PHENOL GLYCOSIDES AND THEIR USE IN THE TREATMENT OF UROLITHIASIS

The present invention relates to novel derivatives of polyphenol glycoside or polyols, pharmaceutical composition on the basis of the new glycosidic derivatives of polyphenols, or polyalcohols and their use, which are capable of dissolving calcium oxalate, applicable in the treatment of urinary tract disorders, in particular urolithiasis.

Urolithiasis is a chronic disease characterized by the formation and presence in the urinary system of inorganic deposits, formed of the components present in normal or pathologically altered urine. The majority of urinary stones are the kidney stones, which consist of inorganic salts, mainly calcium as calcium oxalate (Moe OW; Kidney Stones: an Pathophysiology medical management, Lancet, 2006, Vol 367, pp 333-344) .

In the last few years of the last century there was a rapid development of invasive treatments for urolithiasis. Currently, most patients used invasive methods such as crushing stones with shock waves ESWL (SH Mousavi-Bahar , Mehrabian S, Moslemi MK, Int Urol Nephrol 2011, 43, pp 983- 987), and other endourological treatments PCNL (Rozanski W., Klimek L, M Lipinski, Klis R., Selected examples of complications after minimally invasive treatment for urolithiasis, Central European Journal of Urology, 2012, 65/2, pp 80-83) . These modern methods, however, cause post-operative complications and are characterized by a high percentage of remission over the next five years from the time of surgery (Coe FL, Evan A, Worcester E., Kidney stone disease, J Clin Invest., 2005.115 (10), pp 2598 -608). Accordingly, it is desirable to find an alternative method of treatment of urolithiasis. In recent years, particularly interesting are the search for chemical compounds of natural origin as well as synthetic with kidney stone dissolving properties.

Plant materials play an important role in the prevention of urinary tract, including kidney stones. On the market there were two of them Rubinex plant preparations and Rubiolizyna having in its composition madder root extract (Rubia tinctorum L.) . They were widely used in the treatment of kidney stones, until it senses a mutagenic potential of one of the components. They were then completely withdrawn from the market, and mutagenic compound proved to be lucydyna - 1,3- dihydroxy-2-hydroxymethyl-9 , 1 O-anthraquinone (Westendorf J., Pfau W Schulte, A., Carcinogenicity and DNA adduct formation in the ACI Observed rats after long-term treatment with madder root, Rubia tinctorum L . , Carcinogenesis, 1998, 19 (12), pp 2163-2168) . The studies conducted show that for the inhibition of a kidney stone formation are responsible anthraquinone derivatives, which are capable of complexing calcium ions, due to the presence of the C = 0 and OH (Yasui Y., Takeda N., Identification of a mutagenic substance, in Rubia tinctorum L. (madder) root, as lucidin, Mutation Research, 1983, 121 (3-4), pp 185-190) . Literature sources report that the compounds having a keto group and hydroxyl have complexing properties, in particular with regard to the bivalent metal and have potential use in the treatment of kidney stones (Wai CM, Wang S., Supercritical fluid extraction: metals as complexes, Journal of Chromatography A, 1997, 785, 369-383) .

Plant preparations are available in the pharmacy sales, for use in diseases of the urinary tract, including kidney stones. The most common available in pharmacies (without prescription) are pastes: Fitolizyna and Debelizyna. They are intended for the preparation of oral suspension. They are prepared by extraction of herbal ingredients, and then compacting the resulting extract in vacuum evaporators. The resulting extract is gelled using water-soluble substances, usually agar and glycerol [ [Herbapol Warsaw, information on the product manufacturer and Bogdan Kedzia B . ; Vegetable pastes for the preparation of oral suspensions. Phytotherapy progress in 2008 ; (3) : 165-169] . However, these measures are mainly anti-inflammatory and diuretic, in contrast to substances which are the subject of the patent. The latter remove the stones in a chemical form non-toxic complexes with calcium ions, which are then more easily removed from the urinary tract.

The technical problem posed prior to the present invention is to provide such synthetic compounds that will exhibit the properties of forming complexes with calcium ions, thereby affecting the dissolution of the calcium oxalate and inhibit its formation, the resulting compounds would be soluble in water, do not exhibit mutagenicity and would be safe for humans and their production could be done in a simple and fast way, which does not require the use of sophisticated equipment required for the synthesis . Equipment necessary to obtain the compound covered by patent is the basic equipment used in typical organic industry plants.

The first object of invention relates to novel derivatives of polyphenol glycoside or polyalcohols of Formula 1

wherein Rl, R2, R3 is selected from the group consisting of H, OH, C(0)R4, C(0)0R4, 0(GlyH3)nH, wherein n = 0, 1, 2, 3, while R4 is selected from the group consisting of H, alkyl, and Gly is a mono- or disaccharide residue. Preferably the polyphenol of the invention contains at least one aromatic ring. In a further preferred embodiment of the invention the polyphenol contains at least two hydroxyl groups attached to aromatic rings, or one hydroxyl group and one carbonyl group attached to the aromatic rings. In a further preferred embodiment of the invention glycoside contains from one to four sugar units.

A second object of the invention is a pharmaceutical composition comprising a novel derivatives of polyphenol glycoside or polyalcohols , as defined in the first object of the invention.

A third object of the invention is the use of the new glycosidic derivatives of polyalcohols or polyphenols, as defined in the first object of the invention to treat urinary tract disorders, in particular urolithiasis.

The novel polyphenol glycoside of polyalcohols of the invention exhibit properties of forming complexes with calcium ions, which dissolve calcium oxalate and inhibit its formation thus allowing the treatment of kidney stones. Synthetic compounds that constitutes the present invention have in their structure a plurality of hydroxyl groups, carboxyl, ester, ether, or carbonyl, which due to the characteristics coordinating calcium ions bind to the ligands via oxygen atoms. Furthermore, the compounds are soluble in water which allows their use in a wide range of pharmaceutical compositions .

The invention is described in the embodiments and the drawings, in which Fig. 1 shows the structure of the glycosidic derivatives of phenol, and Fig. 2 shows example of cytotoxicity estimation for compound 106 (toxixc) and 101 (no toxic) . Glycoside 106 (at a concentration of 1500 and 750 ug / ml) show toxicity to L929 cell line and the A549, the cells are shrunken, round a granular cytoplasm, a large number of cells is dead. No toxic effect is observed for compound 101.

Example 1 - Synthesis of donors glikozykacji - acetylation of sugars - general procedure S-Ac

To a round bottom flask 56mmol corresponding sugar was added and was dissolved in acetic anhydride. After addition of iodine the mixture was stirred with a magnetic stirrer, until the color turned dark cherry-red solution, which was manifested with a strong heating of the flask contents. It was then quenched with 100 ml of dichloromethane and washed in a separatory funnel with 10% aqueous Na ₂ S ₂ C>3. The organic layer was collected in a flask and the aqueous layer was further extracted twice with dichloromethane (two times 50ml) . The collected organic fractions were combined and washed in a separatory funnel with 10% aqueous Na ₂ CC>3, then with water and dried over anhydrous Na ₂ SC>4. After removal of the drying agent, the solvent was evaporated and the resulting oil was dissolved in hot ethanol and the flask was placed in a refrigerator. After several hours white crystals fell out from the solution, which were filtered under reduced pressure and air-dried for 48 hours. The structure of the resulting products was confirmed by NMR studies. a) 1, 2, 3, 4, 6-penta-O-acetyl- -D-glucopyranose (21). The synthesis of acetylated glucose was carried out according to the general procedure acetylation of sugars (S-Ac) . To the reaction lOg of D-glucose and 40 ml of acetic anhydride and 0.5 g of iodine talyst was used. The washout of iodine was used using 50 ml of 10% aqueous solution of Na ₂ S ₂ C>3. 50 ml of 10% aqueous solution of Na ₂ CC>3 was used to wash out the the acid used. The product was crystallized from 20 ml ethanol. was 74%. Mp = 112 ⁰ C. NMR analysis confirmed receipt of pure product .

1HNMR ( 300MHz, CDC13) , 6.33 (d, 1H, CHO, 3JH-H=3.7Hz), 5.47 (t, 1H, CHO, 3JH-H=9.8Hz) , 5.17-5.08 (m, 2H, CHO) ,

4.27 (dd, 1H, CH20, 2JH-H=12.7Hz, 3JH-H=4.2Hz, 3JH-H=4.0Hz) , 4.15- 4.07(m,lH,CH2O;lH,CHO) , 2.18 ( s, 3H, CH3CO) , 2.09 ( s , 3H , CH3CO ) , 2.04 (s, 3H, CH3CO) , 2.03 ( s, 3H, CH3CO) , 2.02 ( s, 3H, CH3CO) . b) 1, 2, 3, 4, 6-penta-0-acetyl^-D-galactopyranose (22) The synthesis of acetylated galactose was carried out according to the general procedure of acetylation of sugars (S-Ac) . To the reaction lOg of D-glucose and 40 ml of acetic anhydride and 0.5 g of iodine talyst was used. To washout of iodine 50 ml of 10% aqueous solution of Na ₂ S ₂ 0 ₃ was used. The acid used to wash out the 50 ml of 10% aqueous solution of Na2C03. The product was crystallized from 20 ml of ethanol to give 15.3g of pure product as a white solid. Yield was 70%. Mp = 90-92 ⁰ C. NMR analysis confirmed receipt of pure product .

1HNMR (300MHz, CDC13) , 6.37 (d, 1H, CHO, 3JH-H=1.5Hz ) ,

5.50 (d, 1H, CHO, 3JH-H= 1.5Hz), 5.34-5.33 (m, 2H, CHO) , 4.36- 4.32(m,lH), 4.15-4.08 (m, 2H, CH20) , 2.16 ( s, 3H, CH3CO) ,

2.05 (s, 3H, CH3CO) , 2.04 ( s, 3H, CH3CO) , 2.04 ( s, 3H, CH3CO) , 2.00 (s, 3H, CH3CO) . c) 1, 2, 3, 4, 6-penta-0-acetyl^-D-galactopyranose / method with pyridine (22 ')

To a flask of 250 ml lOg (56mmol) of D-glucose and 40 ml of acetic anhydride and 50 ml pyridine was added and the mixture was stirred with a magnetic stirrer. Sugar was dissolved after approx. one day. Then extraction was performed - the reaction mixture is dissolved in 100 ml of dichloromethane and washed twice with 250 ml of oxalic acid solution (10%) and 50 ml of water. The organic layer was washed with 50 ml 10% NaHCC>3 and 50 ml of water. The solution was dried over anhydrous Na ₂ SC>4. After removal of the drying agent the solvent was evaporated and the yellow oil was recrystallized from 50 ml of hot ethanol. After several hours the evolution of a white flaky precipitate was formed which was filtered off with suction and dried in air. 10.5g of pure product were obtained. The reaction yield is 48%. Mp = 89-92 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR (300MHz, CDC13) , 5.67 (d, IH, CHO, 3JH-H=8.3Hz ) ,

5.40 (d, IH, CHO, 3JHH=3.0Hz) , 5.31 (q, IH, CHO, 3JH-H=10.3 , 3JH- H=8.3), 5.05 (dd, IH, CH20, 2JH-H=10.4, 3JHH=3.4, 3JH-H=3.4Hz) ,

4.14-4.00 (m, 2H, CHO; IH, CH20) , 2.14 ( s, 3H, CH3CO) , 2.10

(s, 3H, CH3CO) , 2.02 (s, 6H, CH3CO) , 1.97 ( s, 3H, CH3CO) . d) 2,3,4,6,1 ', 2', 3 ', 6 ' -octa-O-acetyl-4-O- ( -D- galactopyranosyl^-D-glucopyranose) and 2.3, 4,6,1 ', 2', 3 ', 6 ' -octa-O-acetyl-4-O- ( β-D-galactopyranosyl-β-D- glucopyranose ) (23)

The synthesis of acetylated lactose was performed according to the general procedure of sugars acetylation (S-Ac) . To the reaction 19g of D-glucose and 80 ml of acetic anhydride and 1.0 g of iodine talyst was used. To washout of iodine

