CC BY
10DOI: 10.6060/ivkkt.20236605.6784
УДК: 543.645
КОНТРОЛЬ СЕЛЕКТИВНОСТИ РАЗДЕЛЕНИЯ 3-ГЛЮКОЗИДОВ И 3,5-ДИГЛЮКОЗИДОВ АНТОЦИАНИДИНОВ ВИНОГРАДА: ОПРЕДЕЛЕНИЕ АНТОЦИАНОВ ПЛОДОВ ВИНОГРАДОВ, ВЫРАЩЕННЫХ В БЕЛГОРОДСКОЙ ОБЛАСТИ
Я.Ю. Саласина, В.И. Дейнека, И.П. Блинова, Е.Ю. Олейниц, Л.А. Дейнека, С.Л. Макаревич
Ярослава Юрьевна Саласина (ORCID 0000-0002-4118-9941)*, Виктор Иванович Дейнека (ORCID 00000002-3971-2246), Ирина Петровна Блинова (ORCID 0000-00002-4525-4536), Елена Юрьевна Олейниц (ORCID 0000-0003-2065-6296), Людмила Александровна Дейнека (ORCID 0000-0002-4101-2468)
Кафедра общей химии, Белгородский государственный национальный исследовательский университет, ул. Победы, 85, Белгород, Российская Федерация, 308015
E-mail: salasina@bsu.edu.ru*, deineka@bsu.edu.ru, blinova@bsu.edu.ru, oleinits_e@bsu.edu.ru, deyneka@bsu.edu.ru
Сергей Леонидович Макаревич (ORCID 0000-0002-4671-1903)
Белгородская межобластная ветеринарная лаборатория, ул. Студенческая, 32, Белгород, Российская
Федерация, 308023
E-mail: sergmazay@yandex.ru
Впервые предложен новый вариант разделения 3-глюкозидов и 3,5-диглюкозидов пяти основных для плодов виноградов антоцианидинов - дельфинидина, цианидина, пету-нидина, пеонидина и мальвидина в условиях обращенно-фазовой ВЭЖХ. Показано, что замена традиционно используемого ацетонитрила на экологически более приемлемый этанол при подкислении не муравьиной, а ортофосфорной кислотой позволяет существенно изменить селективность разделения двух типов глюкозидов при их совместном присутствии. Предложенный вариант разделения позволяет дифференцировать винограды вида Vitis vinifera, в кожуре плодов которых синтезируются только 3-глюкозиды перечисленных выше антоцианидинов, и винограды иных видов или гибридных сортов винограда. Для полного обзора антоцианового состава необходимо использование градиентного режима, поскольку ацилирование антоцианов уксусной и пара-кумаровой кислотами существенно изменяет липофильность антоцианов. Тип антоцианов анализировали по ранее предложенной системе, учитывающей активность трех типов ферментов: 1) 5-О-гликозил-трансферазы, участвующей в образовании 3,5-диглюкозидов, 2) 3',5'-гидроксилазы ответственной за гидроксилирование кольца B; 3) антоциан O-метилтрансферазы для превращения производных цианидина в производные пеонидина, как и производных дельфинидина в гликозиды петунидина и мальвидина. Метод был использован для определения антоцианов 43 сортов виноградов, выращенных в Белгороде в фермерских и частных хозяйствах. Среди исследованных виноградов обнаружены сорта с плодами винограда с накоплением только 3-глюкозидов, как и одновременно 3-глюкозидов и 3,5-диглюкозидов, с дель-финидиновым и цианидиновым типами антоцианов, и с различной степенью метилирования. Рассчитанные параметры всех сортов виноградов представлены в двух таблицах и приведено краткое обсуждение некоторых сортов.
Ключевые слова: плоды виноградов, антоцианы, ОФ ВЭЖХ, подвижные фазы на основе этанола, три критерия классификации виноградов
Для цитирования:
Саласина Я.Ю., Дейнека В.И., Блинова И.П., Олейниц Е.Ю., Дейнека Л.А., Макаревич С.Л. Контроль селективности разделения 3-глюкозидов и 3,5-диглюкозидов антоцианидинов винограда: Определение антоцианов плодов виноградов, выращенных в Белгородской области. Изв. вузов. Химия и хим. технология. 2023. Т. 66. Вып. 5. С. 72-79. DOI: 10.6060/ivkkt.20236605.6784.
