on hydrolysis with 12% hydrochloric acid. 7. The sum of the water-soluble and acid-soluble pectin (as calcium pectate) of the alcohol-insoluble fraction of the peel of the fumigated and oil-sprayed fruit amounted to 43.79% and 43.95%, respectively. These values are much lower than those for the total pectin calculated from the carbon dioxide and the methoxyl values. 8. To establish criteria of purity, the calcium pectate values are accompanied by corresponding values for carbon dioxide, calcium, and furfural. The percentages of calcium in the calcium pectates are somewhat higher than the 7.50% usually reported for pure calcium pectate. The percentages of carbon dioxide are slightly higher than the 17.40% reported for pure calcium pectate. The furfural values are in accord with those reported by other investigators. 9. The residue that remains after extracting the pectin from the alcohol-insoluble solids of the peel is composed of cellulose and hemicellulose and a comparatively small amount of firmly bound pectin that is difficult to extract and determine quantitatively. 10. The sum of the carbon dioxide in the aqueous extract, acid extract, and residue amounted to 94.26% and 95.19% of the total carbon dioxide of the alcoholinsoluble solids in the peel of fumigated and oil-sprayed fruit, respectively. In the aqueous and acid extracts, calcium pectate accounted for more than 90% of the carbon dioxide dissolved by the extractants.

A Quantitative Paper-Chromatographic Study of the Amino Acid Composition of Proteins and Juice of the Orange

1. A quantitative paper-chromatographic analysis of the amino acids in proteins isolated from Washington Navel and Valencia orange fruit indicates that both qualitative and quantitative differences exist between the proteins from the two sources. 2. A similar analysis of juice from these two fruits indicates that not all the amino acids found in the protein could be detected in the free form in the juice. There were also marked differences in amino acid and amide composition of the two juices, but they did not reflect the differences in the composition of the two proteins.

Changes in Pectic Constituents of Valencia Oranges During Growth and Development

1. The alcohol- and acetone-insoluble solids were recovered periodically from peel and pulp of Valencia oranges. These fractions were analyzed for total pectic substances, using the CO2 method. The results of the two were practically the same. The water-soluble pectins were extracted from the alcohol-insoluble solids and determined colorimetrically by the carbazole method, which analyzes directly for the total anhydrogalacturonic acid units present. A comparison of this method was made with the calcium pectate method, which precipitates the pectic substances down to a limited polymer size and which, because of purity, must be further analyzed by means of CO2 determinations, to determine the uronide content. 2. Seasonal changes in concentrations of pectic substances in peel, pulp, and juice of Valencia oranges were followed by determining the total and the water-soluble pectic substances as anhydrogalacturonic acid during growth and development under normal conditions. After a tremendous initial increase in the total pectic substance in peel and pulp, there is a gradual decrease throughout the remainder of the season. The water-soluble pectins of both peel and pulp also show a gradual decline, but, when observed in relation to the total pectic substances of the corresponding component, the percentage is found to increase in the peel and decline in the pulp before eventually increasing also. The pectic substances found in the juice remain relatively constant throughout.

Polyuronide Fraction and Soluble and Insoluble Carbohydrates of Orange Peel

1. The whole peel of Valencia oranges and the albedo of navel oranges were separated into alcohol-soluble and alcohol-insoluble fractions by extraction of the fresh material with hot 80% ethyl alcohol. The principal constituents of the two fractions were determined, and some of their chemical properties studied. 2. The alcohol-soluble fraction of the whole peel of Valencia orange and of the albedo of navel orange averaged 57.31% and 55.19%, respectively, of the total dry weights. Soluble sugars accounted for 55.4% of the total extractable material in Valencia peel and for 72.69% in navel albedo. This fraction also contained substances which produced a small amount of carbon dioxide on hydrolysis with 19% hydrochloric acid. This carbon dioxide was probably produced by low polymer galacturonides soluble in hot 80% alcohol. 3. The alcohol-insoluble fraction of Valencia orange whole peel and of navel albedo averaged 42.69% and 44.81%, respectively, of the total dry weights. The sum of the water-soluble and acid-soluble pectic substances (as calcium pectate) of the alcohol-insoluble fraction averaged 39.76% in Valencia peel and 34.87% in navel albedo; the insoluble residues averaged 30.53% and 30.41%, respectively. The undetermined portion consisted of hemicelluloses which contained small amounts of polyuronides and which were rendered soluble by the extraction treatment. 4. The purity of the extracted pectic material (as calcium pectate) was ascertained on each sample by determining contents of carbon dioxide, calcium, and furfural. 5. As the polyuronides yield carbon dioxide quantitatively on hydrolysis with hydrochloric acid, the carbon dioxide values determined are a fair measure of the total pectic substances in the alcohol-insoluble solids. When the carbon dioxide values were accompanied by corresponding values for methoxyl, the degree of methylation of the pectic substances was subsequently calculated by dividing the determined methoxyl by that equivalent to the total carbon dioxide. Methylation in Valencia peel averaged 83.38%, and in navel albedo, 82.39%. 6. The sum of the carbon dioxide in the aqueous extract, acid extract, and residue amounted to 95.20% and 97.47% (mean values) of the total carbon dioxide of the alcohol-insoluble solids in the Valencia orange peel and navel orange albedo, respectively. The total calcium pectate precipitated from the aqueous and acid extracts of Valencia peel yielded 94.12% of the carbon dioxide dissolved by the extractants. The corresponding value for navel albedo was 88.00%. 7. Since the alcohol-insoluble fraction contained the pectic substances, most of the carbon dioxide which could be liberated by hydrolysis with 19% hydrochloric acid, and all the methoxyl groups, occurred in this fraction. The ratio of methoxyl to carbon dioxide was approximately the same in Valencia orange peel and in navel orange albedo. 8. The carbon dioxide equivalent to the sum of the esterified and nonesterified carboxyl groups was equal to the total carbon dioxide of the alcohol-in-soluble fraction obtained on hydrolysis with 19% hydrochloric acid.

