Capillary electrophoresis for evaluating orange juice authenticity: a study on Spanish oranges

Fruit juices have very distinct organic acid profiles that can be used as fingerprints for establishing possible adulteration. Recently, our group developed and validated a capillary electrophoresis method using UV detection for determining citric, isocitric, tartaric, and malic acids in natural and commercial orange juices. Sample treatment consisted of only dilution and centrifugation or filtration. This method has been applied to evaluate these acids and their ratios in 63 samples of Navelina, the most common variety of Spanish oranges, over a three month period. This evaluation has been conducted to establish ranges of acid concentrations and to compare them with those found in commercial juices. The more reliable parameter, because of the lower variability in fresh samples, was found to be the citrate/isocitrate ratio with a value of 113 (RSD = 10%). Only one of nine ramdonly selected commercial juices presented values within the range of those of the population of just-pressed Navelina orange juice. Moreover, three of them had measurable tartrate values, which is not a natural component of orange juice, showing mixtures with cheaper fruits.     

Identification of TLC markers and quantification by HPLC-MS of various constituents in noni fruit powder and commercial noni-derived products

The composition of noni (Morinda citrifolia) products has been investigated. TLC profiles of several commercial juices and capsules were compared. 3-Methyl-1,3-butanediol was identified as a typical marker in noni juices. The presence of sorbic acid (E200) was detected in one juice declared as additive free. Quantitative data have been obtained by HPLC-MS. A method for the quantification of characteristic noni constituents, such as iridoid glucosides, scopoletin, rutin, fatty acid glucosides, and anthraquinones, was developed and validated. The separation was performed on a C18 column with a gradient of acetonitrile in water containing 0.1% formic acid. Detection was carried out with ESI-MS in the negative ion mode. Significant differences were observed between the products. Asperulosidic acid, deacetylasperulosidic acid, and rutin were present in all samples analyzed, but their concentrations differed considerably between the products. Fatty acid glucosides, noniosides B and C, were present in capsules and most juices. Scopoletin was mainly found in juices. The anthraquinone alizarin, which has been reported from roots and leaves, was not detected in the samples investigated.     

Gas-liquid chromatographic determination of benzoic acid and sorbic acid in foods: NMKL collaborative study

A gas-liquid chromatographic method for the simultaneous determination of benzoic acid and sorbic acid in foods was collaboratively studied by 8 laboratories. Benzoic and sorbic acids are isolated from food by successive extractions with ether, sodium hydroxide, and methylene chloride, converted to trimethylsilyl (TMS) esters, and determined by gas-liquid chromatography. Phenylacetic acid and caproic acid are used as internal standards for benzoic acid and sorbic acid, respectively. Seven samples were collaboratively studied: almond paste, fish homogenate, and apple juice with benzoic and sorbic acid levels from 0.04 to 2 g/kg. Average recoveries (%) for benzoic and sorbic acids were as follows: almond paste, 99.6 and 101.2; fish homogenate, 99.2 and 97.4; and apple juice 98.2 and 106.6. The reproducibility coefficients of variation (%) for benzoic and sorbic acids at 0.5-2 g/kg levels were 3.5-6.1 and 5.2-9.0; and at the 0.04 g/kg level, 14.7 and 23.3, respectively. The method has been adopted official first action at 0.5-2 g/kg levels.     

Extraction of organic acids by ion-pair formation with tri-n-octylamine. Part 6. Determination of sorbic acid, benzoic acid, and saccharin in yogurt

The sorbate content of commercial yogurt samples is determined by reverse phase liquid chromatography following ion-pair extraction with tri-n-octylamine. Mean recoveries (70-88%), precision (1.1-3.3% RSD), and detection limit of the method are presented for sorbic acid, benzoic acid, and saccharin.     

Rapid quantitative method for simultaneous determination of benzoic acid, sorbic acid, and four parabens in meat and nonmeat products by liquid chromatography

A liquid chromatographic (LC) method for the simultaneous determinations of benzoic acid, sorbic acid, and methyl, ethyl, propyl, and butyl parabens (methyl, ethyl, propyl, and butyl-p-hydroxybenzoates) in meat and nonmeat products was developed. Benzoic acid, sorbic acid, and parabens were extracted from meat and nonmeat products with 70% ethanol. After filtration, extracts were analyzed by reverse phase liquid chromatography. Homogeneously ground samples of fresh sausage and hamburger were fortified with benzoic acid, sorbic acid, and each paraben at 5 different concentrations. Average recovery (after discarding outliers) for each preservative at all 5 levels was greater than 95% with a coefficient of variation less than 5%.     