For citation:
Salasina Ya.Yu., Deineka V.I., Blinova I.P., Oleinits E.Yu., Deineka L.A., Makarevich S.L. Control of the selectivity of the separation of grape anthocyanidin 3-glucosides and 3,5-diglucosides: determination of anthocyanins in grape fruit grown in the Belgorod region. ChemChemTech [Izv. Vyssh. Uchebn. Zaved. Khim. Khim. Tekhnol.]. 2023. V. 66. N 5. P. 72-79. DOI: 10.6060/ivkkt.20236605.6784.
CONTROL OF THE SELECTIVITY OF THE SEPARATION OF GRAPE ANTHOCYANIDIN
3-GLUCOSIDES AND 3,5-DIGLUCOSIDES: DETERMINATION OF ANTHOCYANINS IN GRAPE FRUIT GROWN IN THE BELGOROD REGION
Ya.Yu. Salasina, V.I. Deineka, I.P. Blinova, E.Yu. Oleinits, L.A. Deineka, S.L. Makarevich
Yaroslava Yu. Salasina, (ORCID 0000-0002-4118-9941)*, Victor I. Deineka (ORCID 0000-0002-3971-2246), Irina P. Blinova (ORCID 0000-00002-4525-4536), Elena Yu. Oleinits (ORCID 0000-0003-2065-6296), Lyudmila A. Deineka (ORCID 0000-0002-4101-2468)
Department of General Chemistry, Belgorod State National Research University, Pobedy st., 85, Belgorod, 308015, Russia
E-mail: salasina@bsu.edu.ru*, deineka@bsu.edu.ru, blinova@bsu.edu.ru, oleinits_e@bsu.edu.ru, deyneka@bsu.edu.ru
Sergey L. Makarevich (ORCID 0000-0002-4671-1903)
Belgorod Interregional Veterinary Laboratory, Studencheskaya st., 32, Belgorod, 308023, Russia E-mail: salasina@bsu.edu.ru
For the first time, a new version of the separation of 3-glucosides and 3,5-diglucosides of the five main anthocyanidins for grape fruits - delphinidin, cyanidin, petunidin, peonidin and mal-vidin under reverse-phase HPLC conditions was proposed. It has been shown that the replacement of the traditionally used acetonitrile with environmentally more acceptable ethanol upon acidification with ortho-phosphoric rather than formic acid makes it possible to significantly change the selectivity of the separation of two types of glucosides at their joint presence. The proposed separation option allows differentiating between Vitis vinifera grapes, in the fruit peel of which only 3-glucosides of the above anthocyanidins are synthesized, and grapes of other species or hybrid grape varieties. For a complete review of the anthocyanin composition, it is necessary to use a gradient mode, since the acylation of anthocyanins with acetic andp-coumaric acids significantly changes the lipophilicity of anthocyanins. The type of anthocyanins was analyzed according to the previously proposed system, which takes into account the activity of three types of enzymes: 1) 5-O-glucosyltransferase that leads to 3,5-diglucosides synthesis; 2) 3',5'-hydroxylase responsible for ring B hydroxylation; 3) anthocyanin O-methyltransferase for converting cyanidin into peonidin derivatives as well as delphinidine into petunidin and malvidin glycosides. The method was used to determine anthocyanins in 43 grape varieties grown in Belgorod on farms and private farms. Among the studied grapes, varieties were found with grape fruits with the accumulation of only 3-glucosides, as well as 3-glucosides and 3,5-diglucosides simultaneously, with delphinidin and cyanidin types of anthocyanins, and with different degrees of methylation. The calculated parameters for all grape varieties are presented in two tables and a brief discussion of some varieties is given.