100 mL of 10% aqueous solution of Na ₂ S ₂ 0 ₃ were used. The product was crystallized from 40 ml of ethanol to give 23.4g of pure product as a white solid, as a mixture of anomers a and β (77% and 23%). Yield was 60%. Mp = 76-77 ⁰ C. NMR analysis confirmed receipt of pure product. 1HNMR ( 300MHz, CDC13) , 6.24 (d, 1H, CH O, 3JH-H=3.6Hz) ,

5.65 (d, 1H, ΟΗβΟ, 3JHH=8.4Hz) , 5.45 ( t , 1H, CHO, 3JH-H=10.2Hz ) ,

5.34 (d, 1H, CHO, 3JH-H=2.4Hz) , 5.144.92 (m, 3H, CHO) , 4.49-

4.41 (m, 2H, CH20) , 4.17-3.97(m,2H, CHO; 2H, CH20) , 3.903.67(m,3H,CHO) , 2.17 ( s, 3H, CH3CO) , 2.15 ( s, 3H, CH3CO) ,

2.12 (s, 3H, CH3CO) , 2.06 (s,3H, CH3CO) , 2.05 ( s, 3H, CH3CO) ,

2.04 (s, 3H, CH3CO) , 2.00 ( s, 3H, CH3CO) , 1.96 (s,3H, CH3CO) . e) 2,3,4,6,1 ', 2', 3 ', 6 ' -octa-O-acetyl-4-O- ( -D- glucopyranosyl- -D-glucopyranose ) and 2.3, 4,6,1 ', 2', 3 ', 6 ' -octa-O-acetyl-4-O- ( β-D-glucopyranosyl-a-D- glucopyranose (24)

The synthesis of acetylated maltose was carried out according to the general procedure of sugars acetylation (S-Ac) . To the reaction 19g of D-glucose and 80 ml of acetic anhydride and 1.0 g of iodine talyst was used. To washout of iodine

100 ml of 10% aqueous solution of Na ₂ S ₂ 03 was used. The product was crystallized from 40 ml of ethanol to give 24. lg of pure product as a white solid. The yield was 62%. Mp = 156-158 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR ( 300MHz, CDC13) , 6.23 (d, 1H, CHaO, 3JH-H=3.9Hz),

5.73 (d, 1H, ΟΗβΟ, 3JHH=8. lHz) , 5.61-5.25 (m, 4H, CHO) ,

5.07 (t, 1H, CHO, 3JH-H=9.9Hz), 4.95(dd,lH, CHO, 3JHH=10.2Hz, 3JH-

H=3.6Hz), 4.86(dd,lH, CHO, 3JH-H=10.5Hz, 3JH-H=4.2Hz), 4.44(dd,lH, CH20, 2JH-H=12.3Hz , 3JH-H=2.4Hz ) , 4.26-

4.18 (m, 2H, CH20) , 4.13-3.84 (m, 2H, CHO, 2H, CH20) ,

2.21 (s, 3H, CH3CO) , 2.13 (s, 3H, CH3CO) , 2.09 (s, 3H, CH3CO) ,

2.06 (s, 3H, CH3CO) , 2.02 ( s, 3H, CH3CO) , 2.01 ( s, 3H, CH3CO) ,

2.00 (s, 3H, CH3CO) , 1.98(s,3H, CH3CO) . Example 2 - Synthesis of glycosylation donors - Brominations of acetylated sugars - general procedure S-Br In a round bottom flask 26mmol of appropriate acetylated sugar was dissolved in glacial acetic acid with stirring on a magnetic stirrer. Then, at low speed stirrer, slowly 33% solution of bromine in acetic acid was added dropwise and the mixture was allowed while stirring for 15 minutes. At this time, the solution adopted a dark yellow color. To the mixture 100 ml of dichloromethane was added and the mixture was transferred to a separatory funnel, where the mixture was washed with two portions (200 ml) of cold water. The organic layer was washed with two portions of 10% NaHCO 3 and then 100 ml of water. The organic layer was dried over anhydrous sodium sulfate. After filtering off the drying agent the solvent was evaporated on a rotary evaporator and the residual thick oil acetylated sugar bromide was dried in a desiccator over P ₂ O ₅ for 3-7dni period until the compound was completely dry.

a) 2, 3, 4, 6-tetra-O-acetyl- -D-glucopyranose (31)

The synthesis was performed according to the general procedure of bromination of acetylated sugars (S-Br) . To the reaction lOg of 1 , 2 , 3 , 4 , 6 -penta-O-acetyl- -D-glucose was used, which was dissolved in 20ml acetic acid and then 30 ml of 33% HBr/ CH3COOH was added dropwise. To washout 200 ml of 10% NaHC0 ₃ was used to give 9.3g of pure product as a hard light gray mass. Yield was 88%. Mp = -. NMR analysis confirmed receipt of pure product .

1HNMR (300MHz, CDC13) , 6.61 (d, IH, CHBr , 3JH-H=4.0Hz ) ,

5.56 (t, IH, CHO, 3JH-H= 9.7Hz), 5.16 ( t , IH, CHO, 3JH-H=9.9Hz ) , 4.84 (dd, IH, CH20, 2JH-H=10.0Hz, 3JH-H=4. lHz, 3JHH=4.0Hz ) , 4.36- 4.24 (m, 2H, CHO) , 4.13 (dd, IH, CH20, 2JH-H=1.9Hz , 3JH-H=12.3Hz ) , 2.10 (s, 3H, CH3CO) , 2.09 ( s, 3H, CH3CO) , 2.05 ( s, 3H, CH3CO) ,

2.03 (s, 3H, CH3CO) . b) 2, 3, 4, 6-tetra-O-acetyl-a-D-galactopyranose (32)

The synthesis was performed according to the general procedure of bromination of acetylated sugars (S-Br) . The lOg of 1 , 2 , 3 , 4 , 6 -penta-0-acetyl- β-D-galactose was dissolved in 20 ml of acetic acid and then 30 ml of 33% HBr / CH3COOH was added dropwise. To washout 200 ml of 10% NaHCC>3 was used to give 8.5g of pure product as a hard light gray mass. Yield was 81%. Mp = -. NMR analysis confirmed receipt of pure product.

1HNMR ( 300MHz, CDC13) , 6.68 (d, 1H, CHO, 3JH-H=3.9Hz),

5.50 (d, 1H, CHO, 3JH-H= 3.0Hz), 5.38 ( dd, 1H, CH20, 2JH-H=10.8Hz , 3JH- H=3.3Hz, 3JH-H=3.3Hz) , 5.03 (dd, 1H, CH20, 2JH-H=10.6Hz , 3JH-

H=3.9Hz, 3JH-H=3.9Hz) , 4.46 ( t , 1H, CHO, 3JH-H=6.3Hz ) , 4.20-4.06 (m,2H,CHO), 2.13 ( s, 3H, CH3CO) , 2.09 ( s, 3H, CH3CO) ,

2.04 (s, 3H, CH3CO) , 1.99 ( s, 3H, CH3CO) . c) bromide 2,3,4,6,2 ', 3', 6 ' -hepta-O-acetyl-4-O- ( -D- galactopyranosyl^-D-glucopyranose ) ( 33 ) The synthesis was performed according to the general procedure of bromination of acetylated sugars (S-Br) . The 18g 2,3,4,6,1 ', 2', 3 ', 6 ' -octa-O-acetyl-4-Olaktopiranozy was dissolved in 40 ml of acetic acid and 60 ml of 33% HBr / CH3COOH was added dropwise. To washout 400ml of 10% NaHC0 ₃ was used to give 15.5g of pure product as a hard light gray mass. Yield was 82%. Mp = -. NMR analysis confirmed to obtain pure product.

1HNMR ( 300MHz, CDC13) , 6.52 (d, 1H, CHaO, 3JH-H=3.9Hz),

5.55(t,lH,CHO,3JH-H= 9.6Hz), 5.54 (d, 1H, CHO) , 5.16- 5.10 (m, 1H, CHO) , 4.96 ( dd, 1H, CH20, 2JH-H=10.2Hz , 3JHH=3.9Hz , 3JH-

H=3.3Hz), 4.75 (dd, 1H, CH20, 2JH-H=19.9Hz, 3JH-H=3.9Hz) , 4.52- 4.48(m,2H, CHO), 4.23-4.05 (m, 2H, CHO; 2H, CH20) , 3.88-

3.86 (m, 2H, CHO) , 2.16 ( s, 3H, CH3CO) , 2.13 ( s, 3H, CH3CO) , 2.09 ( s, 3H, CH3CO) , 2.07 ( s, 3H, CH3CO) , 2.06 (s,3H, CH3CO) ,

2.05 (s, 3H, CH3CO) , 2.0 (s, 3H, CH3CO) . d) bromide 2,3,4,6,2 ', 3', 6 ' -hepta-O-acetyl-4-O- ( -D- glucopyranosyl- -D-glucopyranose ) (34) The synthesis was performed according to the general procedure of bromination of acetylated sugars (S-Br) . The 18g 2,3,4,6,1 ', 2', 3 ', 6 ' -octa-O-acetyl-4-0 maltopiranozy was dissolved in 40 ml of acetic acid and then 60 ml of 33% HBr / CH3COOH was added dropwise. To washout 400ml of 10% NaHCC>3 was used to give 14.3 g of pure product as a hard light gray mass. Yield was 75%. Mp = -. NMR analysis confirmed to obtain pure product .

1HNMR ( 300MHz , CDC13 ) , 6.49 (d, 1H, CHO, 3JH-H=4.2Hz ) , 5.63-

5.57 (m, 1H, CHO) , 5.415.33 (m, 2H, CHO) , 5.06 ( t , 1H, CHO, 3JH- H=9.9Hz), 4.85 (dd, 1H, CHO, 3JH-H=10.5Hz, 3JHH=3.9Hz) ,

4.70 (dd, 1H, CHO, 3JH-H=9.9Hz, 3JH-H=4.2Hz) , 4.51 (m, 1H,CH20) ,

4.284.21 (m, 3H, CH20) , 4.08-4.02 (m, 3H, CHO) , 3.97-3.92 (m, 1H, CHO) , 2.13 (s, 3H, CH3CO) , 2.08 ( s, 3H, CH3CO) , 2.07 ( s, 3H, CH3CO) ,

2.06 (s, 3H, CH3CO) , 2.03 ( s, 3H, CH3CO) , 2.02 ( s, 3H, CH3CO) , 2.00 (s, 3H, CH3CO) .

Example 3 - Synthesis of glycosides derivatives of phenols - Glycosylation polyphenols and their derivatives - general procedure S-KK

To a round bottom flask 5.0 g (12mmoli) acetylated sugar bromide was added and dissolved in 15 ml of quinoline grinding the bromide pellets using a glass spatula. After complete dissolution, llmmoli the appropriate phenol was added and the flask was placed on a magnetic stirrer. To the mixture 3.0 g of silver oxide (13mmol) was added. After a few minutes the mixture had become very thick - in the case where stirring was stopped, additional amount of quinoline was added. During the reaction color of mixture turn brown. The reaction was monitored by TLC (CHC13 / MeOH 10: 1, silica gel, 12, combustion H2S04) . After approx. 2 hours to the reaction mixture 100 ml of chloroform was added and the mixture was filtered from the precipitate of silver salt. To washout the quinoline the solution was washed with 5% sulfuric acid solution (twice molar excess with respect to quinoline used, ie. 17ml of acid for each 1ml of quinoline), added in two portions. If emulsion was formed the mixture was allowed to stand for several hours to separate the layers. The organic layer was finally washed with 50 ml of water and dried over anhydrous calcium chloride. After filtration of the drying agent and solvent evaporation an oil as obtained which was recrystallised from methanol, ethanol or a mixture of methanol-ether or etanol-eter (ether was added to alkohol solution until slight turbidity) and the flask was placed in a refrigerator. Some compounds crystallize in the form of crystals and other as an oil. If the product fell out of solution in the form of crystals it were filtered under reduced pressure and dried at room temperature. On the other hand, in the case where the product came out in the form of an oil, it was purified by column chromatography (eluent CHC13 / MeOH 10: 1) . Glycoside fractions were collected, evaporated and recrystallized from methanol or ethanol in the chamber with diethyl ether. As a result of the process an oil was formed which was decanted from the solution and the residue was dried under vacuum.

a) 2-methoxycarbonylphenyl-0- (2,3,4, 6-tetra-0-acetyl^-D- glucopyranoside )

The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.67g (llmmol) of o-hydroxybenzoate (methyl salicylate), and 3.0 g (13mmol) of silver oxide was used. 3.4g of a white solid product was obtained as a result of the crystallization from ethanol solution. The yield was 64%. Rf = 0.77 (CHC13 / MeOH 20: 1, silica gel, 12, combustion H2S04). Mp = 158-159 ⁰ C. NMR analysis confirmed to obtain pure product .

