RU2576658C2 - Triphenylphosphonium salts of lupane and ursane triterpenoids, method of production and use for treating bilharzia - Google Patents

Abstract

FIELD: chemistry.

SUBSTANCE: invention employs triphenylphosphonium salts lupane and ursane triterpenoids of formula 1-11 as agents with schistosomicide activity, novel compounds 8-11, and describes a method for production thereof

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EFFECT: use of invention widens range of medicinal agents derived from natural products for treatment of schistosomiasis - a dangerous parasitic disease of humans and animals.

7 cl, 1 dwg, 3 tbl, 11 ex

Description

The invention relates to the field of bioorganic and medical chemistry, in particular to the creation of new drugs for the treatment of parasitic (invasive) diseases of humans and animals, and specifically relates to triphenylphosphonium derivatives of lupanic and ursanic triterpenoids of formula 1-11, which exhibit schistosomicidal activity against schistosomules (newly transformed schistosoma) and adult Schistosoma Manson worms (Schistosoma mansoni) in low micromolar concentrations

Triphenylphosphonium salts of formula 1-11 are derivatives of betulin of formula 12, betulinic acid of formula 13, 20,29-dihydrobetulinic acid of formula 14, 20,29-dihydro-3-epi-betulinic acid of formula 15 and ursolic acid of formula 16.

Using the invention will expand the range of drugs derived from natural products for the treatment of schistosomiasis - a dangerous parasitic disease in humans and animals.

Schistosomiasis is a chronic parasitic disease caused by blood flukes (trematodes) of the genus Schistosoma. Of the existing parasitic infections, schistosomiasis is in second place after malaria in terms of its negative impact on public health and the economy. Today, treatment is required for at least 243 million people and 779 million people are at risk. Schistosomiasis transmission has been documented in almost all countries located in subtropical and tropical climates [WHO Newsletter 2013, No. 215]. Chronic schistosomiasis can trigger other serious infections, such as HIV infection and malaria. Schistosome S. Haematobium is considered as a high risk group for the development of bladder cancer. Intestinal schistosomiasis (Schistosoma mansoni) can cause colon cancer and hepatocellular carcinoma. In Africa, more than 200 thousand people die from schistosomiasis annually. There are two main forms of schistosomiasis - intestinal and urogenital, caused by the five main types of obligate parasite. Intestinal schistosomiasis is caused by blood flukes of the species Schistosoma msnsoni, Schistosoma japonicum, Schistosoma mekongi, Schistosoma intercalatum, which live in the veins of the large intestine and abdominal cavity. Urogenital schistosomiasis is caused by Schistosoma haematobium trematodes. Among the five types of human schistosomiasis listed, S. mansoni is the most common disease. All types of schistosomes parasitizing in humans have a rather complex life cycle. At the invasive stage, free-living cercariae, after exiting a freshwater mollusk, penetrate intact human or animal skin and turn into schistosomules. After 2-3 days, the schistosomes migrate through the blood vessels into the lungs, and then into the portal vein, where mature schistosomes are copulated. Then, depending on the type of schistosome, they migrate into the venules of the mesentery, bladder or ureter and begin to lay eggs. For S. mansini, the periods of migration and maturation of parasites are 4-5 weeks.

The list of modern medicines recommended by WHO for the treatment of schistosomiasis includes two drugs - praziquantel and oxamnihin. The most widely used preparation is praziquantel (2- (cyclohexylcarbonyl) -1,2,3,6,7,11b-hexahydro-4H-pyrazino [2,1-a] isoquinolin-4-one) [R. Abdul-Ghani, N. loutfy, A. El Sahn, A. Hassan. Current chemotherapy arsenal for schistosomiasis mansoni: alternatives and challenges. Parasitol Res., 2009, 104, 955-965; D. Ndjonka, L.N. Rapado, A.M. Silber, E. Liebau, C. Wrenger. Natural Products as a source for treating neglected parasitic diseases. Int. Mol Sci, 2013, 14, 3395-3439]