Key words: grape fruits, anthocyanins, RP HPLC, ethanol-based mobile phases, three criteria for grape classification
INTRODUCTION
Grape is one of the most popular fruit crops cultivated worldwide; according to literature data world grape production reached 25.62 million metric tons in 2021/2022 season [1]. Vitaceae (the grape fam-
ily) consists of about 14 genera and 900 species primarily distributed in tropical regions [2]. Grapes are explored for mainly winemaking - about 80%, 13% is sold as table while the remaining grapes are used to produce raisins, juice and the other products [3]. Since ancient times in Europe (in Mediterranean region)
namely Vitis vinifera species are cultivated. But in the second half of the 19th century, the European vineyards were attacked with accidentally introduced from North America phylloxera and the problem to withstand to the attacks was resolved by employing American grapevines as rootstocks for V. vinifera [4].
The dark coloration with blue or red shades of grape fruits is obliged to the biosynthesis (ordinary in the skin) of anthocyanins that may be utilized to prepare natural food colorants (E163). Meanwhile there is a distinct difference between anthocyanin types as V. vinifera and non-V. vinifera species [5]. For the former the synthesis of 3-O-glucosides (3Glu) of five antho-cyanidines: delphinidin (Dp), cyanidin (Cy), petunidin (Pt), peonidin (Pn) and malvidin (Mv) is characteristic, Fig. 1. For the latter addition to 3Glu-types of 3,5-O-diglucosides (3,5diGlu) of the same anthocyanidins was found [6-10].
Fig. 1. Structures of five main grape fruit anliiocyanidins and positions of glucosylation Рис. 1. Структуры пяти основных антоцианидинов плодов винограда и положения глюкозилирования
Thus, the detection of 3,5-O-diglucosides can be used for differentiation of V. vinifera and V. rotun-difolia, V. labrusca, V. coignetiae, V. rupestris, V. amurensis as well as interspecific hybrid grapes. The ten 3-glucosides and 3,5-diglucosides are supplemented with set of anthocyanins acylated with acetic and para-coumaric acids (sometimes also acylated with caffeic one) that have highly enhanced lipophilic-ity. Thus, a problem of anthocyanins separation for a subsequent quantification is not a simple task. Re-versed-phase HPLC (RP HPLC) in gradient elution mode is a method ordinary used for grape anthocyanins analysis [11-14]. Though acylated anthocyanins are interesting for preparation of differently colored encapsulated anthocyanins [15] the problem of Vitis species differentiation may be established by the determination only of non-acylated compounds. The five 3-glucosides as well as the five 3,5-diglucosides are easily separated by acidified mobile-phases with acetonitrile or methanol as organic modifiers eluting in the order of retention times growth:
Dp3Glu - Cy3Glu - Pt3Glu - Pn3Glu - Mv3Glu, Dp3,5diGlu - Cy3,5diGlu - Pt3,5diGlu - Pn3,5diGlu -Mv3,5diGlu.
In the case of joint presence of 3-glucosides and 3,5-diglucosides some separation problems arise in isocratic as well as in gradient elution with mobile phases containing acetonitrile and formic acids as mobile phase modifiers are used [13, 14, 16]. Meanwhile acetonitrile is a toxic solvent for men and environment, thus giving rise to find more ecologically suitable conditions for RP HPLC.
Thus, the objective of this study was to find more ecologically friendly mobile phase modifiers for separation of the ten non-acylated anthocyanins for differentiation of V. vinifera and hybrid varieties of the grapes grown in Belgorod.
EXPERIMENTAL TECHNIQUES
Samples of grape were obtained from peasant farms of Belgorod and the region. Grape fruits accumulate anthocyanins mainly in fruit skin like black currant [17] and some other fruits. Thus, anthocyanins were extracted by maceration of grape fruit skins in 0.1 M HCl water solution for 20 h. The samples were partially cleaned up by solid phase extraction on DIAPACK C18 cartridges as follows: cartridges were activated by passing 3 ml of acetone, conditioned by passing 9 ml of 0.1 M HCl water solution. After sorption of anthocyanins from the extracts, anthocyanins were de-sorbed from cartridges by passing 3 ml of solution 30 vol. % of CH3CN, 30 vol.% of HCOOH and 40 vol. % of water. The obtained solution was diluted with water 1:2 by volume for subsequent injection into the chromatograph.