IHNMR ( 300MHz, CDC13) , 7.73 (dd, IH, 3JH-H=1.7Hz, 3JH-H=10Hz) ,

7.42 (t≡ddd, IH, ArH, 3JH-H=8.4Hz , 3JH-H=7.3Hz , 4JH-H=1.6Hz) , 7.14(d,lH, ArH, 3JH-H=7.5Hz ) , 7.12 ( t , IH, 3JHH=7.5Hz ) , 5.36- 5.29 (m, 2H,CHO) , 5.26-5.09 (m,2H,CH20),

4.29 (dd, IH, CH20, 2JHH=12.2Hz, 3JH-H=5.3Hz), 4.20(dd,lH, CH20, 2JH- H=12.2Hz, 3JH-H=2.4Hz) , 3.8 (s,3H, CH300C) , 2.10 ( s, 3H, CH3CO) , 2.08 (s, 3H, CH3CO) , 2.05 ( s, 3H, CH3CO) , 2.04(s,3H, CH3CO) . 13CNMR( 300MHz, CDC13) , 170.56(CO), 170.26(CO), 169.42(CO), 169.35(CO), 166.54(CO), 155.56 (COOCH3 ) , 133.09 ( 1C, ArC) , 131.38 (1C, ArC) , 123.11 ( 1C, ArC) , 122.42 (ArC), 116.83(ArC), 99.44(CHO), 72.73(CHO), 72.09(CHO), 70.85(CHO), 68.29(CHO), 61.97(CH20), 52.18(CH3), 20.28-20.63 ( 4C, COOCH3 ) .

b) 1 , 3-fenylobis-O- ( 2 , 3 , 4 , 6-tetra-0-acetyl^-D- glucopyranoside (202)

The synthesis was performed according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.21g (llmmol) of resorcinol, 3.0 g (13mmol) of silver oxide and 15 ml of quinoline was used. During crystallization from methanol the product fell in the form of an oil, it was the purified by column chromatography (eluent CHC13 / MeOH 10: 1) . Glycoside fractions were collected, evaporated and recrystallized from methanol in a chamber with diethyl ether. As a result, crystallization from methanol- ether the precipitate fell out from solution in the form of crystals. 0.98g of pure product was obtained in the form of two anomers. Yield was 20%. Rfl = 0.58, Rf2 = 0.37 (CHC13 / MeOH 20: 1, silica gel, 12, combustion H2S04). Mp = 78-86 ⁰ C NMR analysis confirmed the presence of both glycoside anomers. 1HMR ( 300MHz , CDC13 ) , 7.21 ( t , 1H, ArH, 3JH-H=8.4Hz ) ,

6.70 (dd, 2H, ArH, 3JHH=8.4Hz, 4JH-H=2.4Hz), 5.33-5.09(m,8H, CHO) ,

4.28 (dd, 2H, CH20, 2JH-H=12.3Hz, 3JH-H= 5.4Hz) ,

4.15 (dd, 2H, CH20, 2JH-H=12.0Hz, 3JH-H=2.7Hz), 3.90- 3.84 (m, 2H, CHO) , 2.08(s, 6H, CH3CO) , 2.03 ( s, 6H, CH3CO) , 2.05 (s, 6H, CH3CO) , 2.03 ( s, 6H, CH3CO) . c) 5-methyl-l , 3-fenylobis-O-2 (, 3 , 4, 6-tetra-0-acetyl^-D- glucopyranoside ) and 3-hydroxy -5-methyl-3-phenyl-0- (2,3,4, 6-tetra-0-acetyl^-D-glucopyranoside ) (203)

The synthesis was performed according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.36 g (llmmol) orcine, 3.0 g (13mmol) of silver oxide and 15 ml of quinoline was used. During crystallization from ethanol the product came out in the form of an oil, which was purified by column chromatography (eluent CHC1 3 / MeOH 8: 1) . Glycoside fractions were collected, evaporated and recrystallized from ethanol in the chamber with diethyl ether. As a result the resulting oil was dried under vacuum. 1.10g of pure product obtained in the form of monoglucoside of orcine and diglucoside of orcine. Yield was 22%., Rf mono = 0.39 Rf di = 0.48 (CHC13 / MeOH 20: 1, silica gel, 12, combustion H2S04) . Mp = 84-89 ⁰ C NMR analysis confirmed the presence of both anomers the molar ratio 0.66 (di) : 0.34 (mono) .

1HMR ( 300MHz , CDC13 ) , 6.52-6.33 (m, 1H, ArH) , 5.33-5.05 (m, 8H, CHO) , 4.29-4.14 (m,4H,CH20), 3.76-3.69 (m, 2H, CHO) , 2.30 (s,3H,CH3ArbisGly) , 2.26 ( s, 3H, CH3ArmonoGly) ,

2.09 (s, 6H, CH3CO) , 2.05 ( s, 6H, CH3CO) , 2.04 ( s, 6H, CH3CO) ,

2.03 (s, 6H, CH3CO) . d) 4-0-acetyl-phenyl-2 , 3, 4, 6-tetra-0-acetyl^-D- glucopyranoside (204) .

The synthesis was performed according to the general procedure S-KK. To the reaction 8. Og (19 mmol) acetylated sugar bromide, 3.0 g (18mmol) monoacetylohyrdochinonu, 4.8g (21mmol) of silver oxide and 20 ml of quinoline was used. Recrystallization from methanol in a chamber with diethyl ether yield product as beige crystals with a weight of 2.8 g. Yield was 35%. Rf = 0.82 (CHC13 / MeOH 20: 1, silica gel, 12, combustion H2S04) . Mp = 146 ⁰ C. NMR analysis confirmed the presence of pure glycoside.

1HMR ( 300MHz , CDC13 ) , 7.00 ( s, 4H, ArH) , 5.29-5.26 (m, 2H, CHO) ,

5.16 (t, 1H, CHO) , 5.03 (d, 1H, CHO, 3JH-H=7.8Hz) , 4.29 (dd, 1H, CH20, 2JH-H=12.3Hz, 3JH-H=5. lHz ) ,

4.16 (dd, 1H, CH20, 2JH-H=12.3Hz, 3JH-H=2.4Hz ) , 3.87-

3.81 (m, 1H, CHO) , 2.29 ( s, 3H, CH3 ) , 2.08 ( s, 3H, CH3CO) ,

2.06 (s, 3H, CH3CO) , 2.05 ( s, 3H, CH3CO) , 2.03 ( s , 3H, CH3CO ) .

13CMR ( 300MHz , CDC13 ) , 197.75 (CH3COOAr ) , 170.60 ( CH3COO) , 170.26 (CH3COO) , 169.72 (CH3COO) , 169.43 ( CH3COO ) ,

169.32 (CH3COO) , 154.49(ArC), 146.28(ArC), 122.61(ArC),

118.04(ArC), 99.53(CHO), 72.74(CHO), 72.10(CHO), 71.18(CHO), 68.29(CH20), 61.96(CH20), 21.09 ( CH3COO ) , 20.74 (CH3COO) , 20.66 (CH3COO) .

e) 4-methoxycarbonyl-phenyl-O- ( 2 , 3 , 4 , 6 -tetra-O-acetyl-β-Ο- glucopyranoside ) (205)

The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.67g (llmmol) of methyl p-hydroxybenzoate and 3.0 g (13mmol) of silver oxide was used. 1.33g of white solid was otained, yield 25%. Rf = 0.47 (CHC13 / MeOH 20: 1, silica gel, 12, combustion H2S04) Mp = 141-143 ⁰ C. NMR analysis confirmed pure product.

1HNMR ( 300MHz, CDC13) , 7.93 (d, 2H, ArH, 3JH-H=8.7Hz),

7.09 (d, 2H, ArH, 3JH-H=8.7Hz), 5.70(d,lH, CHO, 3JH-H=7.8Hz) ,

5.40 (t, 1H, CHO, 3JH-H=9.6Hz) , 5.01-4.97 (m, 4H, CHO) , 4.30-

4.24 (m, 1H, CHO) ,4.21-4.15(m,lH, CH20) , 4.05 (dd, 1H, CH20, 2H- H=12.3Hz, 3JH-H= 2.1Hz), 3.81 ( s, 3H, CH3COO) , 2.00 ( s, 6H, CH3CO) , 1.99 (s, 3H, CH3CO) , 1.96(s,3H, CH3CO) .

f) 3-methoxycarbonyl-phenyl-O- ( 2 , 3 , 4 , 6 -tetra-O-acetyl-β-Ο- glucopyranoside ) (206)

The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.67g (llmmol) of m-hydroxybenzoate and 3.0 g (13mmol) of silver oxidewas used. 1.21g of white solid was obtained, yield 23%. Rf = 0.51 (CHC13 / MeOH 20: 1, silica gel, 12, visualisation H2S04) Mp = 111-112 ⁰ C. NMR analysis confirmed pure product.

1HNMR ( 300MHz, CDC13) , 7.76 (dd, 1H, ArH, 3JH-H=7.8Hz, 4JH-H=1.2Hz), 7.65 (d, 1H, ArH, 3JH-H=1.5Hz ) , 7.36 ( t , 1H, ArH, 3JH-H=8. lHz ) , 7.20- 7.16 (m, 1H, ArH) , 5.32-5.13 (m, 5H, CHO), 4.31-4, 15 (m, 2H, CH20) , 3.91 (s, 3H, CH3COO) , 2.08 ( s, 3H, CH3CO) , 2.06(s,3H, CH3CO) , 2.05 (s, 3H, CH3CO) , 2.04 (s, 3H, CH3CO) . g) 2-methoxycarbonyl-l , 3-fenylobis-O- ( 2 , 3 , 4 , 6 -tetra-O- acetyl^-D-glucopyranoside) (208) The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.85g (llmmol) of 2 , 6-dihydroxybenzoate and 3.0 g (13mmol) of silver oxide was used. The 1.51g of pale pink solid was obtained. The yield of this preparation process was 17%. Rf = 0.66 (CHC13 / MeOH 20: 1, silica gel, 12, visualisation H2S04) Mp = 128-132 ⁰ C. NMR analysis confirmed pure product . lHNMR(300MHz, DMSO) , 7.43 ( t , 1H, ArH, 3JH-H=8.4Hz ) , 6.86

(d, 2H, ArH, 3JH-H=8.4Hz ) , 5.51 (d, 2H, CHO, 3JH-H=8. lHz ) ,

5.38 (t, 2H, 3JH-H=9.6Hz) , 5.01-4.94 (m,4H,CHO),

4.264.06 (m, 2H, CHO; 4H, CH20) , 3.66 (s, 1H, CH3COO) , 2.01 (s, 6H, CH3CO) , 1.98(s,9H, CH3CO) , 1.98 ( s, 3H, CH3CO) ,

1.90 (s, 3H, CH3CO) , 1.94 (s, 3H, CH3CO) . h) 2-nitro-phenyl-O- ( 2 , 3 , 4 , 6 -tetra-O-acetyl-β-Ο- glucopyranoside ) (209)

The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.53g (llmmol) of o-nitrophenol and 3.0 g (13mmol) Ag20_was used Recrystallization from ethanol gave 3.46g dark yellow solid, yield was 67%. Rf = 0.79 (CHC13 / MeOH 10: 1, silica gel, 12, visualisation H2S04, Mp = 158-159 ⁰ C. NMR analysis confirmed pure product.