The drug was received in the mid-1970s. It is a safe and inexpensive medicine that is effective in treating all forms of schistosomiasis. Juvenile parasites (ages 7-21 days) are less sensitive to the drug than adults. Despite prolonged use of praziquantel, the mechanism of its multifunctional effect on schistosomes has not been fully studied. It is assumed that praziquantel interacts with specific parasite proteins, which modulate the transmembrane potential of the cell, are responsible for the activity of voltage-gated calcium channels, and control the value of the Ca ²⁺ current. Under its influence, an excess amount of Ca ²⁺ ions enters the cell, which leads to disturbances in the specific functions of the cell. In addition, under the influence of praziquantel, the worm membrane is broken and access to antigens that cause the host's immune response opens [K. Abdul-Ghani, N. Loutfy, A. El Sahn, A. Hassan. Current chemotherapy arsenal for schistosomiasis masnsoni: alternatives and challenges. Parasitol Res., 2009, 104, 955-965].

The second drug used in clinical practice as a reserve drug is oxamniquin, synthetic tetrahydroquinoline (1,2,3,4-tetrahydro-2 - {[((1-methylethyl) amino] methyl} -7-nitro-6-quinolinomethanol)

This compound is only effective in S. mansoni infection. In Egypt and South America, persistent varieties of the pathogen are found, which requires high doses of the drug. The process of obtaining oxamnichine includes five synthetic stages using enzymatic catalysis in the final stage, which makes the production of the drug difficult and expensive.

Metrifonate (chlorophos, vermicide-Bayer 2349) is an O, O-dimethyl-2,2,2-trichloro-1-hydroxyethyl phosphate and was used as an insecticide to protect plants from agricultural pests before use in medical practice

It exhibits low efficiency in intestinal schistosomiasis, suitable only for the treatment of urogenital schistosomiasis S. haematobium. The use of metrifonate leads to serious side effects in connection with the suppression of acetylcholinesterase activity and the accumulation of acetylcholine in nerve synapses. This drug is currently undergoing clinical trials in Ghana, Zambia and other African countries.

Antimony potassium tartrate and other trivalent antimony derivatives have been used to treat schistosomiasis since 1918 for fifty years

Currently, these extremely toxic medications that cause serious side effects (nausea and vomiting, diarrhea, joint pain, hepatitis) are not recommended for use.

Thus, the set of modern drugs used in clinical practice for the treatment of schistosomiasis is very limited. The weak interest of pharmaceutical companies in the creation of new drugs against schistosomiasis and other "neglected" tropical diseases is explained by the relatively low price of products in this pharmaceutical field.

Schistosomiasis is a serious, widespread disease in the world, with a constantly expanding geography of endemic areas due to developed tourism and mass labor migration. To date, praziquantel is almost the only effective and non-toxic drug suitable for mass use. Since this drug has been used in chemotherapy for schistosomiasis since the mid-seventies, there is a serious problem of parasite resistance to it. In this regard, studies on the development and implementation of new effective schistosomicidal drug agents are extremely important.

It seems promising to search for new cheap drugs based on available biologically active substances of plant origin and their directed chemical modification, taking into account the specific characteristics of the parasite’s organism [D. Ndjonka, L.N. Rapado, S.M. Silber, E. Liebau, C. Wrenger. Natural products as a source for treating neglected parasitic diseases. Int. J. Mol. Sci., 2013, 14, 3395-3439]. In vitro laboratory experiments studied the extracts of essential oils of various medicinal plants, vegetable oil extracts and individual biologically active compounds belonging to the class of coumarins, flavanoids, derivatives of anthracene and sesquiterpene lactones. Antiparasitic compounds were identified in a number of these natural substances, but their antishistosomal activity was weakly expressed. Thus, when adults act on curcumin, hydroxychrysophanol, quercetin, and vernodalin on S. S. mansoni adults, high parasite mortality was observed at concentrations in the range of 40–100 μg / ml. A more significant schistosomicidal effect was exerted by the well-known antimalarial drugs, such as the natural compound artemisinin (tricyclic sesquiterpene with a peroxide bridge), its water-soluble semisynthetic analogue artesunate, and oil-soluble artemisinin methyl ester - artemether [D. Ndjonka, L.N. Rapado, S.M. Silber, E. Liebau, C. Wrenger. Natural products as a source for treating neglected parasitic diseases. Int. J. Mol. Sci., 2013, 14, 3395-3439; J. Utzinger, X. Shuhua, E.K. N'Goran, R. Bergquist, M. Tanner. The potential of artemethen for the control of schistosomiasis. International Journal for Parasitology 2001, 31, 1549-1562]