For gradient elution two mobile phases were used - phase A, 8 vol.% of ethanol and 1 vol.% of H3PO4 in water; phase B, 20 vol.% of ethanol and 1 vol.% of H3PO4 in water. Gradient mode: 0 min 0% B; 10 min 10% B, 30 min 100 % B, 31 min 0% B; 40 min 0% B.
Separations were performed on an Agilent 1200 Infinity chromatograph with a diode array detector (DAD). A chromatographic column 150^4.6 mm SymmetryTMC18, 3.5 ^m was used for separation of anthocyanins.
Utilization of DAD permits to differentiate 3-glucosides by characteristic electronic absorption spectra that are almost the same for Cy3Glu and Pn3Glu (solutes are mentioned according to the order of elution) with absorption maxima near 515 nm, Fig. 2. Spectra of Dp3Glu, Pt3Glu and Mv3Glu have absorption maxima shifted bathochromically to 524-526 nm region, while the methylation of OH-groups in B-ring leads to small sequential 1.0 nm shift of the maxima. 3,5-diglucosides are also easily differentiated by absence of local maxima at 420-450 nm in electronic ab-
sorption spectra fixtures. Moreover, peaks of 3,5-diglucosides have extra broadening compared to that of 3-glucosides [18].
Fig. 2. Electronic absorption spectra of anthocyanins in the mobile phase. Compounds: 1 - Dp3Glu, 2 - Cy3Glu, 3 - Pt3Glu, 4 - Pn3Glu, 5 - Mv3Glu, 6 - Mv3,5diGlu. Рис. 2. Электронные спектры поглощения в подвижной фазе. Антоцианы: 1 - Dp3Glu, 2 - Cy3Glu, 3 - Pt3Glu, 4 - Pn3Glu, 5 - Mv3Glu, 6 - Mv3,5diGlu
The mobile phases were prepared using ace-tonitrile (HPLC Gradient grade, Fisher Chemical, Germany), ethanol (95%, Hippocrates LLC, RF) ortho-phosphoric acid (85%, RUSHIM, RF), distilled water.
RESULTS AND DISCUSSION
The most popular mobile phases for anthocya-nins separations are based on acetonitrile as an organic modifier and formic acid for pH correction to transfer anthocyanins into flavylium form. The separation map for 3-glucosides and 3,5-diglucosides of five common for grape fruits anthocyanidins for mobile-phase system "10 vol. % of formic acid-(6-10 vol. %) acetoni-trile - water" and Symmetry TMC18, 3.5 цш column is proposed in Fig. 3.
It is evident that for the system there is a problem for separation Pt3Glu and Mv3,5diGlu. The problem is complicated because of the latter compound' peak broadening being the intrinsic property of peaks of solutes with 3,5-diglycosylation. This problem has a great value since the presence of namely malvidin-3,5-diglucoside is commonly the main criteria for detection of hybrid or non-vinifera grape cultivars. The problem has some ways of decisions [16, 18, 19], but lowering the temperature of separation and exchange of formic acid with phosphoric acid buffer [18] permits not to separate all the ten compounds. Thus another approach must be found to solve the problem.
Ethanol seems to be the most «green» solvent for the substitution of ecologically unfavorable ace-tonitrile [20]. But the exchange of acetonitrile with eth-anol demands also to exchange formic acid for ortho-
phosphoric one to prevent esterification reaction. To maintain high mobile phase acidity 1 vol. % of the acid must be added. The separation map for mobile phase system "10-20 vol. % of ethanol - 1 vol.% of ortho-phosphoric acid in water" is presented in Fig. 4.
