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In continuation of studies of plants of the genus Polygonatum (Convallariaceae), we studied the flavonoid compositions of roots from the two species P. polyanthemum (Bieb.) A. Dietr and P. glaberrimum C. Koch [1]. The plants were collected in May–June in Georgia (Kodzhori, Saguramo). Total flavonoids (400 g) were obtained from dried and ground rhizomes of each plant by extraction (3 ) with MeOH (80%) for 1 h under reflux. The alcohol extracts were evaporated to an aqueous residue and worked up with CHCl3. The purified aqueous extract was extracted exhaustively with EtOAc, which was evaporated. Total flavonoids were precipitated with CHCl3. Pure compounds were isolated by preparative chromatography over a column of polyamide that was formed in CHCl3. The column was eluted by CHCl3 and CHCl3:EtOH in various ratios. The separation was monitored by paper chromatography using n-BuOH:HOAc:H2O (4:1:2), C6H6:EtOAc:HOAc:H2O (50:50:1:1), and formamide:EtOH (1:3). Similar fractions were combined, evaporated to dryness, and recrystallized from MeOH. Fractions containing a mixture of compounds were rechromatographed over a column of polyamide. The resulting compounds 1–7 were additionally purified by recrystallization from MeOH. The isolated compounds were identified by physicochemical methods (UV, IR, PMR spectroscopy). A comparison of the results with the literature and with data for authentic samples identified 1 as quercetin [2, 3]; 2, isoquercitrin [4, 5]; 3, hyperin [6]; 4, rutin [5, 7]; 5, kaempferol [3, 6]; 6, astragalin [3, 5]; and 7, kaempferol-3-O-D-arabinopyranoside [6, 8]. All aforementioned flavonoids were isolated from P. polyanthemum. The flavonoid composition of P. glaberrimum was qualitatively and quantitatively much poorer. Only 1, 2, 5, and 6 were observed in it. Quercetin (3,5,7,3 ,4 -pentahydroxyflavone) (1), C15H10O7, bright yellow needle-like crystals, mp 312–314°C (MeOH), [M]+ 302. UV spectrum (EtOH, max, nm): 257, 372; +CH3COONa: 270, 406. IR spectrum (KBr, , cm –1): 3450– 3200 (OH), 1665 ( -pyrone C=O), 1615, 1565, 1515 (C=C). PMR spectrum (300 MHz, Py-d5, , ppm, J/Hz): 13.83 (br.s, 5-OH), 11.72 (br.s, 3-OH), 8.46 (1H, d, J = 2.3, H-2 ), 7.95 (1H, dd, J = 2.3, 8.4, H-6 ), 7.23 (1H, d, J = 8.4, H-5 ), 6.64 (1H, d, J = 2.5, H-8), 6.58 (1H, d, J = 2.5, H-6). Quercetin-3-O-D-glucopyranoside (isoquercitrin) (2), C21H20O12, yellow crystals, mp 225–227°C (MeOH), [M]+ 464. UV spectrum (MeOH, max, nm): 255, 265, 360. IR spectrum (KBr, , cm –1): 3200–2900 (OH), 1650 (C=O), 1615–1450 (C=C), 1085, 1055, 1010, 890. PMR spectrum (300 MHz, DMSO-d6, , ppm, J/Hz): 12.44 (1H, br.s, 5-OH), 7.64 (1H, dd, J = 2.4, 8.8 H-6 ), 7.60 (1H, d, J = 2.4, H-2 ), 6.86 (1H, d, J = 8.8, H-5 ), 6.42 (1H, d, J = 2.0, H-8), 6.21 (1H, d, J = 2.0, H-6), 5.62 (1H, d, J = 7.6, glucose H-1 ), 3.2–4.8 (m, glucose 6H). Quercetin-3-O-D-galactopyranoside (hyperin) (3), C21H20O12, yellow crystals, mp 235–236°C (MeOH), [M]+ 464. UV spectrum (MeOH, max, nm): 260, 362; +CH3COONa: 276, 395; +CH3ONa: 276, 405; +CH3COONa + H3BO3: 272, 375. IR spectrum (KBr, , cm–1): 3300 (OH), 1665 (C=O), 1615, 1565, 1515 (C=C), 1095, 1030 (C–O). PMR spectrum (300 MHz, CD3OD, , ppm, J/Hz): 7.56 (1H, dd, J = 2.0, 8.9, H-6 ), 7.44 (1H, d, J = 2.0, H-2 ), 6.79 (1H, d, J = 8.9, H-5 ), 6.42 (1H, d, J = 2.0, H-8), 6.15 (1H, d, J = 2.0, H-6), 5.61 (1H, d, J = 7.8, galactose H-1 ), 3.30–4.6 (m, galactose 6H). Quercetin-3-O-[( -L-rhamnopyranosyl-(1 6)]-D-glucopyranoside (rutin) (4), C27H30O16, yellow powder, mp 215–216°C, [M]+ 610, [ ]D 20 –11.9° (c 0.08, EtOH). UV spectrum (MeOH, max, nm): 258, 360. IR spectrum (KBr, , cm–1): 3450, 1650, 1610, 1520. PMR spectrum (300 MHz, DMSO-d6, , ppm, J/Hz): 7.58 (1H, d, J = 2.0, 8.5, H-6 ), 7.50 (1H