The antishistosomal activity of artemisinin, artesunate and artemether was discovered by Chinese scientists in the 1980s while studying the schistosome of S. japonicum. The most chemically stable and promising drug in this group of compounds, artemether, in animal trials showed high activity against S. mansoni, S. japonicum and S. haematobium. Juvenile schistosomes (1-3 weeks old) were significantly more sensitive to artemether compared to praziquantel. In the combined chemotherapy of schistosomiasis, the simultaneous use of artemether and praziquantel has shown encouraging results in laboratory studies and clinical trials [J. Utzinger, J. Keiser, X. Shuhua, M. Tanner, B.H. Singer. Combination chemotherapy of schistosomiasis in laboratory studies and clinucal trials. Antimicrobial agents and chemotherapy, 2003, 1487-1495]. However, the use of artemisinin to control schistosomiasis in malaria endemic areas can lead to an irreversible increase in the resistance of malaria plasmodium to this drug.

The schistosomicidal activity of available pentacyclic triterpenoids - betulin, betulinic and ursolic acids and their analogues has not been previously studied. However, these plant metabolites are known for a wide range of biological properties: anti-inflammatory, antiviral (anti-HIV), antitumor, antimolar, anti-leishmaniasis and trypanocidal [G.A. Tolstikov, O.B. Flechter, E.E. Schulz, L.A. Baltina, A.G. Tolstikov. Chemistry for Sustainable Development, 2005, 13, 1; P. Yogeeswari, D. Sriram, Current Medicinal Chemistry, 2005, 12, 657; G. NS da Silva, N. RG Marial, DC Schuch, LN Cruz, M. S de Moraes, M. Nakabashi, C. Graebin, G. Gosmann, C. RS Garcia, S. CB Gnoatto. Two series of new semisynthetic triterpene derivatives: differences in anti-malarial activity, cytotoxicity and mechanism of action. Malaria Journal, 2013, 12, 1573-1582; S. Alakurtti, T. Heiska, A. Kiriazis, N. Sacerdoti-Sierra, CL Jaffe, J. Yli-Kauhaluoma. Synthesis and anti-leishmanial activity of heterocyclic betulin derivatives. Bioorganic & Medicinal Chemistry. 2010, 18, 1573-1582; DB Domínguez-Carmonaa, F. Escalante-Erosaa, K. García-Sosaa, G. Ruiz-Pinellb, D. Gutierrez-Yapub, MJ Chan-Bacabc, A. Giménez-Turbab, LM Pena-Rodrígueza. Antiprotozoal activity of betulinic acid derivatives. Phytomedicine. 2010, 17, 379-382]. In vitro laboratory experiments, betulinic and ursolic acids showed moderate antimalarial activity against Plasmodium falciparum 3D7 with an IC _{50 of} 13.9 μM (for betulinic acid) and 36 μM (for ursolic acid) [G. NS da Silva, N. RG Marial, DC Schuch, LN Cruz, M. S de Moraes, M. Nakabashi, C. Graebin, G. Gosmann, C. RS Garcia, S. CB Gnoatto. Two series of new semisynthetic triterpene derivatives: differences in anti-malarial activity, cytotoxicity and mechanism of action. Malaria Journal, 2013, 12, 1573-1582]. As a result of active research on the synthesis and study of the antimalarial activity of a large group of semisynthetic analogues of betulinic and ursolic acids, new potential antimolar agents were identified that showed in vitro activity at low micromolar or nanomolar concentrations [HL Ziegler, H. Franzyk, M. Sairafianpour, M. Tabatabai, , MD Tehrani, K. Bagherzadeh, H. Hagerstrand, D. Stark, JW Jaroszewski. Erythrocyte membrane modifying agents and the inhibition of Plasmodium falciparum growth: structure-activity relationships for betulinic acid analigues. Bioorganic & Medicinal Chemistry. 