Fig. 3. Separation map of 3-glucosides and 3,5-diglucosides in mobile phase system "6 - 9.6 vol.% of CH3CN, 10 vol.% of HCOOH in water". Compounds: 1 - Dp3,5diGlu; 2 - Cy3,5diGlu, 3 - Pt3,5diGlu, 4 - Pn3,5diGlu, 5 - Mv3,5diGlu, 6 - Dp3Glu; 7 - Cy3Glu, 8 - Pt3Glu, 9 - Pn3Glu, 10 - Mv3Glu Рис. 3. Карта разделения 3-глюкозидов и 3,5-диглюкозидов в
системе подвижных фаз "6 - 9,6 об.% CH3CN, 10 об.% HCOOH в воде". Соединения: 1 - Dp3,5diGlu; 2 - Cy3,5diGlu, 3 - Pt3,5diGlu, 4 - Pn3,5diGlu, 5 - Mv3,5diGlu, 6 - Dp3Glu; 7 - Cy3Glu, 8 - Pt3Glu, 9 - Pn3Glu, 10 - Mv3Glu
Fig. 4. Separation map of 3-glucosides and 3,5-diglucosides in mobile phase system "12 - 16 vol.% of CH3CH2OH, 1 vol.% of
H3PO4 in water". For compounds numbering see Fig. 2 Рис. 4. Карта разделения 3-глюкозидов и 3,5-диглюкозидов в системе подвижных фаз "12 - 16 об.% CH3CH2OH, 1 об.% H3PO4 в воде". Нумерация соединений как на Рис. 2
In this case, selectivity of 3Glu and 3,5diGlu compounds separation changes markedly. At low ethanol concentration (10 vol. %) in a mobile phase Mv3,5diGlu elutes after Pt3Glu and Pn3,5diGlu. At high mobile phase elution strength solutes are grouped according to number of OH-groups in B-ring with tendency for reversal of the elution order in pairs Mv3Glu + +Pn3Glu and Pt3Glu + Cy3Glu. Nevertheless, the elu-tion mode in mobile phases, based upon ethanol is highly favorable for grape type differentiation by HPLC method.
In this case, selectivity of 3Glu and 3,5diGlu compounds separation changes markedly. At low eth-anol concentration (10 vol. %) in a mobile phase Mv3,5diGlu elutes after Pt3Glu and Pn3,5diGlu. At high mobile phase elution strength solutes are grouped according to number of OH-groups in B-ring with tendency for reversal of the elution order in pairs Mv3Glu + +Pn3Glu and Pt3Glu + Cy3Glu. Nevertheless, the elu-tion mode in mobile phases based upon ethanol is highly favorable for grape type differentiation by HPLC method.
Thus, for investigation of V. vinifera grape an-thocyanins simple gradient modes with CH3CN, CH3OH or C2H5OH may be composed, but for non-V. vinifera and hybrid grape cultivars the mode of elution must be carefully selected, e.g. gradient mode proposed in the experimental part. The separation of grape fruits anthocyanins in this mode of elution is presented in Fig. 5.
The developed in this paper method was applied for analysis of anthocyanins composition in fruits of 43 grape cultivars grown and harvested in Belgorod region. For grape classification we used criteria developed in paper [21] for peak areas of non acylated anthocyanins.
The first criterion is devoted to estimate the inheritance (or absence) of 5-0-glucosyltransferase in anthocyanins biosynthesis from non-V. vinifera species by equation (1):
where X S3Glu (i) + X S3,5 diGlu (i) - is sum of areas
3 3
of peaks of delphinidin, petunidin and malvidin 3 -glucosides and their 3,5-diglucosides, ^ S3Glu( j) +
2
+ X S35diGlu (j) - is sum of areas of peaks of cyanidin
2
and peonidin 3-glucosides and their 3,5-diglucosides.