We investigated the composition of anthocyanins from fruit of Rosa spinosissima L. and R. hracziana Tamam. (Rosaceae Juss.) growing near Bata-Bat, Shakhbuz Region, Republic of Azerbaidzhan. Fruit was collected on Sept. 16, 2009, during ripening. Freshly collected fruit (100 g each) was frozen with liquid N2 and pulverized. Pulp was separated from stones. Pulp (50 g each) was ground and extracted at room temperature for 2 d twice with EtOH (95%) and once with MeOH containing HCl (1%). The extracts were combined, filtered, and evaporated to dryness in vacuo at 30°C. The dry solid was treated with CHCl3 and then dissolved in MeOH (50 mL) containing HCl (0.5%). A part of the extract was hydrolyzed by HCl (2 N, 1:1 ratio) on a boiling-water bath for 60 min under reflux [1]. The hydrolysate was cooled. Aglycones were extracted by isoamyl alcohol for 20 h (shaking periodically). The isoamyl extract was studied by 2D chromatography on FN16 paper using Formic and Forestal systems [2]. Two aglycones were detected in the chromatogram. Preparative column chromatography using the Formic system isolated two pure aglycones. Aglycone-1 with Rf 0.32 (Formic) and 0.48 (Forestal) gave absorption maxima with fuchsine dye at 276 and 535 nm (MeOH containing 0.01% HCl). Adding AlCl3 produced a bathochromic shift of 20 nm. Aglycone-2 with Rf 0.12 and 0.31 (in the same systems, respectively) gave absorption maxima with purple dye at 277 and 546 nm. Addition of AlCl3 produced a bathochromic shift of 22 nm. Based on the chromatographic and spectral data and a comparison with authentic samples and the literature [2, 3], the first aglycone was identified as cyanidin; the second, delphinidin. Preparative 2D paper chromatography using n-BuOH:AcOH:H2O (4:1:2) (first direction) and AcOH:HCl (conc.):H2O (15:3:82) (second direction) showed that pulp of R. spinosissima fruit contained three; of R. hracziana, four anthocyanin glycosides. The first three components were identical in both species. Therefore, total anthocyanins from fruit pulp of R. hracziana was analyzed in detail. Preparative column chromatography (acidic cellulose powder) [4] and rechromatography on paper produced four pure anthocyanins. The pure anthocyanins were studied by chromatography and spectroscopy [2, 3, 5]. The anthocyanins were hydrolyzed as described above by neutralizing the hydrolysate with ion-exchange resin. Paper chromatography with authentic sugar samples identified D-glucose in hydrolysates of all glycosides. The chromatographic mobilities of anthocyanins 1 and 2 suggested that they were monoglucosides; 3 and 4, diglucosides. This agreed with results of stepwise acid analysis. The aglycone and sugar were cleaved after stepwise hydrolysis of anthocyanins 1 and 2 for 20 min. However, anthocyanins 3 and 4 formed four components. The first component corresponded to the starting glucoside; the second and third, to a monoglucoside; the fourth, the aglycone. The second spot did not fluoresce. Its Rf value indicated that it was the 3-glucoside. The third spot did fluoresce in UV light with emission analogous to the starting glucoside. It corresponded to the 5-glucoside according to fluorescence and chromatographic mobility. Oxidation of the anthocyanins with H2O2 produced glucose and confirmed that it was bonded to C3 in 1 and 2 and to C3 and C5 in 3 and 4 [1–3, 5, 6]. Thus, cyanidin-3-glucoside, cyanidin-3,5-diglucoside, delphinidin-3-glucoside, and delphinidin-3,5-diglucoside were identified for the first time in total anthocyanins from fruit pulp of R. hracziana using chromatography, spectroscopy, total and stepwise hydrolysis, oxidation by H2O2, and comparison with authentic samples. All components with the exception of delphinidin-3,5-diglucoside were found in R. spinosissima anthocyanidins. The monoglucosides of cyanidin and delphinidin dominated quantitatively the anthocyanin complex from fruit of both Rosa species.