Simultaneous gas chromatographic determination of carboxylic acids in soft drinks and jams

A method was developed for simultaneous gas chromatographic determination of sorbic acid, dehydroacetic acid, and benzoic acid used as preservatives, and succinic acid, fumaric acid, malic acid, and tartaric acid used as acidulants in soft drinks and jams. A sample was dissolved in NH4OH-NH4Cl pH 9 buffer solution, and an aliquot of the solution was passed through a QAE-Sephadex A 25 column. The column was washed with water, and the carboxylic acids were eluted with 0.1N HCl. Sorbic acid, dehydroacetic acid, and benzoic acid were extracted with ethyl ether-petroleum ether (1 + 1), and determined on a 5% DEGS + 1% H3PO4 column. Succinic acid, fumaric acid, malic acid, and tartaric acid in the lower layer were derivatized with N,O-bis(trimethylsilyl)acetamide and trimethylchlorosilane, and determined on a 3% SE-30 column. Recoveries from soft drink and jam samples fortified with 0.1% each of 7 carboxylic acids ranged from 92.4 to 102.6% for preservatives, and from 88.1 to 103.2% for acidulants.     

Chemometric characterization of fruit juices from Spanish cultivars according to their phenolic compound contents: I. Citrus fruits

The data set composed by phenolic compound profiles of 83 Citrus juices (determined by HPLC-DAD-MS/MS) was evaluated by chemometrics to differentiate them according to Citrus species (sweet orange, tangerine, lemon, and grapefruit). Cluster analysis (CA) and principal component analysis (PCA) showed natural sample grouping among Citrus species and even the Citrus subclass. Most of the information contained in the full data set can be captured if only 15 phenolic compounds (concentration ≥10 mg/L), which can be quantified with fast and accurate methods in real samples, are introduced in the models; a good classification which allows the confirmation of the authenticity of juices is achieved by linear discriminant analysis. Using this reduced data set, fast and routine methods have been developed for predicting the percentage of grapefruit in adulterated sweet orange juices using principal component regression (PCR) and partial least-squares regression (PLS). The PLS model has provided suitable estimation errors.     

Measurement of soluble solids contents and pH in orange juice using chemometrics and vis-NIRS

The potential of visible and near-infrared reflectance spectroscopy (vis-NIRS) was investigated for its ability to nondestructively detect soluble solids contents (SSC) and pH in orange juices. A total of 104 orange juice samples were used for vis-NIRS at 325-1075 nm using a field spectroradiometer. Wavelet packet transform, standard normal variate transformation (SNV), and Savitzky-Golay first-derivative transformation were applied for the preprocessing of spectral data. The chemometrics of partial least-squares (PLS) regression analysis was performed on the processed spectral data. The evaluation of SSC and pH in orange juices by PLS regression with SNV showed the highest accuracy of the three preprocessing methods. The correlation coefficient (r), standard error of prediction, and the root-mean-square error of prediction for SSC were 0.98, 0.68, and 0.73, respectively, whereas those values for pH were 0.96, 0.06, and 0.06, respectively. The "fingerprint" representing features of orange juices or reflecting sensitivity to some elements at a certain band was proposed on the basis of regression coefficients. It is very useful in the field of food chemistry and further research on other materials. It is concluded that the vis-NIRS technique combined with chemometrics is promising for the fast and nondestructive detection of chemical components in orange juices or other materials.     