X SGu
a, = =-=-r • 100, % (1)
X S3,5diGlu (i) + X S3Glu (i) 5 5
where X S35AG&(i) - is sum of areas of peaks of all five
5
anthocyanidins, glucosylated at positions 3 and 5;
X S3GIU (i) - is sum of areas of peaks of all five an-
5
thocyanidins, glucosylated at position 3. For V. vinifera grapes a1 = 100.
mAU
0 ^—1—1—1—1—I—J—1—1—1—I—1—1—J—J—I—1—1—1—1—I—'—J—J—J—I—1—1—1—1—h
0 5 10 15 20 25
Fig. 5. Separation of hybrid grape anthocyanins in gradient mode with eluents, based upon ethanol and phosphoric acid in water. Extracts of fruits of grape cultivars: A - Mucuzani; B - Chernyj
sultan; C - Regent. For compounds numbering see Fig. 2 Рис. 5. Разделение антоцианов гибридных сортов в градиентном режиме с элюентами на основе этанола и фосфорной кислоты. Экстракты плодов винограда сортов: A - Мукузани, B - Черный султан, C - Регент. Нумерация соединений как на Рис. 2
The second criterion differentiates grapes on delphinidin or cyanidin types pointing out relative activity of flavonoid 3',5'-hydroxylase (F3'5'H):
(2)
According to our experience the grape of species with a(F3'5'H) > 50% have dark blue coloration while in the case of a(F3'5'H) > 50% coloration is dark or light red.
The third criterion will determine the relative degree of activity of anthocyanin O-methyltransferase (AOMT):
X S3Glu (i) + X S3,5diGlu (i)
a 2 -;-^-;-^-:—^-r • 100, %
X S3Glu (i) + X S3,5diGlu (i) + X S3Glu ( j) + X S3,5diGlu ( j) 3 2 2
3
X S (Pn ) i + X S (pt )j + 2 -X S (Mv) ,,
a, =
2-X S(Dp)y +X S(Cy) y + 2-X S(Pt) y +X S(Pn) y + 2-X S(Mv),
-100,%
(3)
where ^s(X)r - is sum of areas of peaks of 3-gluco-
2
side and 3,5-diglucoside of anthocyanidins indicated in brackets. Common European V. vinifera grapes belong to malvidin type due to high values of parameters a2 and аэ.
The results of grape anthocyanins determination are summered in Table 1 and Table 2. In the Tables parameters а1-а3 are calculated as well as the mole fraction of the main anthocyanin. The first table includes grapes without inheritance of glycosylation of OH-group in position 5 of the aglycons. The Table 2 contents grape cultivars with typical for hybrid culti-vars anthocyanin types.
Table 1
Characterization of anthocyanin biosynthesis in fruits of some grape cultivars without inheritance of 3,5-diglucosides biosynthesis Таблица 1. Характеристики биосинтеза антоцианов в плодах некоторых сортов виноградов без наследования биосинтеза 3,5-диглюкозидов
The main an-
No. Grape cultivar thocyanin mole fraction, % a1 a2 аэ
1 Chernaya magiya Mv3Glu (46.1) 100 93.2 61.4
2 Furor Mv3Glu (86.6) 100 91.5 96.6
3 Bagira Mv3Glu (80.0) 100 88.6 93.2
4 Charli (Antracit) Mv3Glu (43.9) 100 87.5 62.4
5 Kodryanka Mv3Glu (72.2) 100 85.7 88.8
6 Tornado Mv3Glu (70.7) 100 85.1 88.3
7 Atos Mv3Glu (59.1) 100 80.2 80.9
8 Lorano Mv3Glu (64.3) 100 80.2 86.0
9 Yupiter Mv3Glu (31.6) 100 75.6 53.0
10 Magiya Dp3Glu (38.6) 100 66.8 32.6
11 Yasya Mv3Glu (42.9) 100 58.7 80.1
12 Gurzufskij rozovyj Pn3Glu (28.5) 100 44.1 48.8
13 Kishmish Lu-chistyj Cy3Glu (36.9) 100 32.2 44.9
14 Polonez Cy3Glu (50.4) 100 30.2 31.1
15 Koronnyj Cy3Glu (57.1) 100 21.6 29.9
16 Chernyj zhemchug Pn3Glu (63.5) 100 20.2 80.9
17 Pamyati uchitelya Pn3Glu (72.4) 100 20.0 88.5
18 Preobrazhenskij Pn3Glu (75.6) 100 12.2 84.9
19 Podarok Vinokuru Pn3Glu (79.6) 100 7.4 85.3
20 Ametist Pn3Glu (91.1) 100 0.7 91.7
In spite of 3,5-diglucosides absence in antho-cyanin set of fruits presented in Table 1 most of the species are mostly of hybrid nature. Thus, grape cultivar No. 1 ("Chernaya magiya") entirely has not 3,5-
2 2 diglucosides but is relatively resistant to mildew, gray mold and oidium, as a hybrid type cultivar. This property is highly valuable for grape cultivation in our region. But there is a difference from conventional "malvidin" V. vinifera grapes due to reduced AOMT activity. In fruits of also hybrid "Bagira" grape cultivar mole fraction of Mv3Glu is appreciably higher and the grape has high frost resistance but loose stability towards phylloxera. Anthocyanins biosynthesis is mainly located in the fruit skin. As a rule, delphinidin (or malvidin) grape types have dark blue coloration while intensity of red coloration is significantly reduced for cyanidin (peonidin) types.