In continuation of research on the chemical composition of plants from the genus Astragalus [1], we studied polyphenolic compounds of A. asper Jacq growing in Caucases foothills on Stavropol highlands near Stavropol [2]. Raw material was collected during flowering. Total polyphenolic compounds were worked up by known methods [1, 3]. For this, air-dried raw material (1 kg, aerial part) was ground to 3-5 mm and extracted with EtOH (70%, 1:10, 4 ). The yield of biologically active compounds was about 94% [3]. The extracts were combined. EtOH was distilled off. The extracts were condensed in vacuo (pressure 4.0 104 N/m2). The condensed extract was purified of lipophilic substances using pentane and heptane. The principal groups of biologically active compounds were separated by selective extraction by CHCl3, EtOAc, BuOAc, and BuOH [4]. Total polyphenolic compounds were obtained from the EtOAc and BuOAc extracts by distilling the solvent, combining them, and drying in a vacuum drying cabinet at 45–50°C and 2.6 104 N/m2 [2]. The yield was 5.2% of air-dried mass of raw material. Paper chromatography (PC) using systems I (BAW, BuOH:AcOH:H2O, 4:1:2), II (AcOH, 15%), and III (AcOH, 2%) and specific detectors established that the polyphenols consisted of 10 compounds. Column chromatography over polyamide using water and a water:EtOH gradient (90:10 10:90) isolated eight pure compounds. Compound 1. C9H8O4, mp 196–198°C (MeOH); Rf 0.83 (system I), 0.28 (system III). UV spectrum (EtOH, max, nm): 217, 234, 300sh, 235. IR spectrum (KBr, max, cm –1): 3350 (OH), 1650-1630 (C=O), 1610-1575-1540 (Ar), 870. Alkaline fusion gave protocatechoic acid. The melting point of the product agreed with that of caffeic acid. Melting point depression was not observed. Compound 1 was identified as caffeic acid by comparison with the literature [4]. Compound 2. C16H18O9, mp 203–206°C (MeOH); Rf 0.63 (system I), 0.66 (system III). UV spectrum (EtOH, max, nm): 245, 292, 328. IR spectrum (KBr, max, cm –1): 2900–2780 (OH), 1740–1715 (ester), 1660-1625 (C=O), 1605–1550 (Ar), 840. Alkaline hydrolysis formed caffeic acid and D-quinic acid. One spot was observed in a chromatogram of a mixed sample of 2 and chlorogenic acid. Melting point depression also was not observed for 2 and chlorogenic acid. This compound was identified as chlorogenic acid [1, 4]. Compound 3. C15H10O7, mp 310–312°C (MeOH); Rf 0.64 (system I), 0.04 (system II). UV spectrum (EtOH, max, nm): 375, 265. IR spectrum (KBr, max, cm –1): 3380–3300 (OH), 1665 (C=O), 1615-1565-1515 (Ar). Fusion with crystalline KOH formed phloroglucinol and protocatechoic acid. Melting point depression of 3 and authentic quercetin was not observed. The compound was identified as quercetin [4]. Compound 4. C21H20O11, mp 240–242°C (MeOH); [ ]D 20 –59° (c 0.1, DMF). UV spectrum (EtOH, max, nm): 372, 257. Acid hydrolysis of the compound (1% H2SO4) formed an aglycon with mp 310–312°C that was identified by UV and IR spectroscopy as quercetin. The sugar component was identified by PC as D-glucose. The site of attachment of the sugar component to the aglycon was determined using UV spectroscopy with ionizing and complexing reagents. Comparison of molecular rotations in the studied compound established the -configuration for the glycoside bond. Based on the results the compound was identified as quercimeritrin [4]. Compound 5. C21H30O16, mp 190–192°C (MeOH); Rf 0.45 (system I), 0.51 (system II). UV spectrum (EtOH, max, nm): 256, 264sh, 354. IR spectrum (KBr, max, cm –1): 3595-3400 (OH), 1660-1658 (C=O), 1605–1575–1510 (Ar), 1085, 1062, 1025, 980, 900 (sugar). Acid hydrolysis (0.25% H2SO4, stepwise hydrolysis) formed quercetin-3-O-glycoside and L-rhamnose; then quercetin, D-glucose, and L-rhamnose. This was confirmed by PC with authentic samples. The compound was identified as rutin (quercetin-3-rutinoside) [4].