Phloroglucinol from phlorin hydrolysis for testing quality of commercial orange juices and beverages

The European Association of Juices and Nectars Producers (AIJN) is evaluating the opportunity to introduce the content of phlorin (3,5-dihydroxyphenyl beta-D-glucopyranoside), a peel marker for oranges, as a parameter for testing the quality of orange juices. Because of the lack of a commercial standard of phlorin and its laborious isolation procedures, in this contribution is developed a simple and reliable method for measuring the phlorin level as the corresponding aglycon phloroglucinol, obtained after a total enzymatic hydrolysis of the sample. The method was applied to the quantification of phloroglucinol in several industrial and commercial blond and pigmented orange juices and beverages based on 12% orange juice. Under the same extraction procedure, the phloroglucinol content in the pigmented juices was higher than in the blond ones. No significant difference was obtained between not from concentrate juices and reconstituted from concentrate juices. The marker amount increases in the highly processed orange fruits and in the byproducts of citrus processing due to the contact of the juice with the albedo, which is the major source of phlorin. In orange-based beverages the phloroglucinol content revealed a large heterogeneity and a poor quality of the raw juices used.     

Multielement analysis of argentinean lemon juices by instrumental neutronic activation analysis and their classification according to geographical origin

Multielement analysis of lemon juices from different Argentinean regions was carried out by instrumental neutronic activation analysis (INAA) with the aim at developing a reliable method in the traceability of the origin of lemon juices. This work presents a characterization of 44 lemon juice samples selected from three different geographical origins in the Northwest region of Argentina. Multivariate chemometric techniques such principal component analysis and lineal discriminant analysis (LDA) were used with the aim of classifying the juices and identifying the most significant parameters. Eleven elements were determined (Br, As, Na, Rb, La, Cr, Sc, Fe, Co, Zn, and Sb). The analytical method was validated by analyzing the standard reference material IAEA V-10 (hay powder); the results are within +/-10% of the reported values for the majority of the elements. Biplots of LDA scores for the INAA data illustrate clear separation between each sample.     

Orange, mandarin, and hybrid classification using multivariate statistics based on carotenoid profiles

A study was undertaken to differentiate citrus on the basis of a carotenoid profile obtained from the HPLC determination of 12 carotenoids found in saponified fresh juice. Mandarin, orange, and various hybrid varieties were analyzed to determine their carotenoid profiles. The resulting peak areas were analyzed using principal component analysis (PCA), canonical discriminate analysis (CDA), and Mahalanobis distances. These were used to develop models for classifying the juices into appropriate groups. Thirty-two samples were analyzed to determine classification techniques. Mandarin and orange juices were quite distinct, with the hybrids scattered throughout the mandarin and orange clusters using PCA. CDA resulted in three distinct groups with only a few of the hybrids in the orange grouping.     

Separation of flavanone-7-O-glycoside diastereomers and analysis in citrus juices by multidimensional liquid chromatography coupled with mass spectrometry

The major flavanone-7-O-glycoside constituents in citrus fruit juices (naringin, hesperidin, neohesperidin, narirutin, and eriocitrin) were separated as diastereomers by multidimensional liquid chromatography. The method consisted of coupling two HPLC columns: a reversed-phase (RP(18)) column was used for the separation of flavanone glycosides, which were, then, individually switched into a carboxymethylated beta-cyclodextrin (beta-CD)-based column and resolved as the corresponding stereoisomers. The method was used for the full analysis of flavanone glycosides in fresh hand-squeezed and commercial fruit juices by combining the quantitative estimation with the diastereomeric analysis. Quantitative data were in general consistent with previously reported data in this field. CC-LC isomer analysis was carried out by coupling the beta-CD column with a mass spectrometer operated with negative ion electrospray ionization (ESI-MS). The results showed that hesperidin was present in orange juices almost exclusively as the 2S isomer, whereas narirutin had mainly the 2R configuration. In grapefruit juices (2S)-naringin prevailed with the respect to the 2R isomer, whereas the opposite was true for narirutin. Lemon juices contained eriocitrin stereoisomers in equal amount (50% each), but hesperidin was almost exclusively found as the 2S isomer. Significant differences of the diastereomeric ratios were observed between freshly squeezed juices and juices from commercial sources.     