1HNMR ( 300MH, CDC13, ) , 7.79 (dd, 1H, ArH, 3JH-H=8. lHz, 4J=1.5Hz)

7.56-7.53 (ddd≡m, 1H, ArH, 4JH-H=1.8Hz ) , 7.35 (d, 1H, ArH, 3JH

H=8.4Hz), 7.21 (t, 1H, ArH, 3J=8. lHz) , 5.325.30 (m, 2H, CHO) , 5.22 5.12 (m, 2H, CHO) , 4.27-4.24 (m, 2H, CH20) , 3.90-3.84(m, 1H,CH0) 2.13 (s, 3H, CH3C0) , 2.09 (s, 3H, CH3CO) , 2.05 (s, 3H, CH3CO) ,

2.04 (s, 3H, CH3CO) . i) 4-nitro-phenyl-O- (2,3,4, 6-tetra-0-acetyl^-D- glucopyranoside )

The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.53g (llmmol) of p-nitrophenol and 3.0 g (14mmol) Ag20_was used. Recrystallization from ethanol gave 2.32g of a yellow solid, yield was 45%. Rf = 0.75 (CHC13 / MeOH 10: 1, silica gel, 12, visualisation H2S04, Mp = 145-147 ⁰ C. NMR analysis confirmed pure product. 1HNMR (300MHz, CDC13) , 8.21 (d, 2H, ArH, 3JH-H=9.3Hz ) ,

7.08 (d, 2H, ArH, 3JH-H=7.2Hz), 5.33-5.15(m,4H, CHO) ,

4.29 (dd, 1H, CH20, 2JH-H=12.3Hz, 3JH-H=5.4Hz), 4.19 (dd, 1H,

CH20, 3JH-H=14.4Hz, 2JH-H=2.4Hz), 3.96-3.91 (m, 1H, CHO) ,

2.07 (s, 3H, CH3CO) , 2.07 ( s, 3H, CH3CO) , 2.06 ( s, 3H, CH3CO) , 2.05 (s, 3H, CH3CO) . j) 3-nitrophenyl-O- (2,3,4, 6-tetra-0-acetyl^-D- glucopyranoside ) The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) acetylated sugar bromide, 1.53g (llmmol) of o-nitrophenol and 3.0 g (13mmol) Ag20_was used Recrystallization from ethanol gave 2.80g dark beige solid. Yield was 52%. Rf = 0.89 (CHC13 / MeOH 10: 1, silica gel, 12, visualisation H2S04), Mp = 171-172 ⁰ C. NMR analysis confirmed pure product.

1HNMR (300MHz, CDC13) , 7.82-7.80 (m, 2H, ArH) , 7.42 ( t , 1H, ArH, 3JH- H=8.1Hz), 7.337.25(m, 1H, ArH) ,5.23-5.10(m,4H, CHO) ,4.20- 4.11 (m, 2H, CH20) ,3.70-3.64(m,lH, CHO) , 2.06 (s, 3H, CH3CO) ,

2.05 ( s, 3H, CH3CO) , 2.04 ( s, 3H, CH3CO) , 2.02 ( s, 3H, CH3CO) . k) phenyl-O- (2,3,4, 6-tetra-0-acetyl-β-D-glucopyranoside )

(200)

The synthesis was carried out according to the general procedure S-KK. To the reaction 2.8 g (6.8mmol) of the bromide acetylated lactose, 0.59g (6.3 mmol) of phenol and 1.7g (7.5mmol) Ag20, Crystallization afforded 0.99g of a light brown solid. Yield was 33%. Rf = 0.29 (CHC13 / MeOH 10: 1, silica gel, 12, visualisation H2S04), Mp = 120-122 ⁰ C. NMR analysis confirmed pure product.

1HMR ( 300MHz , CDC13 ) , 7.30-7.24 (m, 2H, ArH) , 7.07-6.96 (m, 3H, ArH) , 5.28-5.25 (m, 2H, CHO) , 5.18-5.14 (m, 1H, CHO) , 5.08-5.05 (m, 1H, CHO) , 4.27 (dd, 1H, CH20, 2J=l 2.3Hz , 3J=5.4Hz ) , 4.15 (dd, 1H, CH20,

2J=12.3Hz, 3J=2.4Hz) , 3.86-3.81 (m, 1H, CHO) , 2.06 ( s , 3H , CH3CO ) , 2.04 (s, 3H, CH3CO) , 2.03 ( s, 3H, CH3CO) , 2.02 ( s, 3H, CH3CO) .

1) 2-methoxycarbonylphenyl-O- ( 2 , 3 , 4 , 6 -tetra-O-acetyl-β-Ο- galactopyranoside ) (221)

The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0 g (12 mmol) of the bromide acetylated galactose, 1.67g (llmmol) of methyl- hydroksobenzoesanu (methyl salicylate), and 3.0 g (13mmol) of silver oxide was used. Recrystallization from ethanol yielded 3.1g of a white solid. The reaction yield is 58%. Rf = 0.76 (CHC13 / MeOH 20: 1, silica gel, 12, visualisation H2S04), Mp = 157-159 ⁰ C. NMR analysis confirmed pure product.

1HNMR ( 300MHz, CDC13) , 7.74 (dd, 1H, ArH, 3JH-H=7.8Hz, 4JH-

H=l .8Hz, 4JH-H=1.5Hz) , 7.44 (m≡ddd, 1H, ArH, 4JH-H=1.8Hz ) , 7.14- 7.10 (m, 2H, ArH) , 5.59-5.53 (m,lH,CH20), 5.44 (d, 1H, CH20, 3JH- H=2.7Hz), 5.10-5.03 (m, 2H, CHO) , 4.24-4.06 (m, 3H, CHO) ,

3.84 (s, 3H, CH300C) , 2.17 ( s, 3H, CH3CO) , 2.06 (s,3H, CH3CO) ,

2.05 (s, 3H, CH3CO) , 2.00 (s, 3H, CH3CO) . m) 1 , 3-fenylobis-O- ( 2 , 3 , 4 , 6 -tetra-O-acetyl-β-Ο- galactopyranoside ) and 3-hydroksyfenyloO- (2,3, 4, 6-tetra-

O-acetyl -β-D-galactopyranoside (222)

he synthesis was performed according to the general procedure S-KK. To the reaction 10. Og (24mmol) of the bromide acetylated galactose, 2.42g (22mmol) of resorcinol, 6.1 g (26mmol) and 30ml Ag20 quinoline was used. Recrystallization from methanol product yielded an oil, which was purified by column chromatography (eluent CHC13 / MeOH 10: 1) . Glycoside fractions were collected, the solvent was evaporated and recrystallized from methanol in a chamber with diethyl ether, yielded an oil, which was dried in vacuo. 2.40g of pure product was obtained in the form of two anomers, 25%. Rfl = 0.56, Rf2 = 0.38 (CHC13 / MeOH 20: 1, silica gel, 12, visualisation H2S04), Mp = 82-92 ⁰ C. NMR analysis confirmed the presence of both glycoside anomers.

1HMR ( 300MHz, CDC13) , 7.21 (t, 1H, ArH, 3JH-H=8. lHz) ,

6.71 (dd, 2H, ArH, 3JHH=9 ,0Hz, 4JH-H=2.lHz), 6.65(d, 1H, ArH, 3JH- H=1.2Hz), 5.49-5.43 (m, 2H, CHO; 2H, CH20) , 5.12 (dd, 2H, CH20, 2JH- H=10.5Hz, 3JH-H=3.3Hz), 5.07(d,2H, CHO, 3JH-H=7.8Hz) ,

4.234.07(m,6H,CHO) , 2.18 ( s, 3H, CH3CO) , 2.08 ( s , 9H, CH3CO) ,

2.07 (s, 6H, CH3CO) , 2.05 ( s, 3H, CH3CO) , 2.02 ( s, 3H, CH3CO) .

13CNMR( 300MHz, CDC13) , 170.51 ( CH3COO ) , 170.27 ( CH3COO ) , 170.16 (CH3COO) , 169.40 ( CH3COO ) , 157.76(ArC), 130.19(ArC), 111.06(ArC), 106.28(ArC), 99.11(CHO), 71.08(CHO), 70.85(CHO), 68.57(CHO), 66.9KCHO), 61.35(CH20), 20.80 ( CH3COO ) ,

20.71 (CH3COO) , 20.64 ( CH3COO ) . n) 2-metoksykarbonylofenylo-0- (2, 3, 4, 6, 2', 3', 6' -hepta-O-acetyl-

4-0^-Dgalaktopiranozylo^-D-glukopiranozyd) ( 231 )

The synthesis was carried out according to the general procedure S-KK. To the reaction 8. Og (llmmol) of the bromide acetylated lactose, 1.52g (10 mmol) of methyl- hydroksobenzoesanu (methyl salicylate), and 2.8 g (12 mmol) of silver oxide was used . Recrystallization from ethanol gave 2.00 g of a white solid. The yield 25%. Rf = 0.38 (CHC13 / MeOH 20: 1, silica gel, 12, visualisation H2S04), Mp = 132-133 ⁰ C. NMR analysis confirmed pure product.

1HNMR ( 300MHz , CDC13 ) , 7.73 (dd, 1H, ArH, 3JH-H=7.8Hz , 4JH-

H=l .8Hz, 4JH-H=1.5Hz) , 7.42 (m≡ddd, 1H, ArH, 3JH-H=8. lHz, 4JH-

H=1.8Hz), 7.12-7.07 (m, 2H, ArH) , 5.34(d,lH, CHO, 3JH-H=3.3Hz ) , 5.27-5.24 (m, 2H, CHO) , 5.14-5.05 (m, 2H, CHO) , 4.95 (dd, 1H, CHO, 3JH- H=10.5Hz, 4JH-H=3.3Hz) , 4.52-4.49 (m, 2H, CH20) , 4.14- 4.08 (m, 1H, CHO; 2H, CH20) , 3.95-3.76 (m, 3H, CHO) ,

3.82 (s, 3H, COOCH3) , 2.14 ( s, 3H, CH3CO) , 2.06 ( s, 6H, CH3CO) ,

2.05 (s, 6H, CH3CO) , 2.04 ( s, 3H, CH3CO) , 1.95 ( s, 3H, CH3CO) . o) 1, 3-fenylobis-O- (2,3,4,6,2 ', 3', 6 ' -hepta-O-acetyl-4-O- -D-galactopyranosyl- β -D-glucopyranoside ) and 1 3- fenylobis-0 (2,3,4,6,2 ', 3', 6 ' -hepta-O-acetyl-4-Ο-β-

Dgalaktopiranozylo^-D-glucopyranoside) (232a) and 1, fenylobis-3-O- (2,3,4,6,2 ', 3', 6 ' -hepta-O-acetyl-4-Ο-β- D-galactopyranosyl-β -D-glucopyranoside) (232b) The synthesis was performed according to the general procedure

5-KK. To the reaction 8. Og (llmmol) of the bromide acetylated lactose, 1.10g (10 mmol) of resorcinol, 2.8 g (12 mmol) of silver oxide and 20 ml of quinoline was used. The product fell out from methanol as an oil which was purified by column chromatography (eluent CHC13 / MeOH 10: 1) . The oil resulting from evaporation of the glycoside-containing fractions was recrystallized from methanol in a chamber with diethyl ether. An oil, was decanted from the solution and dried under vacuum. 1.8g of pure product obtained in the form of two anomers. Yield was 24%. Rfl = 0.46, Rf2 = 0.33 (CHC13 / MeOH 20: 1, silica gel, 12, combustion H2S04) . The product obtained was separated by column chromatography (eluent CHC13 / MeOH 20: 1) giving two fractions, one containing the pure β anomer and a fraction containing a mixture of anomers a and β. After evaporation of both fractions was in form of an oil. The sample was crystallized from methanol in a chamber with ether solutions yielding an oil which was dried in vacuo. 0.35g rof product was obtained as a mixture of anomers a and β glycosides and 1.30g of pure β anomer glycoside. Mp / β = 138-155 ⁰ C, Mp β = 155-156 ⁰ C. NMR analysis confirmed the presence of glycosides obtained.