Eight alkaloid derivatives of tropane, among which the major ones were convolvine and convolamine; the minor ones, phyllalbin, convolidine, nortropine, phyllalbin N-oxide, convolvidine, and conpropine, have been isolated previously from the aerial part of Convolvulus subhirsutus growing in Uzbekistan [1, 2]. Quantitative analysis found 0.6% total bases in the aerial part; 1.6%, in the roots. The contents of the major alkaloids convolvine and convolamine were 45-50% of the total mixture. In continuation of research on alkaloids of this plant collected near Tepar, Tashkent Oblast′, a base (15 mg) with mp 144–145°C that was identified as the known alkaloid convolicine was isolated by chromatography over a column of Al2O3 from the non-phenol part of total alkaloids after removal of convolvine and convolamine [3]. The benzene eluates contained an alkaloid (10 mg) with mp 178–179°C that was identified as confolidine. The extract was neutralized to pH 7 after removal of the alkaloids, diluted with water in a 1:1 ratio, and treated successively with hydrocarbons, EtOAc, and CHCl3. Paper chromatography and comparison with authentic samples detected in the remaining aqueous phase D-mannose, D-glucose, and D-galactose. Crystals with mp 136°C that were identified as β-sitosterol were isolated from the hydrocarbon fraction. The mother liquor was treated with acetone, evaporation of which produced crystals with mp 81–82°C that were identified as acetamide. Treatment of the mother liquor with acetone:MeOH isolated veratric acid with mp 181–182°C. The CHCl3 fraction was also treated with acetone, evaporation of which produced crystals with mp 201–202°C, Rf 0.5 (system 1). The IR spectrum of the compound contained absorption bands for active H (3115 cm −1, –OH), –CH (3045), an aromatic ring (1605, 1562, 1506), and a conjugated carbonyl (1702) [4, 5]. The UV spectrum showed absorption maxima at 297 and 345 nm that were characteristic of 6,7-dihydroxycoumarins [6]. The PMR spectrum of the coumarin had resonances for aromatic protons at 6.15 and 7.79 ppm as two doublets (J = 9.6 Hz) for the C-3 and C-4 protons; at 7.05, a singlet for the C-5 proton; at 6.71, a singlet for the C-8 proton. A 3H singlet for the aromatic methoxyl was observed at stronger field (3.85). Resonances for H-4 and H-8 were broadened due to through-space spin—spin coupling between them with J of H-4 and H-8