Determination of seven artificial sweeteners in diet food preparations by reverse-phase liquid chromatography with absorbance detection

The artificial sweeteners aspartame, saccharin, cyclamate, alitame, acesulfam-K, sucralose, and dulcin are determined in diet soft drinks and tabletop sweetener preparations. Samples are diluted, filtered, and analyzed directly by liquid chromatography on a C-18 reverse-phase column with a mobile phase gradient ranging from 3% acetonitrile in 0.02M KH2PO4 (pH 5) to 20% acetonitrile in 0.02M KH2PO4 (pH 3.5). Diet puddings and dessert toppings are extracted with ethanol, filtered, and diluted with mobile phase for analysis. The sweeteners, except sucralose and cyclamate, were detected by UV absorbance at either 200 or 210 nm. Sucralose was determined at 200 nm or by refractive index. Cyclamate was determined after post-column ion-pair extraction. The sweeteners stevioside and talin were not detected. Additives such as caffeine, sorbic acid, and benzoic acid did not interfere.     

Betaines in fruits of Citrus genus plants

Numerous compounds, many of them osmolytes, were quantified in natural juices and in frozen concentrate juices from fruits of plants of the Citrus genus. L-proline, N-methyl-L-proline (hygric acid), N,N-dimethyl-L-proline (stachydrine), 4-hydroxy-L-prolinebetaine (betonicine), 4-hydroxy-L-proline, γ-aminobutyric acid (Gaba), 3-carboxypropyltrimethylammonium (GabaBet), N-methylnicotinic acid (trigonelline), and choline in the fruit juices of yellow orange, blood orange, lemon, mandarin, bitter orange (Citrus aurantium), chinotto (Citrus myrtifolia), and grapefruit were analyzed by sensitive HPLC-ESI-tandem mass spectrometry procedure. It was found that the most represented osmolytes in the juices, that is, L-proline, stachydrine, and betonicine, can be quantified with minimal sample preparation and short analysis time (about 1 min) also by flow injection analysis (FIA) ESI-MS/MS with the same results as obtained by HPLC ESI-MS/MS. In all of the juices, discrete amounts of choline and trigonelline were present. Conversely, GabaBet was always below detection limits. Notably, N-methyl-L-proline and 4-hydroxy-L-prolinebetaine, which were discovered for the first time in the juice of bergamot (Citrus bergamia Risso et Poit), are also present in all of the citrus juices examined.     

Reverse-phase liquid chromatographic determination of benzoic and sorbic acids in foods

An isocratic liquid chromatographic (LC) technique is described for the determination of benzoic acid and sorbic acid in foods such as beverages, fruits, seafood, vegetables, sauces, and dairy, bakery, and confectionery products. A C18 column is used with methanol-phosphate buffer (5 + 95) as mobile phase and 4-hydroxyacetanilide or 3,5-dinitrobenzoic acid as internal standard. Sample preparation is simple, rapid, and produces a sample extract that has a minimum effect on the column performance and life. Specificity of the method was checked against common food additives such as L-ascorbic acid, caffeine, artificial sweeteners (saccharin, cyclamate, aspartame), antioxidants (BHT, BHA) and artificial colors. Also described are 2 procedures for confirmation of the preservatives, using either redox reaction of sorbic acid with potassium permanganate or gas chromatography/mass spectrometry. Mean recoveries of 90-105% were obtained with a precision of 1-6% and a detection limit of 20 mg/kg for the 2 preservatives.     

Effect of processing techniques at industrial scale on orange juice antioxidant and beneficial health compounds

Phenolic compounds, vitamin C (L-ascorbic acid and L-dehydroascorbic acid), and antioxidant capacity were evaluated in orange juices manufactured by different techniques. Five processes at industrial scale (squeezing, mild pasteurization, standard pasteurization, concentration, and freezing) used in commercial orange juice manufacturing were studied. In addition, domestic squeezing (a hand processing technique) was compared with commercial squeezing (an industrial FMC single-strength extraction) to evaluate their influences on health components of orange juice. Whole orange juice was divided into soluble and cloud fractions after centrifugation. Total and individual phenolics were analyzed in both fractions by HPLC. Commercial squeezing extracted 22% more phenolics than hand squeezing. The freezing process caused a dramatic decrease in phenolics, whereas the concentration process caused a mild precipitation of these compounds to the juice cloud. In pulp, pasteurization led to degradation of several phenolic compounds, that is, caffeic acid derivatives, vicenin 2 (apigenin 6,8-di-C-glucoside), and narirutin (5,7,4'-trihydroxyflavanone-7-rutinoside) with losses of 34.5, 30.7, and 28%, respectively. Regarding vitamin C, orange juice produced by commercial squeezing contained 25% more of this compound than domestic squeezing. Mild and standard pasteurization slightly increased the total vitamin C content as the contribution from the orange solids parts, whereas concentration and freezing did not show significant changes. The content of L-ascorbic acid provided 77-96% of the total antioxidant capacity of orange juice. Mild pasteurization, standard pasteurization, concentration, and freezing did not affect the total antioxidant capacity of juice, but they did, however, in pulp, where it was reduced by 47%.     