1HMR ( 300MHz , CDC13 ) , 7.20-7.06 (m, 1H, ArH) , 6.58-6.51 (m, 3H, ArH) , 5.68(d,lH, CH O, 3JH-H=4.5Hz ) , 5.36-4.95 (m, 12H, CHO) ,

4.63 (d, 1H, CHO, 3JH-H=7.8Hz) , 4.524.49 (m, 2H, CHO; 2H, CH20) , 4.34- 4.26 (2H, CHO; 6H, CH20) , 4.08-3.78 (m, 6H, CHO), 2.16 ( s, 3H, CH3CO) , 2.15 (s, 3H, CH3CO) , 2.13 ( s, 3H, CH3CO) , 2.12(s,3H, CH3CO) ,

2.09 (s, 6H, CH3CO) , 2.08 ( s, 3H, CH3CO) , 2.07 ( s, 3H, CH3CO) , 2.06(s,3H, CH3CO) , 2.05 ( s, 6H, CH3CO) , 2.05 ( s, 6H, CH3CO) ,

2.04 (s, 3H, CH3CO) , 1.97 ( s, 3H, CH3CO) , 1.97 ( s , 3H, CH3CO ) - anomer /β.

1HMR ( 300MHz , CDC13 ) , 7.15-7.09 (m, 1H, ArH) , 6.55-6.51 (m, 3H, ArH) , 5.55-4.74(m, 12H,CHO), 4.52-4.49 (m, 2H, CHO; 2H, CH20) , 4.15- 4.12 (m, 2H, CHO; 4H, CH20) , 3.893.77 (m, 6H, CHO) , 2.15 ( s, 3H, CH3CO) , 2.14 (s, 3H, CH3CO) , 2.13 ( s, 3H, CH3CO) , 2.12(s, 3H,CH3CO),

2.11 (s, 3H, CH3CO) , 2.09 ( s, 3H, CH3CO) , 2.07 ( s, 3H, CH3CO) ,

2.07(s,3H, CH3CO), 2.06 ( s, 3H, CH3CO) , 2.05 ( s, 6H, CH3CO) ,

2.04 (s, 3H, CH3CO) , 1.97(s,3H, CH3CO) - anomer β. p) 2-nitro-phenyl-2 , 3 , 4 , 6 , 2 ', 3', 6 ' -hepta-O-acetyl-4-0- ( β-D-galactopyranosyl-β -D-glucopyranoside ) (234) The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0g (6.8mmol) of the bromide acetylated lactose, 0.87 g (6.3 mmol) of o-nitrophenol and 1.7g (7.5mmol) Ag20 was used. Crystallization afforded 2.8 g of a yellow solid. Yield was 59%. Rf = 0.86 (CHC13 / MeOH 10: 1, silica gel, 12, visualisation H2S04), Mp = 206-208 ⁰ C. NMR analysis confirmed pure product.

1HNMR ( 300MHz, CDC13) , 7.79 (dd, 1H, ArH, 3JH-H=8. lHz, 4JH-H=1.8Hz), 7.52 (ddd≡t, 1H, 3JH-H=8.2Hz , 4JH-H=1.8Hz , 4JH-H=1.5Hz ) , 7.31- 7.26 (m, 1H, ArH) , 7.22-7.16 (ddd≡t , 1H, ArH, 3JH-H=7.8H, 4JH-

H=l .2Hz, 4JH-H=0.9Hz) , 5.37-4.99 (m, 6H, CHO, CH20) ,

4.604.53(m,2H, CH20) , 4.14-3.83 (m, 4H, CHO, 2H, CH20) ,

2.16 (s, 3H, CH3CO) , 2.13(s,3H, CH3CO) , 2.11 ( s, 3H, CH3CO) , 2.10 (s, 3H, CH3CO) , 2.05 ( s, 3H, CH3CO) , 2.04(s,3H, CH3CO) ,

1.98 (s, 3H, CH3CO) .

r) 4-nitro-phenyl-2, 3, 4, 6, 2 ', 3', 6 ' -hepta-O-acetyl-4-O- ( β-D-galactopyranosyl-β -D-glucopyranoside) (235)

The synthesis was carried out according to the general procedure S-KK. To the reaction 5.0g (6.8mmol) of the bromide acetylated lactose, 0.87 g (6.3 mmol) of p-nitrophenol and 1.7g (7.5mmol) Ag20 was used. Crystallization yielded 3.5g of yellow solid, yield was 74%. Rf = 0.73 (CHC13 / MeOH 10: 1, silica gel, 12, visualisation H2S04), Mp = 200-201 ⁰ C. NMR analysis confirmed pure product. 1HNMR (300MHz, CDC13) , 8.21 (d, 2H, ArH, 3JH-H=17.4Hz ) ,

7.06 (d, 2H, ArH, 3J=9.3Hz ) , 5.53-4.96 (m, 6H, CHO) , 4.54-

4.50 (m, 2H, CH20) , 4.16-4.15(m,2H, CH20) ,

4.164.10(m,2H, CHO; 2H, CH20) , 3.92-3.72(m,2H, CHO) ,

2.17 (s, 3H, CH3CO) , 2.09(s,3H, CH3CO) , 2.09 ( s, 3H, CH3CO) ,

2.08 (s, 3H, CH3CO) , 2.07 ( s, 3H, CH3CO) , 1.98(s,3H, CH3CO) ,

1.97(s,3H,CH3CO) .

s ) 2-metoksykarbonylofenylo-O- (2, 3, 4, 6, 2', 3', 6' -hepta-O-acetyl- 4-0^glukopiranozylo-a-D-glukopiranozyd) ( 241 )

The synthesis was carried out according to the general procedure S-KK. To the reaction 8. Og (llmmol) of the bromide acetylated maltose, 1.52g (10 mmol) o-hydroksobenzoesanu (methyl salicylate), and 2.8 g (12 mmol) of silver oxide was used. 1.50 g of white solid was obtained. The reaction yield was 19%. Rf = 0.51 (CHC13 / MeOH 20: 1, silica gel, 12, visualisation H2S04), Mp = 173-175 ⁰ C. NMR analysis confirmed pure product .

1HNMR (300MHz, CDC13) , 7.75 (dd, 1H, ArH, 3JH-H=8. lHz , 4JH-

H=2. lHz, 4JH-H=1.8Hz) , 7.49-7.43 (m, 1H, ArH) , 7.14-

7.11 (m, 2H, ArH) , 5.46 (d, 1H, CHO, 3JH-H=3.9Hz ) , 5.41- 5.29 (m, 2H, CHO) , 5.21-5.14 (m, 2H, CHO) , 5.09-5.02 (m, 1H, CHO) ,

4.86 (dd, 1H, CHO, 3JHH=10.5Hz, 3JH-H=4.2Hz), 4.51(dd,lH, CH20, 2JH- H=ll .7Hz, 3JH-H=3.0Hz, 3JH-H=2.7Hz), 4.30-4.23(m,2H, CH20) ,

4.14 (t, 1H, CHO) , 4.06 (dd, 1H, CH20, 2JH-H=12.3Hz, 3JH-H= 2.1Hz), 4.00-3.83 (m, 2H, CHO) , 3.83 ( s , 3H, COOCH3 ) , 2.10 ( s, 3H, CH3CO) , 2.09(s,3H, CH3CO), 2.05 ( s, 6H, CH3CO) , 2.04 ( s , 3H, CH3CO) ,

2.03 (s, 3H, CH3CO) , 2.01(s,3H, CH3CO) . t) 1,3-fenylobis-O- (2,3,4,6,2 ', 3', 6 ' -hepta-0-acetyl-4- 0- β-D-glucopyranosyl- -D-glucopyranoside ) and 1 3- fenylobis-O- (2,3,4,6,2 ', 3', 6 ' -hepta-O-acetyl-4-Ο-α

D-glucopyranosyl^-D-glucopyranoside) ( 242 )

The synthesis was performed according to the general procedure S-KK. To the reaction 8. Og (llmmol) of the bromide acetylated maltose, 1.10g (10 mmol) of resorcinol, 2.8 g (12 mmol) of silver oxide and 20 ml of quinoline was used. The product was in the form of an oil, which was purified by column chromatography (eluent CHC13 / MeOH 10: 1) . The oil resulting from evaporation of the glycoside-containing fractions was recrystallized from methanol in a chamber with diethyl ether. An oil, which was decanted from the solution and dried under vacuum. 1.90 g of pure product obtained in the form of two anomers, 25%. Rfl = 0.49, Rf2 = 0.35 (CHC13 / MeOH 20: 1, silica gel, 12, visualisation H2S04), Mp = 134-135 ⁰ C. NMR analysis confirmed the presence of both glycoside anomers.

1HMR ( 300MHz, CDC13) , 7.18 (t, 1H, ArH, 3JH-H=8. lHz) ,

6.66 (dd, 2H, ArH, 3JH-H=8. lHz, 4JH-H=2. lHz ) , 6.58 ( t , 1H, ArH, 3JH- H=2.1Hz), 5.36-4.94 (m, 6H, CHO) , 4.51-4.46(m, 2H,CHO), 4.16- 4.88 (m, 4H, CH20) , 3.91-3.69 (m, 2H, CHO; 4H, CH20) , 2.16(s,3H,

CH3CO), 2.08 (s, 3H, CH3CO) , 2.07 ( s, 6H, CH3CO) , 2.06 ( s, 3H, CH3CO) , 2.05(s,3H, CH3CO), 1.97 ( s, 3H, CH3CO) .

Egzample 4 - Deacylacja acetylowanych glikozydow polifenoli SD 2.5g (5.2mmol) of acetylated glycoside was dissolved in 15ml of dichloromethane and 40 ml of methanol was added and stirred with a magnetic stirrer for 20 minutes. Then the catalyst was added 0.36ml of 0.5M (0.18mmol) NaOMe and allowed to stand overnight, while stirring. When TLC analysis (CHC13 / MeOH 5: 1, silica gel, combustion H2S04) showed conversion of starting material to the flask 2 ml (approx. Twentyfold excess of Na +) resin Amberlite IR 120 (H +) was added in order to neutralize the reaction mixture and stirred 20 minutes, then the solution was drained from the resin and evaporated on a rotary evaporator .

On the other hand, when the TLC plate indicated still visible substrate, another portion of 0.5M NaOMe in methanol was added and product was treated as above The product was crystallized from methanol, yielding white or yellow fine sand like or white flaky like residue. In some cases, the solution fell dark brown and brown oil was obtained, which was evaporated to dryness under vacuum, thereby to give a pink "foamed" precipitate. NMR analysis confirmed the structure of the resulting glycosides.

a) 2-methoxycarbonylphenyl-0^-D-glucopyranoside (101)

The synthesis was performed according to the general procedure SD. To the reaction 3.0 g (6.4mmol) of methyl salicylate, acetylated glucoside (201) was used which was dissolved in 15 ml of dichloromethane; then 40 ml of methanol and 0.44ml (0.22mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 2.5ml resin Amberlite IR 120 (H +) was used. Crystallization yielded 2.09g of a white solid. Yield was 99%. Rf = 0.54 (CHC13 / MeOH 5: 1, silica gel, visualisation H2S04) . Mp = 105-107 ⁰ C. NMR analysis confirmed to obtain pure product .

1HNMR (300MHz, D20) , 7.67 (dd, 1H, ArH, 3JH-H=7.8Hz , 4JH- H=l .8Hz, 4JH-H=1.5Hz) , 7.51-7.45 (m, 1H, ArH) , 7.20-

7.17 (m, 1H, ArH) , 7.11-7.06 (m, 1H, ArH) , 5.03 (d, 1H, CHO, 3JH- H=7.2Hz), 3.82-3.81 (m, 1H, CHO) , 3.81 ( s, 3H, CH300C) , 3.66- 3.38 (m, 4H, CHO; 2H,CH20).