The genus Ammoides (Apiaceae) tribe of Ammineae includes two species in Algeria, one of which is endemic: Ammoides atlantica (Coss. et Dur.) Wolf; the other one, Ammoides pusilla (Brot.) Breistr, is widespread in the Mediterranean region [1]. The aerial parts of Ammoides atlantica were collected from Megress Mountain (Eastern Algeria) at 1500 m above sea level during June 2004 and identified by Dr. H. Laouer. A voucher specimen (B6308) has been deposited in the Museum d’Histoire Naturelle of Nice (France). The air-dried powdered parts (700 g) of A. atlantica were macerated three times in boiling methanolic solution (70%). The MeOH extract was concentrated to dryness, the residue was dissolved in boiling water (600 mL) after filtration, and the residue was extracted successively three times with AcOEt and n-butanol (3×200 mL) to give 2.5 and 22.7 g of the respective residues. Solvents were evaporated and the residues of n-BuOH and AcOEt extracts were dissolved in small volumes of MeOH. Analysis by two-dimensional paper chromatography using 15% AcOH and BAW (n-BuOH–AcOH–H2O, 4:1:5, upper phase) as solvents indicated that extracts of n-BuOH and AcOEt contain identical separation of flavonoids. The AcOEt extract was applied to a column of silica gel and eluted with a gradient of n-hexane–AcOEt–MeOH with increasing polarity to obtain nine fractions (A-I). Concentration under reduced pressure of fractions B, D, and G gave three precipitates, which were filtered and washed with MeOH to obtain three compounds 1 (10 mg), 2 (32 mg), and 3 (40 mg). Fraction C was purified by preparative TLC on silica gel eluted with n-hexane–AcOEt (2:8) to afford compound 4 (10 mg). Final purification of each compound for spectral analysis was carried out using MeOH over Sephadex LH-20 (Pharmacia Fine Chemicals), prior to UV, MS, and 1H spectral analysis [2, 3]. Hydrolysis of the glycosides 3 and 4 (HCl 0.1 N, 2 h) yielded the sugar residues and the aglycones, all of which were chromatographed with authentic samples. 1H NMR spectra were recorded in CD3OD at 300 MHz. All the structures were in good agreement with literature data. Compound 1, C15H10O5, mp 345°C, identified as apigenin [4]. Compound 2, C15H10O6, mp 330°C, identified as luteolin [5]. Compound 3, C21H20O10, mp 220–222°C identified as apigenin 7-O-glucoside [6]. Compound 4, C21H20O11, mp 239–242°C, identified as luteolin 7-O-glucoside [7]. Only one flavonoid, luteolin 7-O-glucoside [7], has recently been reported from an Egyptian A. pusilla. Compounds 1–4 were isolated for the first time from A. atlantica, and compounds 3, 4 are reported for a second time from a Ammoides species [6, 7]. Compounds 1 and 2 are new for the genus Ammoides.