Table 2
Characterization of anthocyanin biosynthesis in fruits of some grape cultivars with inheritance of 3,5-digluco-
sides biosynthesis Таблица 2. Характеристики биосинтеза антоцианов в плодах некоторых сортов виноградов с наследованием биосинтеза 3,5-диглюкозидов
No. Grape cultivar name The main anthocyanin mole fraction, % a1 a2 a3
1 Spokusa Mv3Glu (49.7%) 99.0 69.9 81.0
2 Dubovskij rozovyj Cy3Glu (52.9%) 97.8 29.5 24.9
3 Fioletovyj rannij Mv3Glu (29.6%) 96.9 70.2 57.1
4 Severnyj plechistik Mv3Glu (29.6%) 96.7 69.7 57.9
5 Preobrazhenie Cy3Glu (76.3%) 96.4 14.4 21.2
6 Zarya Nesvetaya Pn3Glu (81.6%) 96.2 8.2 90.9
7 Asya Pn3Glu (81.6%) 95.2 38.5 88.3
8 Zejbel' Mv3Glu (28.2%) 88.0 91.2 53.5
9 Andryusha Pn3Glu (64.1%) 85.4 9.54 85.0
10 Nadezhda AZOS Mv3Glu (33.8%) 78.8 92.3 65.6
11 Markett Dp3Glu (28.9%) 76.9 92.8 53.1
12 Atos Pn3Glu (20.9%) 71.1 43.3 69.6
13 Regent Mv3Glu (22.7%) 67.3 86.1 63.5
14 Vas'kovskij 5 Mv3Glu (25.6%) 67.0 73.6 50.8
15 Livadijskij chernyj Mv3,5diGlu (27.3%) 59.9 94.5 63.4
16 Monarh Mv3,5diGlu (44.8%) 48.5 96.2 78.0
17 GIS 1-31 Mv3,5diGlu (35.1%) 48.0 78.6 78.5
18 Nadezhda rannyaya Mv3,5diGlu (40.8%) 47.4 84.2 90.1
19 Kaberne kortis Mv3,5diGlu (44.3%) 40.1 89.7 79.5
20 Taezhnyj (V. amurensis) Mv3,5diGlu (51.9%) 25.3 89.2 71.6
21 Neretinskij Mv3,5diGlu (70.2%) 25.1 97.6 90.9
22 Seedless Chernyj sultan Dp3,5diGlu (38.5%) 35.2 85.2 16.4
23 Mukuzani ("Oberlin Noir") Mv3,5diGlu (47.1%) 24.6 95 64.1
2
2
2
2
2
2
The first 7 grape species in the Table 2 are closed to V. vinifera types though they belong to different types according to criteria a2 and a3 having Mv3Glu, Cy3Glu or Pn3Glu as a major anthocyanin. The grape "Livadijskij chernyj" begins the list of cul-tivars with Mv3,5diGlu as a predominant anthocyanin. The set includes "Taezhnyj" cultivar which is an example of V. amurensis. The most interesting cultivar was "Mukuzani". According to non-official information available in Internet cites the cultivar was accidentally found in Georgia though the "Saperavi" cultivar is used to make wine under the "Mukuzani" brand. Meanwhile according to our investigation "Mukuzani" is evidently hybrid cultivar with the lowest value of criteria a1 between all cultivars studied in current paper.