13C NMR (300MHz, D20) , 168.72 ( CH300C ) , 155.43 (ArCO) , 134.36(ArC), 131.07 (ArC) , 123.06(ArC), 120.80(ArC), 116.64(ArC), 100.63(CHO), 76.11(CHO), 75.53(CHO), 72.84(CHO), 69.21(CHO), 60.44(CH2O), 52.71 (CH300C) . b) 1, 3-fenylobis-0-β-D-glucopyranoside and 3-hydroxy-0^-D- glucopyranoside (102) The synthesis was performed according to the general procedure SD. To the reaction 1.5g (3.4mmol) acetylated glucoside resorcinol (202) was added, which was dissolved in 10 ml of dichloromethane ; then 20 mL of methanol and 7.0ml (3.5mmol) 0.5M NaOMe was added. For neutralization of sodium ions 40 ml of Amberlite IR 120 resin (H +) was used. In the attempts to crystallize pink oil was obtained from the solution which was dried under vacuum yielding 0.28g of the product. Yield was 38%. Rf = 0.23 (CHC13 / MeOH 5: 1, silica gel, visualisation H2S04) . Mp = 84-85 ⁰ C. NMR analysis confirmed the presence of product . c) 5-methyl-l , 3-fenylobis-O^-D-glucopyranoside and 3- hydroxy-5-methyl-3-phenyl-O^D-glucopyranoside ( 103 )

1HNMR (300MHz, D20) , 7.26-7.21 (m, lHdi, ArH) , 7.17-

7.16 (m, lHmono, ArH) , 6.78-6.75 (m, 2H, ArH) , 6.62-6.52 (m, 1H, ArH) , 5.01 (d, 2H, CHO, 3JH-H=7.5Hz ) , 3.84-3.30 (m, 10H, CHO, 4H, CH20) . The synthesis was performed according to the general procedure SD. To the reaction 1.5g (3.4mmol) acetylated glucoside of orcine (203) was added, which was dissolved in 10 ml of dichloromethane ; then 20 mL of methanol and 7.0ml (3.5mmol) 0.5M NaOMe was added. For neutralization of sodium ions 20ml of Amberlite IR 120 resin (H +) was used. Crystallization afforded 0.22 g of pink solid comprising a mixture of mono- and diglucosideof orcine. Yield was 38%. Rf = 0.22 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 88-89 ⁰ C. NMR analysis confirmed the receipt of the product.

1HNMR (300MHz, D20) , 6.61-6.34 (m, 3H, ArH) , 4.97 (d, 2H, CHO, 3JH— H=7.2Hz), 3.843.23 (m, 8H, CHO; 4H, CH20) , 2.20 ( 2 , 3H, CH3ArbisGly ) , 2.15 (2, 3H, CH3ArmonoGly) . d) 4-O-acetyl-phenyl-O- β-D-glucopyranose (104) The synthesis was performed according to the general procedure SD. To the reaction 2.5g (5.0mmol) of p-acetylated glucoside monoacetylofenolu (204) was added, which was dissolved in 15 ml of dichloromethane; then 40 ml of methanol and 0.5ml (0.25mmol) of 0.5M NaOMe was added.

Next day, TLC analysis still showed the presence of starting material. Therefore, the addition of another 0.5 ml of 0.5M NaOMe and 10 ml of methanol was done. After 2 hours, the spot of starting material on TLC disappeared. For neutralization of sodium ions 6ml resin Amberlite IR 120 (H

+) was used. Crystallization afforded 0.90 g of pure product as a white solid. Yield was 43%. Rf = 0.18 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 198-200 0 C. NMR analysis confirmed to obtain pure product.

1HNMR (300MHz, D20) , 6.94 (d, 2H, ArH, 3JH-H=7.8Hz ) ,

6.76 (d, 2H, ArH, 3JH-H=7.8Hz) , 4.87 (d, 1H, CHO, 3JH-H=7.2Hz ) , 3.83- 3.79 (m, 1H, CHO) , 3.61 (dd, 1H, CH20, 2JH—H=12.0Hz, 3JH—H=5.1Hz), 3.48-3.37 (m, 3H, CHO, 1H, CH20) .

e) 4-methoxycarbonyl-phenyl-0^-D-glucopyranoside (105)

The synthesis was performed according to the general procedure SD. To the reaction 1.33g (2.76mmol) of acetylated glucoside p-hydroxybenzoate (205) was added, which was dissolved in 10 ml of dichloromethane ; then 20 mL of methanol and 0.2ml (O.lOmmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.2ml resin Amberlite IR 120 (H +) was used. Crystallization afforded 0.31g of a white solid. Yield was 34%. Rf = 0.32 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 150 154 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR (300MHz, D20) , 7.92 (d, 2H, ArH, 3JH-H=8. lHz ) ,

7.01 (d, 2H, ArH, 3JH-H=8. lHz) , 5.12 (d, 1H, CHO, 3JH-H=7.2Hz ) , 3.86- 3.81 (m, 1H, CH20) , 3.80 ( s, 3H, CH3COO) , 3.69-3.41 (m, 4H, CHO, 1H, CH20) . f) 3-methoxycarbonyl-phenyl-0^-D-glucopyranoside (106)

The synthesis was performed according to the general procedure SD. To the reaction 1.21g (1.61mmol) of acetylated glucoside glucoside m-hydroxybenzoate (206) was added, which was dissolved in 15 ml of dichloromethane; then 20 mL of methanol and 0.12ml (0.06 mmol) of 0.5M NaOMe was added. For neutralization of sodium ions Amberlite resin IR 120 (H +) was used. Crystallization afforded 0.44 g of a white solid. Yield was 86%. Rf = 0. (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 148-152 ⁰ C. NMR analysis confirmed receipt of pure product. 1HNMR (300MHz, D20) , 7.63-7.58 (m, 2H, ArH) , 7.38 ( t , 1H, ArH, 3JH- H=8.1Hz), 7.287.24 (m, 1H, ArH) , 5.04 (d, 1H, CHO, 3JH-H=6.9Hz ) ,

3.86-3.81 (m, 1H, CH20) , 3.81(s,3H, CH3COO) , 3.67-

3.39 (m, 4H, CHO; 1H, CH20) .

g) 2-methoxycarbonyl-l , 3-fenylobis-0-β-D-glucopyranoside

(108)

The synthesis was performed according to the general procedure SD. To the reaction 1.51g (2.02mmol) of 2 , 6-bis-O-acetylated- -glucoside benzoate (208) was added, which was dissolved in 15 ml of dichloromethane ; then 20 ml of methanol and 0.1 ml (0.05mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 0.7ml resin Amberlite IR 120 (H +) was used. Crystallization afforded 0.38 g of a light brown solid. Yield was 38%. Rf = 0.26 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 141-148 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR (300MHz, D20) , 7.38 ( t , 1H, ArH, 3JH-H=8.4Hz ) ,

6.93 (d, 2H, ArH, 3JH-H=8. lHz) , 5.12 (d, 1H, CHO, 3JH-H=8.4Hz ) ,

5.02 (d, 2H, CH O, 3JH-H=7.5Hz), 4.36 (d, 2H, ΟΗβΟ, 3JHH=7.5Hz),

3.84 (s, 3H, CH3COO) , 3.88-3.23 (m, 8H, CHO, 4H, CH20) . h) 2-nitrophenyl-0^-D-glucopyranoside (109) The synthesis was performed according to the general procedure SD. To the reaction 1.9g (4.0mmol) acetylated glucoside o- nitrophenol (209) was added, which was dissolved in 10 ml of dichloromethane; then 20 mL of methanol and 0.28ml (0.14mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.6 ml of the resin Amberlite IR 120 (H +) was used. Crystallization afforded 0.81g of pure product as a pale yellow solid. Yield was 64%. Rf = 0.24 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 166 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR (D20, 300MHz) , 7.89 (d, 1H, 3JH-H=8. lHz ) ,

7.64 (t≡ddd, 1H, ArH, 3JH-H=8.0) , 7.38 (d, 1H, ArH, 3JH-H=8.5Hz),

7.22 (t≡ddd, lH,3JH-H=7.8Hz) , 5.19 (d, 1H, CHO, 3JHH=4.8Hz ) , 3.90- 3.30 (m, 4H, CHO; 2H, CH20) .

i) 4-nitrophenyl-0^-D-glucopyranoside (110)

The synthesis was performed according to the general procedure SD. To the reaction 1.9g (4.0mmol) of acetylated glucoside p- nitrophenol (210) was added, which was dissolved in 10 ml of dichloromethane ; then 20 mL of methanol and 0.28ml (0.14mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.6 ml of the resin Amberlite IR 120 (H +) was used. Crystallization afforded 1.0 g of pure product as a pale yellow solid. Yield was 79%. Rf = 0.25 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 166-168 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR (300MHz, D20) , 8.15 (d, 2H, ArH, 3JH-H=9.3Hz ) ,

7.12 (d, 2H, ArH, 3JH-H=9.3Hz) , 5.15 (d, 1H, CHO, JH-H=6.3Hz ) , 3.84- 3.40 (m, 4H, CHO; 2H, CH20) . j) 3-nitrophenyl-0^-D-glucopyranoside (111)

The synthesis was performed according to the general procedure SD. To the reaction 1.9g (4.0mmol) of acetylated glucoside m- nitrophenol (211) was added, which was dissolved in 10 ml of dichloromethane; then 20 mL of methanol and 0.28ml (0.14mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.6 ml of the resin Amberlite IR 120 (H +) was used. Crystallization afforded 0.89g of pure product as a pale yellow solid. Yield was 70%. Rf = 0.24 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 164-165 ⁰ C. NMR analysis confirmed receipt of pure product. 1HNMR(D20, 300MHz) , 7.88-7.82 (m, 2H, ArH) , 7.46 ( t , 1H, ArH,

3J=8.1Hz), 7.417.37 (m, 1H, ArH) , 5.10 (d, 1H, CHO, 3JH-H=7.8Hz ) , 3.84 (dd, 1H, CHO, 3JH-H=11.7Hz, 3JH-H= 1.5Hz), 3.67-

3.35(m,3H,CHO;2H,CH20) . k) phenyl-O^-D-glucopyranoside (100)

The synthesis was performed according to the general procedure SD. To the reaction 1.0 g (1.8mmol) acetylated glucoside phenol (200) was added, which was dissolved in 15 ml of dichloromethane ; then 30ml of methanol and 0.22ml (O.llmmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.6 ml of the resin Amberlite IR 120 (H +) was used. Crystallization afforded 0.58g of pure product as a light brown solid. Yield was 98%. Rf = 0.20 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 168-169 ⁰ C. NMR analysis confirmed receipt of pure product.

lHNMR(300MHz, D20) , 7.29-7.23 (m, 2H, ArH) , 7.03-6.99 (m, 3H, ArH) , 5.99 (d, 1H, CHO, 3JH-H=7.2Hz ) , 3.78 ( dd, 1H, CHO, 3JH-H=12.4Hz , 3JH- H=1.9Hz), 3.60 (dd, 1H, CH20, 2JH-H= 12.3Hz , 3JH-H=5.6Hz ) , 3.50- 3.35 (m, 4H, CHO) .

1) 2-methoxycarbonylphenyl-0^-D-galactopyranoside (121)

The synthesis was performed according to the general procedure SD. To the reaction 3. lg (6.4 mmol) of methyl salicylate, acetylated galactoside (221) was added, which was dissolved in 20ml of dichloromethane and 40ml of methanol; then 0.46ml (0.23mmol) 0.5M NaOMe was added. For neutralization of sodium ions 2.7ml resin Amberlite IR 120 (H +) was used. Crystallization afforded 1.73g of a white solid. Yield was 82%. Rf = 053 (CHC1 3 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 174-175 ⁰ C. NMR analysis confirmed receipt of pure product.

lHNMR(300MHz, D20) , 7.67 (d, 1H, ArH, 3JH-H=7.8Hz ) ,

7.47 (ddd≡m, 1H, ArH, 3JHH=1.5Hz, 3JH-H=0.9Hz), 7.18(d, 1H, ArH, 3JH- H=8.4Hz), 7.01-7.05 (m, 1H, ArH) , 4.96(d,lH, CHO, 3JH-H=7.5Hz ) , 3.86 (d, 1H, CHO) , 3.78 ( s, 3H, COOCH3 ) , 3.75-3.60 (m, 3H, CHO; 2H,

CH20) .

m) 1, 3-fenylobis-O^-D-galactopyranoside + S-hydroxy-0-β- Dgalaktopiranozyd (122)

The synthesis was performed according to the general procedure SD. To the reaction 1.5g (3.4mmol) resorcinol acetylated galactoside (222) was added, which was dissolved in 10 ml of dichloromethane; then 20 mL of methanol and 7.0ml (3.5mmol) 0.5M NaOMe was added. For neutralization of sodium ions 20ml of Amberlite IR 120 resin (H +) was used. Crystallization afforded 0.28g pale pink solid. The yield of this preparation process was 48%. Rf = 0.23 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 115-116 ⁰ C. NMR analysis confirmed the receipt the pure product as bisglikozydu monoglikozydu and resorcinol in a molar ratio of 0.76 (di -) 0.24 (mono-) . 1HNMR (300MHz, D20) , 7.27 (m, IHdi , ArH, 3JH-H=4.2Hz ) ,