In continuation of examination of plants of the Alhagi genus, we investigated the aerial part of Persian camel’s thorn (A. persarum) collected during flowering in western Kazakhstan District [1-3]. Two-dimensional paper chromatography of the aqueous acetone (1:1) extract with elution by butan-1-ol—acetic acid—water (40:12.5:29) and 15% acetic acid detected in the aerial part 12 flavonoids, 7 of which were isolated pure. Finely ground air-dried raw material was exhaustively extracted with aqueous acetone (1:1). The extract was concentrated, distilled, and purified of lipophilic substances. The total flavonoids were extracted with ethylacetate and separated over a polyamide column. The following flavonoids were isolated pure: Compound 1, C H O , mp 306-308°C, UV (λ , C H OH, nm): 375, 265. Characterized as quercetin. 15 10 7 max 2 5 Compound 2, C H O , mp 303-307°C, UV (λ , C H OH, nm): 370, 268 (sh), 255. Identical to isorhamnetin [4]. 16 12 7 max 2 5 Compound 3, C H O , mp 182-184°C, [α] -175° (ethanol), UV (λ , C H OH, nm): 350, 256. Identified as 21 20 11 D max 20 2 5 quercetin 3-O-α-L-rhamnopyranoside. Compound 4, C H O , mp 238-240°C, [α] -52° (ethanol), UV (λ , C H OH, nm): 360, 258. Hydrolysis 20 20 12 D max 20 2 5 produced quercetin and arabinose. The constants of 4 correspond with those of quercetin-3-O-α-L-arabofuranoside. Compound 5, C H O , mp 220-222°C, [α] -56° (ethanol), UV (λ , C H OH, nm): 368, 270 (sh), 254. 22 22 12 D max 20 2 5 Characterized as isorhamnetin 3-O-β-D-glucopyranoside [1, 4]. Compound 6, C H O , mp 257-259°C, [α] -87° (ethanol), UV (λ , C H OH, nm): 364, 272 (sh), 258. 21 20 20 D max 20 2 5 Identified as isorhamnetin 3-O-α-L-arabopyranoside. Compound 7, C H O , mp 165-167°C, [α] -64.5° (ethanol), UV (λ , C H OH, nm): 362, 278 (sh), 256, IR 36 46 20 D max 20 2 5 spectrum (KBr, ν, cm ): 3300, 3400, 2945, 1665, 1070, 1060, 1055, 1004, 890, 835, 866. FAB/MS, m/z 798, m/z of the -1

In continuation of the study of the flavonoid content of Astragalus L. species, we investigated the aerial part of Astragalus tana L. (Fabaceae L.) collected during flowering in Kartli in Tana region (Georgia). The extract was purified by the literature method [1]. Two-dimensional paper chromatography revealed at least six flavonoids. It was established preliminarily that the starting material contained mainly kaempferol glycosides. First the extract was fractionated on a polyamide column with elution by water, aqueous alcohol (45%), and alcohol (96%). Fraction 2 contained the bulk of the flavonoglycosides. Rechromatography on the column of this fraction produced 1-4. Compound 1, C H O , mp 196-198(C (ethanol), [ ] -19( (c 0.1, ethanol), 375, 268 nm. A dilute acid 21 20 11 D max 20 EtOH

The structure of philoside B — a gypsogenin nonaoside isolated from the roots ofGypsophila patrinii — has been established.

1. The peptides of a tryptic hydrolyzate of the 7S-globulin have been isolated and characterized.

The peptides of thyroglobulin obtained by enzymatic hydrolysis with pronase and by two-dimensional paper chromatography have been investigated by neutron activation analysis in order to determine the micro amounts of iodine in each peptide. The greatest amount of iodine is present in the 2nd, 7th, 12th, 13th, 14th, and 15th peptides. The bulk of the iodine is present in the 14th and 15th peptides, which contain 17.7 and 34.3%, respectively, of the total iodine. There is a definite direct correlation between the concentrations of iodine in them.

The water-soluble polysaccharides of the rhizomes of thePolygonatum sewerzowii Regl. have been studied. A method has been developed for separating the polysaccharide fraction. A new native acetylated glucomannan has been isolated which has been called severan and consists of manno- and glucopyranose residues linked with one another by β-1 → 4 bonds in a ratio of 15.4:1.

1. The nitrobenzene oxidation of the natural lignins of kenaf stems and of DLA fractions has been performed. A study of the oxidation products by gas-liquid chromatography has shown that the kenaf lignins are constructed of three types of structural units: p-coumaryl, guaiacyl, and syringyl, with the latter predominating. 2. Among the products of the nitrobenzene oxidation of fractions of kenaf dioxane lignin paper chromatography has shown the presence of dihydrodivanillin, which is a representative of structures with a biphenyl bond.