ЛИТЕРАТУРА
1. Shahbandeh M. Global grape production 2112/12-2022/23. https://www.statista.com/statistics/237600/world-grape-pro-duction-in-2007-by-region/ (доступ 07.02.2023).
2. Sasidharan H., Varghese A.P. Taxonomy of selected species of Cissus (Vitaceae) from Thrissur district. South Indian J. Biolog. Sci. 2016. V. 2. P. 222-228. DOI: 10.22205/SIJBS/2016/V2/I1/100403.
3. Mazza G. Anthocyanins in Grapes and Grape Products. Crit. Rev. Food Sci. Nutrit. 1995. V. 35. P. 341-371. DOI: 10.1080/10408399509527704.
4. Ardenghi N.M.G., Galasso G., Banfi E., Zoccola A., Foggi B., Lastrucci L. A taxonomic survey of the genus Vitis L. (Vitaceae) in Italy, with special reference to Elba Island (Tuscan Archipelago). Phytotaxa. 2014. V. 166. P. 163-198. DOI: 10.11646/phytotaxa. 166.3.1.
5. He F., Mu L., Yan G-L., Liang N.-N., Pan Q.-H., Wang J. Reeves M.J., Duan C.-Q. Biosynthesis of Anthocyanins and Their Regulation in Colored Grapes. Molecules. 2010. V. 15. P. 9057-9091. DOI: 10.3390/molecules15129057.
6. He J.-J., Liu Y.-X., Pan Q.-H., Cui X.-Y., Duan C.-Q. Different Anthocyanin Profiles of the Skin and the Pulp of Yan73 (Muscat Hamburg x Alicante Bouschet) Grape Berries. Molecules. 2010. V. 15. P. 1141-1153. DOI: 10.3390/molecules15031141.
7. Granese T., Cardinale F., Cozzolino A., Pepe S., Ombra M.N., Nazzaro F., Coppola R., Fratianni F. Variation of Polyphenols, Anthocyanins and Antioxidant Power in the Strawberry Grape (Vitis labrusca) after Simulated GastroIntestinal Transit and Evaluation of in Vitro Antimicrobial Activity. Food Nutr. Sci. 2014. V. 5. P. 60-65. DOI: 10.4236/fns.2014.51008.
8. Flamini R., Tomasi D. The anthocyanin content in berries of the hybrid grape cultivars Clinton and Isabella. Vitis. 2000. V. 39. P. 79-81. DOI: 10.5073/vitis.2000.39.79-81.
9. Zhang K., Yuan L., Li Q., Wang R., Zhang Z.-Z. Comparison of the anthocyanins composition of five wine-making grape cultivars cultivated in the Wujiaqu area of Xinjiang, China. OENO One. 2019. V. 3. P. 549-559. DOI: 10.20870/oeno-one.2019.53.3.2460.
10. Benmeziane F., Cadot Y., Djamai R., Djermoun L. Determination of major anthocyanin pigments and flavonols in red skin of some table grape varieties (Vitis vinitera sp.) by highperformance liquid chromatography - photodiode array detection (HPLC-DAD). OENO One. 2016. V. 50. P. 125-135. DOI: 10.20870/oeno-one.2016.50.3.56.
CONCLUSION
The new RP HPLC method is developed for separation of 3-glucosides and 3,5-diglucosides of five common grape fruit anthocyanidins - delphinidin, cy-anidin, petunidin, peonidin and malvidin. The method as applied for investigation anthocyanin set in grape fruit skins of 43 grape cultivars grow in Belgorod region. The results are discussed according three previously proposed criteria.
The authors declare the absence a conflict of interest warranting disclosure in this article.
Авторы заявляют об отсутствии конфликта интересов, требующего раскрытия в данной статье.
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Поступила в редакцию 12.12.2022 Принята к опубликованию 08.02.2023
Received 12.12.2022 Accepted 08.02.2023