7.18 (m, IHmono, ArH, 3JH-H= 3.9Hz), 6.81-6.79 (m, 3Hdi, ArH) , 6.58- 6.56 (m, 3Hmono, ArH) , 4.98 (d, 2H, CHO, 3JH-H= 3.9Hz),

4.96 (d, 2H, CHO, 3JH-H=3.6Hz) , 3.92 ( 2H, CHO, 3JH-H=1.5Hz ) , 3.86- 3.47(m,6H, CHO, 4H, CH20) .

n) 2-methoxycarbonylphenyl-O- ( β-D-galactopyranosyl-β-D- glucopyranoside ) (131)

The synthesis was performed according to the general procedure SD. To the reaction 2.00 g (2.5mmol) of methyl salicylate, acetylated lactoside (231) was added, which was dissolved in 10 ml of dichloromethane ; then 20 mL of methanol and 0.18ml (0.09mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.1ml resin Amberlite IR 120 (H +) was used. After evaporation of the solvent 0.93 g of product contaminated with starting material was obtained as determined by TLC analysis (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . The precipitate was redissolved in dichloromethane / methanol and after stirring for 20 minutes 0.1 ml of 0.5M NaOMe was added. The next day the solution was neutralized with 0.6mL Amberlite resin IR 120 (H +) . Crystallization afforded 0.99g of a white solid. The yield of this preparation process was 78%. Rf = 0.22 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 178-179 ⁰ C. NMR analysis confirmed receipt of pure product.

lHNMR(300MHz, D20) , 7.62 ( dd, 1H, ArH, 3JH-H=7.8Hz , 3JH-H=1.5Hz ) ,

7.51 (ddd≡t, 1H, ArH, 3JH-H=1.5Hz ) , 7.26 (d, 1H, ArH, 3JH-H=8.4Hz ) , 7.10 (dd≡t, 1H, ArH, 3JH-H=7.5Hz ) , (d, 1H, CHO, 3JH-H=7.8Hz) 4.23 (d, 1H, CHO, 3JH-H=6.3Hz), 3.78 (s, 3H, COOCH3) 3.783.12 (m, 8H, CHO; 4H, CH20) .

o) 1 , 3-fenylobis-O- ( β-Ο^Β ΐ ΒθΙ οργΓ Βηο ε γ Ι-β-Ο- glucopyranoside ) and 1 , 3-fenylobis-O- ( D- galactopyranosyl^-D-glucopyranoside) - anomer / β (132a )

The synthesis was performed according to the general procedure SD. To the reaction 0.3 g (0.28 mmol) acetylated lactoside resorcinol (232a) was dissolved in 2 ml of dichloromethane ; then 4 ml of methanol and 0.6mL (0.29mmol) of 0.5M NaOMe was added. For neutralization of the sodium ions of the resin 3.4 mL of Amberlite IR 120 (H +) was used. In the attempts to crystallize dark pink oil was obtained which was dried under vacuum. It was resulted in light brown sludge, which weighed 0.12g. Yield was 57%. Rf = 0.20 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 115-116 ⁰ C. NMR analysis confirmed the receipt of pure product in the form of two a and β anomers in a molar ratio of 0.34 (a) : 0.66 (β) .

1HNMR (300MHz, D20) , 7.12 ( t , 1H, ArH, 3JH-H=8.4Hz ) , 6.59-

6.51 (m, 3H, ArH) , 5.09(d, 1H, CHO, 3JH-H=3.9Hz ) , 5.00 (d, 1H, CHO, 3JH-H=8. lHz) , 4.53 (d, 1H, CHO, 3JH-H=8. lHz ) ,

4.33 (t≡dd, 2H, CHO, 3JH-H=8. lHz, 3JH-H=8.4Hz), 3.88- 3.42 (m, 16H, CHO; 8H, CH20) .

p) 1 , 3-fenylobis-O- ( β-D-galactopyranos^

glucopyranoside ) - β anomer (132b) The synthesis was performed according to the general procedure SD. To the reaction 1.35g (1.3mmol) acetylated lactoside resorcinol (232b) was dissolved in 8 ml of dichloromethane; then 16ml of methanol and 2.6ml (1.3mmol) 0.5M NaOMe was added. For neutralization of sodium ions 16ml of Amberlite IR 120 resin (H +) was used. The attempts to crystallize afforded a brown oil which, after drying under vacuum weights 0.58g (brown solid) . The yield of this preparation process was 58%. Rf = 0.23 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 155-156 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR (300MHz, D20) , 7.16 (t, IH, ArH, 3JH-H=8.6Hz) , 6.63-

6.54 (m, 3H, ArH) , 5.03 (d, IH, CHO, 3JH-H=7.8Hz), 4.38 (d, IH, CHO, 3JH-H=7.8Hz) , 3.92-3.44 (m, 18H, CHO; 8H, CH20) . r . 2-nitro-phenyl-O- ( β-D-galactopyranosyl-β-D- glucopyranoside ) (134)

The synthesis was performed according to the general procedure SD. To the reaction 1.6g (2.0mmol) acetylated lactoside o- nitrophenol (234) was added, which was dissolved in 10 ml of dichloromethane; then 20 mL of methanol and 0.14ml (0.07mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.6 ml of the resin Amberlite IR 120 (H +) was used. Crystallization afforded 0.48 g of pure product as a pale yellow solid. The process efficiency was 52%. Rf = 0.25 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 197-198 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR ( 300MHz, D20) , 7.90 (d, IH, ArH, 3JH-H=8. lHz) , 7.64 (t≡ddd, IH, ArH, 3JH-H= 8.0Hz), 7.39 (d, IH, ArH, 3JH-H=8.4Hz ) , 7.23 (t, IH, ArH, 3JH-H=7.8Hz) , 5.23 (d, IH, CHO, 3JHH=7.2Hz ) ,

4.44 (d, IH, CHO, 3JH-H=7.8Hz), 3.96-3.51(m,8H, CHO; 4H, CH20) . s) 4-nitro-phenyl-0- ( β-D-galactopyranosyl-β-D- glucopyranoside ) (135)

The synthesis was performed according to the general procedure SD. To the reaction 1.4g (1.8mmol) lactoside acetylated p- nitrophenol (235) was added, which was dissolved in 15 ml of dichloromethane ; then 30ml of methanol and 0.22ml (O.llmmol) of 0.5M NaOMe was added. For neutralization of sodium ions 1.6 ml of the resin Amberlite IR 120 (H +) was used. Crystallization afforded 0.47g of pure product as a pale yellow solid. The yield of this preparation process was 56%. Rf = 0.25 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 114-115 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR ( 300MHz, D20) , 8.22 (d, 2H, ArH, 3JH-H=9.8Hz),

7.21 (d, 2H, ArH, 3JH-H=9.2Hz), 5.25(d,lH, CHO, 3JH-H=7.5Hz),

4.44 (d, 1H, CHO, 3JH-H=7.5Hz) , 3.88-3.60 (m, 8H, CHO ; 4H,CH20). 13CNMR( 300MHz, D20) , 151.25(ArC), 150.39(ArC), 118.42(ArC), 116.22(ArC), 101.32(CHO), 76.04(CHO), 75.54(CHO), 72.97(CHO), 69.43 (CHO) , 60.54 (CHO) . t) 2-methoxycarbonylphenyl-O- ( β-D-glucopyranosyl- -D- glucopyranoside ) (141) The synthesis was performed according to the general procedure SD. To the reaction 1.50 g (1.9mmol) of methyl salicylate, acetylated maltoside was dissolved in 15 ml of dichloromethane; then 30ml of methanol and 0.14ml (0.068mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 0.8ml resin Amberlite IR 120 (H +) was used. After evaporation of the solvent gave 0.67 g of product contaminated with starting material as determined by TLC analysis (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . The precipitate was redissolved in dichloromethane / methanol and after stirring for 20 minutes 0.1 ml of 0.5M NaOMe was added. The next day the solution was neutralized with 0.6mL Amberlite resin IR 120 (H +) was used. Crystallization afforded 0.54g of a white solid. The yield of this preparation process was 56%. Rf = 0.23 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 207-208 ⁰ C. NMR analysis confirmed receipt of pure product.

1HNMR (300MHz, D20) , 7.62 (dd, 1H, ArH, 3JH-H=7.8Hz , 3JH-H=1.5Hz ) 7.547.48 (ddd≡m, 1H, ArH, 3JH-H=7.8Hz , 3JH-H=2.7Hz , 3JH-H=1.8Hz ) , 7.19 (d, 1H, ArH, 3JHH=13.5Hz), 7.10(t, 1H, ArH, 3JH-H=7.5Hz)

5.32 (d, 1H, CHO, 3JH-H=3.9Hz) , 5.04(d,lH, CHO, 3JH-H=7.8Hz )

3.79 (s, 3H, COOCH3) , 3.83-3.31 (m, 8H, CHO; 4H, CH20) .

u) 1 , 3-fenylobis-O- ( β-D-glucopyranosyl- -D-glucopyranoside ) (142)

The synthesis was performed according to the general procedure SD. To the reaction 1.7g (2.2mmol) of acetylated maltoside resorcinol was dissolved in 10 ml of dichloromethane; then 20ml of methanol 6. OmL (3.0 mmol) of 0.5M NaOMe was added. For neutralization of sodium ions 18ml of Amberlite IR 120 resin (H +) was used. In the attempts to crystallize afforded a brown oil which, after drying under vacuum, weighed 0.53g andi was semi solid. The yield of this preparation process was 56%. Rf = 0.23 (CHC13 / MeOH 5: 1 was added, silica gel, visualisation H2S04) . Mp = 102-104 ⁰ C. NMR analysis confirmed receipt of pure product. 1HNMR (300MHz, D20) , 7.17-7.11 (m, 1H, ArH) , 6.61-6.52 (m, 3H, ArH) , 5.02(d,lH, CHO, 3JH-H=7.8Hz ) , 4.36 (d, 1H, CHO, 3JH-H=7.5Hz ) , 3.90- 3.23 (m, 16H, CHO; 4H, CH20) .

Example 5 - Glycosylation alcohol derivatives - general procedure SF

A round bottom flask was provided with 3.6 g D-glucose (20 mmol) lOOmmoli, the corresponding alcohol and 100 mg H2S04 / silica gel. The mixture was stirred at 90-100 ⁰ C until dissolution of glucose (approx. 1 hr . ) . The reaction mixture was dissolved in methanol, filtered by silica gel and the solvent was evaporated. In order to get rid of the unreacted alcohol mixture is quenched with a suitable solvent (diethyl ether, acetone, or chloroform) added and stirred on a magnetic stirrer for approx. 1 hour. The insoluble precipitate contained product - glycoside corresponding alcohol and unreacted sugar as determined by TLC analysis (CH2C12 / MeOH 3: 1, silica gel, visualisation H2S04 and CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) . A mixture of glycoside and glucose were separated by column chromatography using the eluent dichloromethane / methanol 3: 1. The resulting product was crystallized from methanol in a chamber with diethyl ether. In each case, the alcohol glycoside fell on as very thick oil which was dried from solvent residues in vacuo. The obtained products were analyzed by NMR.

a) Sulfuric acid immobilized on silica gel To the round-bottomed flask 10 g silica gel (200-400 mesh) and 50 ml diethyl ether and 3 ml of concentrated sulfuric acid (VI) was added. After stirring for 5 minutes on a magnetic stirrer the solvent was evaporated to dryness and the silica gel immobilized thereon H2S04 was dried at 110 ⁰ C for 3 hours . b) butyl-0- / β-D-glucopyranoside (191)

The synthesis was performed according to the general procedure (SF) was performed with 9.1ml of alcohol. The reaction was carried out at 90 ⁰ C. The resulting pellet containing the glycoside and the unreacted sugar alcohol was washed withss diethyl ether yielding 1.42g of the product The yield was 30%. Rf = 0.62 (CH2C12 / MeOH 3: 1 was added, silica gel, visualisation H2S04) Rf = 0.77 (CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) NMR analysis confirmed receipt of pure product.