锦橙汁辐照和巴氏灭菌处理后相关品质的分析

为探讨辐照和巴氏杀菌对橙汁品质的影响,对锦橙汁分别进行1.4、2.8、5.6kGy3种不同剂量辐照及巴氏灭菌处理,使用固相微萃取-气质联用技术对挥发性成分进行分析,测定橙汁色度、pH值和Vc含量,并对橙汁香气进行感官评定。在鲜橙汁、巴氏灭菌汁和3种辐照样品中分别检测到了54、47、57、55和53种成分,2.8kGy剂量辐照处理后橙汁挥发性成分总峰面积最高,β-月桂烯、柠檬烯和γ-松油烯等橙汁特征香气物质经辐照后保持率高于巴氏灭菌。各种处理后Vc含量均有所下降,橙汁橙香均减弱。1.4kGy辐照后的橙汁感官评价结果最好,因此可以对橙汁进行低剂量辐照灭菌。     

Detection of orange juice adulteration with beet medium invert sugar using anion-exchange liquid chromatography with pulsed amperometric detection

Carbohydrate analysis of 5 beet medium invert sugar (BMIS) samples and 10 pure orange juices was carried out using anion-exchange chromatography with a pulsed amperometric detector. This analysis revealed the presence of several oligosaccharides in BMIS that were in either low concentration or nonexistent in the orange juice samples. These oligosaccharides may be naturally present in sugar beets or synthesized during the acid and/or enzyme catalyzed hydrolysis of sucrose during the production of BMIS. BMIS was intentionally added to pure orange juice at levels of 5, 10, 15, and 20%. Subsequent liquid chromatographic (LC) analysis of these intentionally adulterated samples revealed that detection of 5% BMIS in orange juice was possible.     

Red grape juice inhibits iron availability: application of an in vitro digestion/caco-2 cell model

Adequate bioavailable Fe intake is essential for optimal growth and intellectual development of infants and children. Fruit juices are nutritious and popular drinks for infants and children and are known to contain Fe uptake inhibitors (e.g., polyphenolic compounds) and a dominant promoter, ascorbic acid. Ascorbic acid is naturally present in fruit juices and is added during processing to almost all juices found in supermarkets. With these facts taken into account, an in vitro digestion/Caco-2 cell culture model was developed to compare the effects of apple, pear, white grape, red grape, prune, grapefruir, and orange juices on iron bioavailability. In two series of experiments, juices from a local supermarket were combined with FeCl(3) or commercial infant cereal fortified with elemental iron and subject to simulated gastric and intestinal digestion. Caco-2 cell ferritin formation in response to exposure to the digests served as the measure of Fe uptake. The pear, apple, grapefruit, orange, and white grape juice significantly increased Fe bioavailability from FeCl(3). For the infant cereal studies, the apple, orange, pear, and white grape juices increased the Fe bioavailability of the infant cereal. In contrast, the red grape juice and prune juice had profound inhibitory effects on iron bioavailability. These inhibitory effects were likely due to high levels of polyphenolic compounds that bind and thereby prevent absorption of soluble Fe. These inhibitory compounds appeared to counteract the promotional effects of ascorbic acid as they were in considerable molar excess relative to ascorbic acid and Fe in the digest. From a nutritional standpoint, the results suggest that individuals in need of optimal Fe absorption should avoid red grape and prune juice or at least vary the types of juices consumed. Alternatively, individuals seeking to limit Fe uptake (e.g., hemochromatitics and astronauts) may be able to utilize red grape or prune juice as effective inhibitors of Fe uptake. Consumers should be aware that the compounds that inhibit Fe availability are also linked to anticancer benefits; thus, a dietary balance of the above juices may be optimal.