Alcohol-Insoluble Solids of Juice Vesicles and Pulp of Citrus Fruit

1. The alcohol-insoluble fraction of the juice vesicles of Valencia and navel oranges, and of the pulp of navel oranges and lemons, has been isolated from the fresh material by extraction with 80% ethyl alcohol. This fraction, which contained the cellulose, most of the hemicellulose, and pectin, was lowest in Valencia orange vesicles (mean, 9.87%) and highest in navel orange pulp (mean, 15.56%), on a dry-weight basis. 2. Since the alcohol-insoluble fraction contains the pectin, most of the carbon dioxide which can be liberated by hydrolysis with 12% HCl, and all the methoxyl groups, occur in this fraction. The ratio of methoxyl to CO2 was much lower in Valencia orange vesicles than in navel orange vesicles, navel orange pulp, or lemon pulp. 3. The carbon dioxide equivalent to the sum of the esterified and non-esterified carboxyl groups was equal to the total carbon dioxide of the alcohol-insoluble fraction obtained on hydrolysis with 12% hydrochloric acid. 4. The sum of the water-soluble and acid-soluble pectin (as calcium pectate) of the alcohol-insoluble fraction of the vesicles and pulp samples ranged between means of 26.30% in navel orange vesicles and 36.47% in lemon pulp. These values are much lower than those for the total pectin calculated from the carbon dioxide and methoxyl values. 5. To establish criteria of purity, the calcium pectate values are accompanied by corresponding values for carbon dioxide, calcium, and furfural. The percentages of calcium in the calcium pectates are generally somewhat higher than the 7.50% usually reported for pure calcium pectate, and the percentages of carbon dioxide are generally slightly higher than the 17.40% reported for the pure compound. The furfural values are in accord with those reported by other investigators. The ratios of calcium to carbon dioxide of the calcium pectates from the water-soluble pectin show satisfactory agreement with the theoretical value; except in lemon pulp, the ratios of these substances from the acid-soluble pectin tend to be lower than the theoretical value. 6. The residue that remains after extracting the pectin from the alcohol-insoluble solids of the vesicles and pulp is composed of cellulose and hemicellulose and a comparatively small amount of firmly bound pectin that is difficult to extract and determine quantitatively. 7. The sum of the carbon dioxide in the aqueous extract, acid extract, and residue amounted to 95.4% and 100.9% (mean values) of the total carbon dioxide of the alcohol-insoluble solids in the vesicles and pulp, respectively. In the aqueous and acid extracts, calcium pectate accounted for more than 85% of the carbon dioxide dissolved by the extractants. 8. As a measure of the degree of esterification of the alcohol-insoluble fractions, the ratio of methoxyl to galacturonic acid was lowest in the Valencia orange vesicles (mean, 0.107) and highest (mean, 0.134) in navel orange pulp. In a polygalacturonic acid, the number of carboxyl groups that are free, esterified, or combined with cations is not a constant amount but varies considerably with the method of preparation.

Pectic Substances of Juice Vesicles of Grapefruit

1. Juice vesicles of grapefruit were separated into alcohol-soluble and alcohol-insoluble fractions by extraction with 80% ethyl alcohol. The pectic substances, which are insoluble in alcohol, were thereby separated from the soluble carbohydrates. The alcohol-insoluble solids averaged 7.39% of the dry weight and 0.79% of the fresh weight of the vesicles. All analyses were made on this material. 2. Carbon dioxide was determined quantitatively by boiling with 19% HCl, as a measure of the uronide content. Total carboxyl groups, esterified and nonesterified, were titrated also as a check on CO2 determinations. Results were in close agreement. 3. Methoxyl determinations were made and reported both as percentages of the alcohol-insoluble solids and as percentages of the total carboxyl groups esterified. 4. Furfural is produced by uronic acid anhydrides, as well as by pentoses and pentosans, by boiling with a mineral acid. Recovery is not quantitative but is constant for a definite procedure. Total furfural determined was 12.97% of the alcohol-insoluble solids. Furfural equivalent to the uronic anhydrides was, by calculation, 6.65%, leaving 6.32% undetermined. Expressed as pentosans, it amounted to 10.86%. 5. Pectic substances were obtained by extracting the material, first with hot water, and then with hot 0.1 N HCl. Total pectic material in each extract was determined by precipitating and weighing as calcium pectate. Calcium pectate from water-soluble pectic substances amounted to 11.78% of the alcoholinsoluble solids, and from acid-soluble material it amounted to 24.81%. The purity of the calcium pectates is indicated by accompanying values for carbon dioxide, calcium, and furfural. Percentages of both carbon dioxide and calcium were higher in the water-soluble calcium pectate than in the acid-soluble extract. Furfural values were nearly the same in the two calcium pectates. It was concluded that either carbon dioxide or calcium determinations could be used to calculate the galacturonide content of a calcium pectate.