Scientific Papers Series B Horticulture

found to prevent the formation of N-nitrosocompounds, the cancer causing substancesfrom nitrates and nitrites found in preservedmeats and some drinking water (Du et al.,2009). Vitamin C has many biological activities(reducing carcinogenesis and cardiovasculardiseases, stimulating the immune system) in thehuman body (Simon, 1992).There are several methods to determine vitaminC content; however some of them needsubjective evaluation and some are notpractical (Agar, 1995). Several postharvestfactors influence the vitamin C and organicacid contents of apple. Despite manyinvestigations in the area of nutrition andpostharvest changes of apples, knowledgeabout the determination of vitamin C andorganic acids by using HPLC is inadequate. Ithas been indicated that ascorbic acid content offruit should be measured by HPLC becauseascorbic acid produces an oxidative-reductionreaction (Asami et al., 2003). The aim of thisresearch was to measure the vitamin C(ascorbic acid) and organic acid contents of 20old Romanian apples varieties by using HighPerformance Liquid Chromatography (HPLC)during cold storage period.MATERIALS AND METHODSForty old apples cultivars (Prescurate,Gurguiate, Viesti, Rosii Stetin, Zori, Carla,Mohorat, Gustav Durabil, WachsmanSammeling, Wachsman Amelie, Ancuta,Ardelean, Trotuse,etc.) grown in the researchand application center of HorticultureDepartment in Bucharest located in south ofRomania region were harvested at the last stageof commercial ripeness (red colour with asurface area of > 75-80%). Harvested fruits inthe early morning were transported to the postharvest laboratory within 30 min. Applescultivars were sorted to eliminate fruits withdefects including overripe or too small fruit.Fruits were selected randomly and placed inperforated (8 perforations with 10 mm diameteron each box) plastic boxes (capacity: 750 g) foreach replicates. Four replicates were used pertreatment. Packaged fruits were stored at 0°Ctemperature and 90-95% relative humidity for10 days. Apples fruits cultivars were analyzedat 5 days intervals (0, 5, and 10 days of storage)during cold storage.Vitamin C (Ascorbic acid) and Organic AcidAnalysisThe HPLC analysis was carried out todetermine the vitamin C and organic acids on aShimadzu class LC VP HPLC system withclass LC-VP software, a pump (LC-6AD), anda UV-VIS detector (SPD-10AV VP). Thecolumns used were YMC Pack-ODS (250 mmx 4.6 mm I.D., 5 μm) for organic acids andSGE (250 mm x 4.6 mm I.D., 5 μm) forvitamin C. The mobile phases were wateradjusted to pH 2.2 with trifluoroacetic acid(organic acids) and to pH 3 with phosphoricacid (vitamin C). Separation was carried out byisocratic elution with a flow rate of 0.4 ml min-1 and column temperature was ambient. TheUV detector was set at 210 nm and 254 nm,respectively. Quantitation was based on thepeak area measurement.Sample (10 g) was extracted in 10 ml wateradjusted to pH 1.5 with trifluoroacetic acid fororganic acids and with 10 ml phosphoric acidwater(2%, v/v) for vitamin C. The extractswere filtered through filter paper. Then, 1.5 mlbuffer (0.01 M KH2PO4, pH 8.0) was added to1.5 ml sample extract. From this, 1.5 ml(organic acids) and 1 ml (vitamin C) of thesemixtures were loaded on to C18 cartridges.After loading, 3 ml water adjusted to pH 1.5with trifluoroacetic acid for organic acids and 2ml phosphoric acid-water(2%, v/v) for vitamin C were passed throughthe cartridges. For HPLC, 20 μl of the eluentswere injected.RESULTS AND DISCUSSIONSGenerally, fruits and vegetables show a gradualdecrease in vitamin C content as the storagetemperature or duration increases (Adisa,1986). In this research, change in vitamin C ofapples during cold storage is shown in Figure1. Vitamin C levels decreased for most ofcultivars from the beginning to the end of thestorage, but this reduction was not statisticallysignificant (P