1HNMR ( 300MHz, D20) , 4.78 (d, IH, CH O, 3JH-H=3.6Hz) ,

4.32 (d, IH, ΟΗβΟ, 3JH-H=7.8Hz) , 3.82-

3.09 (m, 4H, CHO; 2H, CH20; IH, CH20) , 1.50-1.45 (m, 2H, CH2 ) , 1.28- 1.23(m,2H, CH2), 0.80-0.75 (m, 2H, CH3 ) . c) 3-hydroxy-butyl-O-a / β-D-glucopyranoside (192)

The synthesis was performed according to the general procedure (SF) adding 9.0ml of alcohol. The reaction was carried out at 95 ⁰ C. The resulting pellet containing the glycoside and the unreacted sugar alcohol washed with acetone yielding 1.08g of the product The yield was 20%. Rf = 0.40 (CH2C12 / MeOH 3: 1 was added, silica gel, visualisation H2S04) Rf = 0.68 (CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) NMR analysis confirmed receipt of pure product.

1HNMR ( 300MHz, D20) , 4.78 (d, IH, CHaO, 3JH-H=3.6Hz) ,

4.33(d,lH,CH30,3JH-H=6.6Hz) , 3.88-3.10 (m, 4H,CHO; 2H,CH20; 2H,CH2; 1H, CH) , 1.69-1.64 (m, 2H, CH2 ) , 1.07(d, 3H, CH3 , 3JH- H=6.0Hz) . d) 3-hydroxy-but-2-yl-0- / β-D-glucopyranoside (193) The synthesis was performed according to the general procedure (SF) with 9.0ml of alcohol. The reaction was carried out at 90 ⁰ C. The resulting pellet containing the glycoside and the unreacted sugar alcohol washed with diethyl ether yielding 0.89g of the product The yield was 16%. Rf = 0.45 (CH2C12 / MeOH 3: 1 was added, silica gel, visualisation H2S04) Rf = 0.51 (CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) The yield was 16%. NMR analysis confirmed receipt of pure product. lHNMR(300MHz, D20) , 5.12 (d, 1H, CH O, 3JH-H=3.6Hz ) ,

4.33 (d, 1H, ΟΗβΟ, 3JH-H=8. lHz) , 3.86- 3.10 (m, 4H, CHO; 2H, CH20; 2H,CH), 1.13-1.01(m,6H, CH3) . e) 3-hydroxy-O- / β-D-glucopyranoside (194) The synthesis was performed according to the general procedure (SF) with 7.2ml of alcohol. The reaction was carried out at 80 ⁰ C. The resulting pellet containing the glycoside and the unreacted sugar alcohol washed with acetone yielding 1.52g of the product The yield was 34%. Rf = 0.26 (CH2C12 / MeOH 3: 1 was added, silica gel, visualisation H2S04) Rf = 0.88 (CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) NMR analysis confirmed receipt of pure product. lHNMR(300MHz, D20) , 4.79 (d, lH,CH O,3JH-H=3.3Hz) ,

4.34(d, lH,CH30,3JH-H=7.8Hz) , 3.91-

3.11 (m, 4H, CHO; 2H, CH20; 4H, CH2) , 1.78-1.65(m,2H, CH2) . f) 5 , 6 -dihydroksyheksylo-O- / β-D-glucopyranoside (195) The synthesis was performed according to the general procedure (SF) with 12.1ml alcohol. The reaction was carried out at 90 ⁰ C. The resulting pellet containing the glycoside and the unreacted sugar alcohol washed with diethyl ether yielding 0.83g of the product The yield was 14%. Rf = 0.62 (CH2C12 / MeOH 3: 1 was added, silica gel, visualisation H2S04) Rf = 0.62 (CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) NMR analysis confirmed receipt of pure product .

lHNMR(300MHz, D20) , 4.81-4.78 (m, 1H, CH O) , 4.36-4.33 (m, 1H, ΟΗβΟ) , 3.82-3.13 (m, 4H, CHO; 2H, CH20; 2H, CH) , 1.58-1.25 (m,8H,CH2). g) 2-methoxyethyl-O-a / β-D-glucopyranoside (196)

The synthesis was performed according to the general procedure (SF) was added, with 7.8ml of alcohol. The reaction was carried out at 90 ⁰ C. The resulting pellet containing the glycoside and the unreacted sugar alcohol was washed with diethyl ether yielding 1.48g of the product The yield was 31%. Rf = 0.42 (CH2C12 / MeOH 3: 1, silica gel, visualisation H2S04) Rf = 0.61 (CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) NMR analysis confirmed receipt of pure product .

lHNMR(300MHz, D20) , 4.82 (d, 1H, CHaO, 3JH-H=3.6Hz ) ,

4.36 (d, 1H, ΟΗβΟ, 3JH-H=8. lHz) , 3.97-

3.55 (m, 3H, CHO; 2H, CH20; 2H, CH2) , 3.45-3.32(m,2H, CH2) ,

3.29 (s, 1H,CH3) , 3.283.15(m, 1H, CHO). h) 2- (2-ethoxy) etoksyenanolo-O-a / β-D-glucopyranoside (197) The synthesis was performed according to the general procedure (SF), with 13.6ml alcohol. The reaction was carried out at 90 ⁰ C. The resulting pellet containing the glycoside and the unreacted sugar alcohol was washed with diethyl ether yielding 2.08g of the product The yield was 35%. Rf = 0.38 (CH2C12 / MeOH 3: 1, silica gel, visualisation H2S04) Rf = 0.58 (CH3COCH3 / BuOH / H20 5: 4: 1, silica gel, 12, visualisation H2S04) NMR analysis confirmed receipt of pure product.

lHNMR(300MHz, D20) , 4.80 (d, 1H, CH O, 3JH-H=3.6Hz ) ,

4.36 (d, 1H, ΟΗβΟ, 3JH-H=7.8Hz) , 3.95-3.12

(m, 4H, CHO; 2H, CH20; 10H, CH2) , 3.05 (t, 1H, CH3 , 3JH-H=6.9Hz) .

Example 6 - Physicochemical characterisation

In order to test the ability of synthesized compounds to form complexes with calcium ions there were performed physico- chemical analysis, such as NMR titration, conductometric analysis and flame photometry. In addition, microscopic observations were performed to illustrate the effect of glycosides to dissolute the calcium oxalate crystals - model of kidney stones. The final step was to test the representative compounds for their effect on dissolution of postoperative kidney stones.

Evaluation of the ability of the compounds to solubilize the calcium oxalate has been made according to the method described in the work: I. Das, SK Gupta, V.N. Pandey, SA Ansari, Journal of Crystal Growth 2004, 267, 654-661, I. Das, SK Gupta, SA Ansari, VN Pandey, RP Rastogi, Journal of Crystal Growth 2005, 273, 546-554, with modifications. The authors of the cited works measured calcium content in the sample containing the compound and the addition of solid calcium oxalate (sample analyzed-A) and another sample containing only a solid calcium oxalate (reference sample-W) . The measurement was done after establishing a state of equilibrium, tracking the impact of compound added to the dissolution process of calcium oxalate in consecutive days. The calcium content was determined as % of emissions in the method of flame photometry. The ability to solubilize calcium salt is expressed as the difference between the calcium content of the analyzed sample and the reference sample (as indicated in Table ΔΙ) . Positive ΔΙ extract is a measure of the compound to dissolve the insoluble calcium salts. The greater the value ΔΙ the greater ability of the tested compound to dissolve the calcium salt. The studies include the amount of calcium-ion content (P) introduced into the system with the tested compound .

ΔΙ = A - W - P

Table 1 shows the numbers of the compounds and the corresponding emissions (ΔΙ) measured by flame photometry.

Table 1

Results of flame photometry

Oznaczenie

Nazwa zwi¾zku ΔΙ referencyj ne

2-metoksykarbonylofenylo-0- β-D-

101 30 glukopiranozyd

1 , 3-fenylobis-O^-D-glukopiranozyd i 3-

102 200 hydroksyfenylo-0- β-D-glukopiranozyd

5-metylo-l, 3-fenylobis-0-β-Ο-

103 glukopiranozyd i 3-hydroksy-5-metylo-3- 156 fenylo-0- β-D-glukopiranozyd

104 4-0-acetylofenylo-0- β-D-glukopiranoza 45

105 4-metoksykarbonylofenylo-0- β-D- 11 glukopiranozyd

3-metoksykarbonylofenylo-0- β-D-

106 19 glukopiranozyd

2-metoksykarbonylo-l , 3-fenylobis-0-β-Ο-

108 86 glukopiranozyd

109 2-nitrofenylo-O- β-D-glukopiranozyd 61

110 4-nitrofenylo-O- β-D-glukopiranozyd 13

111 3-nitrofenylo-O- β-D-glukopiranozyd 30

100 fenylo-O- β-D-glukopiranozyd 12

2-metoksykarbonylofenylo-O- β-D-

121 69 galaktopiranozyd

1 , 3-fenylobis-O^-D-galaktopiranozyd + 3-

122 257 hydroksyfenylo-O- β-D-galaktopiranozyd

2-metoksykarbonylofenylo-O- ( β-D-

131 106 galaktopiranozylo- β-D-glukopiranozyd)

1, 3-fenylobis-O- ( β-D-galaktopiranozylo-β- D-glukopiranozyd) i 1 , 3-fenylobis-O- (a-D-

132a 92 galaktopiranozylo- β-D-glukopiranozyd) - anomer /β

1, 3-fenylobis-O- ( β-D-galaktopiranozylo-β-

132b 133

D-glukopiranozyd) - anomer β

2-nitrofenylo-O- ( β -D-galaktopiranozylo-

134 82 β-D-glukopiranozyd)

4-nitrofenylo-O- ( β-D-galaktopiranozylo-β-

135 34

D-glukopiranozyd)

2-metoksykarbonylofenylo-O- ( β-D-

141 35 glukopiranozylo- -D-glukopiranozyd)

1, 3-fenylobis-O- ( β-D-glukopiranozylo-a-D-

142 130 glukopiranozyd)

Additionally, FIG. 1 shows the structure of the glycosidic derivatives of polyphenols contained in Table 1. Example 7 - Biological studies

The study was performed at the Institute of Immunology and Experimental Therapy -Polish Academy of Science in Wroclaw in accordance with ISO 10993-5. The study used two cell lines:

- L929 - fibroblast cell line obtained from the subcutaneous fat C3H mice (ATCC CCL 1) .

- A549 epithelial carcinoma cell line of lung carcinoma (ATCC CCL 185) .

Cell cultures of A549 and L929 were treated with test compounds at concentrations of 0-1500 g / ml and incubated for 72 hours under suitable conditions (37 ⁰ C, 5% C02) . For three consecutive days, the cytotoxic effect of each concentration was determined for both lines.

Cell viability and proliferation was determined by the method of Mosmann [184] modified by Berg [185] involving the reaction of reduction of the yellow dye MTT (3- [ 4 , 5-dimethylotiazol-2- yl] -2 , 5-difenylotetrazol ) to the dark blue formazan by mitochondrial dehydrogenase of viable cells.

At the 24-hour cultures of the cell lines A549 and L929 test substance were applied in concentration range of 0-1500 g / ml and the samples were incubated under suitable conditions (37 ⁰ C, 5% CO 2 for 3 days) . Morphological changes of cells and their viability was determined under an inverted microscope. In both cell types toxic effect of preparations were not observed .

Only glycosides 105 (at a concentration of 1500 and 750 ug / ml) and 106 (at a concentration of 1500, 750 and 375 ug / ml) show toxicity to cell line L929 and A549. Toxic effect is presented on the pictures in Fig. 2 - cells are shrunk round with granular cytoplasm, a large number of cells is dead. Other preparations do not have a toxic effect on the cells that line the L929 and A549. MTT colorimetric method for determining the viability and proliferation of L929 cells under the influence of the test compounds confirmed the results obtained by biological method - glycosides 105 (at a concentration of 1500 and 750 ug / ml) and 106 (at a concentration of 1500, 750 and 375 ug / ml) show toxic effect to cells L929 line. Other compounds are not toxic for the cell line L929 (FIG. 2) .