Validation of the alkalimetry method for the quantitative determination of free organic acids in raspberry leaves




raspberry; leaves; free organic acids; alkalimetry; validation


 Organic acids are a large group of biologically active compounds that perform important functions in the plant organism. Moreover, all plants, regardless of the species and family, contain organic acids to a small or large extent as organic acids belong to intermediate metabolites arising from the oxidation of proteins and amino acids, fats and carbohydrates.
Aim. To validate the method of alkalimetry proposed with potentiometric detection of the end-point for the quantitative determination of free organic acids in raspberry leaves.
Results and discussion. The method proposed was validated according to the International Conference on Harmonization (ICH) guidelines. The linearity was in the concentration range of 40 – 200 % (r2 = 0.9991). The percentage of recovery was found to be in the range of 98.77 – 102.48 %. The repeatability and intermediate precision were 1.58 % and 1.74 %, respectively. The method is accurate and reliable, with the relative standard deviation of less than 2 %.
Experimental part. Leaves of raspberry were collected in the Kharkiv region during the period of full ripening. A Hanna 2550 pH-meter with a HI 1131P potentiometric electrode was used for alkalimetric titration of free organic acids. The titration was carried out using a microburette with Class A accuracy.
Conclusions. The alkalimetry method for the quantitative determination of free organic acids in raspberry leaves has been proposed and validated according to the following parameters: specifcity, linearity, accuracy, repeatability, intermediate precision, robustness. It has been confrmed that the method is simple, reliable, accurate and cost-effective.
Key words: raspberry; leaves; free organic acids; alkalimetry; validation


Padmanabhan, P.; Correa-Betanzo, J.; Paliyath, G. Berries and Related Fruits. In Encyclopedia of Food and Health; Caballero, B.; Finglas, P. M.; Toldrá, F., Eds. Academic Press: Oxford, 2016; pp 364 – 371.

Sariburun, E.; Şahin, S.; Demir, C.; Türkben, C.; Uylaşer, V. Phenolic Content and Antioxidant Activity of Raspberry and Blackberry Cultivars. J. Food Sci. 2010, 75 (4), C328 – C335.

Velichko, V. V.; Makarova, D. L. Comparative pharmacognostic analysis of leaves and fruit of raspberry ordinary. Medicine and Education in Siberia 2015, 4.

Дергачева, Ж. М.; Гурина, Н. С.; Мушкина, О. В. Фитохимический анализ листьев малины обыкновенной (Rubi Idaeus Folia). Рецепт 2015, 6, 64 – 74.

Bobinaitė, R.; Viškelis, P.; Venskutonis, P. R., Chapter 29 – Chemical Composition of Raspberry (Rubus spp.) Cultivars. In Nutritional Composition of Fruit Cultivars; Simmonds, M. S. J.; Preedy, V. R., Eds.; Academic Press: San Diego, 2016; pp 713-731.

Ferlemi, A.-V.; Lamari, F. N. Berry Leaves: An Alternative Source of Bioactive Natural Products of Nutritional and Medicinal Value. Antioxidants 2016, 5 (2), 17.

Сергунова, Е. В.; Марахова, А. И.; Аврач, А. С. Методы количественного определения органических кислот в лекарственном растительном сырье и водных извлечениях. Фармация 2013, 4, 8 – 11.

Gu, Y.; Li, J.; Song, W.; Zhang, X. Determination of C1-C6 organic acids in the products from syngas to olefins by ion chromatography. Chinese journal of chromatography 2014, 32 (2), 204 – 209.

Mortera, P.; Zuljan, F. A.; Magni, C.; Bortolato, S. A.; Alarcón, S. H. Multivariate analysis of organic acids in fermented food from reversed-phase high-performance liquid chromatography data. Talanta 2018, 178, 15 – 23.

Nogueira, T.; Lago, C. L. d. Determination of Ca, K, Mg, Na, sulfate, phosphate, formate, acetate, propionate, and glycerol in biodiesel by capillary electrophoresis with capacitively coupled contactless conductivity detection. Microchem. J. 2011, 99 (2), 267 – 272.

Zhilkina, V. Yu.; Marakhova, A. I.; Stanishevskiy, Ya. M. Qualitative and quantitative analysis of organic acids in mixture of multivitamin raw material. Drug development & registration 2016, 1, 156 – 159.

Umarov, U. A.; Maslov, O. Y.; Kolisnyk, S. V.; Fathullaeva, М. Development and Validation of The Conductometric Titration Method of Quantitative Determination of Free Organic Acids in The Anise Fruits. European Journal of Molecular & Clinical Medicine 2020, 7 (3), 3874 – 3883.

Sochorova, L.; Torokova, L.; Baron, M.; Sochor, J. Electrochemical and others techniques for the determination of malic acid and tartaric acid in must and wine. Int. J. Electrochem. Sci. 2018, 13 (9), 9145 – 9165.

Strömberg, N.; Sahlin, E. Determination of the short–chain fatty acid pattern in biodiesel using high throughput syringe solvent extraction and ion exclusion chromatography. Fuel 2012, 97, 531 – 535.

The International Council for Harmonisation. Quality Guidelines. (accessed Jan 10, 2021), Validation of analytical procedures: text and methodology Q2(R1).

Державна фармакопея України: в 3 т., 2-е вид.; Державне підприємство «Український науковий фармакопейний центр якості лікарських засобів»: Харків, 2015; Т. 1.