2004. 12. 119-127; SCB Gnoatto, S. Susplugas, LD Vechia, TB Ferreira, A. Dassonville-Klimpt, KR Zimmer, C. Demailly, S. Da Nascimento, J. Guillon, P. Grellier, H. Verli, G. Gosmann, P. Sonnet . Pharmacomodulation on the 3-acetylursolic acid skeleton: design, synthesis, and biological evaluation of nivel N- {3- [4- (3-aminopropyl) piperazinyl] propyl} -3-O-acetylursolamide derivatives as antimalarial agents. Bioorganic & Medicinal Chemistry. 2008.16.1671-182; AM Innocente, GNS Silva, LN Cruz, MS Moraes, M. Nakabashi, P. Sonnet, G. Gosmann, CRS Garcia, SCB Gnoatto. Synthesis and antiplasmodialactivity of betulinic acid and ursolic acid analog. Molecules 2012, 17, 12003-12014]. Thus, hybrid compounds containing several pharmacophore groups (fragments of ursolic or betulinic acid and substituted piperazine) showed high antimolar activity against plasmodium “Plasmodium falciparum 3D7” (IC ₅₀ = 175-220 nM), showed low toxicity to kidney embryonic cells human HEK293T (IC ₅₀ = 4 μM) and acceptable therapeutic selectivity index (SI 18-23) [SCB Gnoatto, S. Susplugas, LD Vechia, TB Ferreira, A. Dassonville-Klimpt, KR Zimmer, C. Demailly, S. Da Nascimento, J. Guillon, P. Grellier, H. Verli, G. Gosmann, P. Sonnet. Pharmacomodulation on the 3-acetylursolic acid skeleton: design, synthesis, and biological evaluation of nivel N- {3- [4- (3-aminopropyl) piperazinyl] propyl} -3-O-acetylursolamide derivatives as antimalarial agents. Bioorganic & Medicinal Chemistry. 2008.16.1671-182; AM Innocente, GNS Silva, LN Cruz, MS Moraes, M. Nakabashi, P. Sonnet, G. Gosmann, CRS Garcia, SCB Gnoatto. Synthesis and antiplasmodialactivity of betulinic acid and ursolic acid analog. Molecules 2012, 17, 12003-12014].

In recent years, lipophilic triphenylphosphonium cations as molecules for studying the functions of mitochondria and as a means of delivering antioxidants, antimicrobial and antitumor agents to mitochondria have been attracting increased attention of researchers [J.S. Modica-Napolitano, J.R. Aprille, Adv Drug Delivery Rev, 2001, 49, 63; F. Wang, M.A. Ogasawara, P. Huang, Mol Aspect of Medicine, 2010, 31, 75; L. Biasutto, L-F. Dong, M. Zoratti, J. Neuzil, Mitochondrion, 2010, 10, 670; B. Bachowska, J. Kazmierczak-Baranska, M. Cieslak, B. Nawrot, D. Szczesna, J. Skalik, P. Balczewski, Chemistryopen, 2012, 1, 33]. A large group of mono- and bisphosphonium salts of benzophenones was studied in vitro against resistant lines of African trypanosomes and protozoan parasites of Leishmania [J.R. Luque-Ortega, P. Reuther, L. Rivas, C. Dardonville. New benzophenone-derived bisphosphonium salt as leishmanicidal leads targeting mitochondria through of respiratory complex II. J. Med. Chem. 2010 53, 1788-1798; A. Taladriz, A. Healy, E.J.F. Pérez, V.H. Garcia, C.R. Martнnez, A.A.M. Alkhaldi, A.A. Eze, M. Kaiser, H.P. de Koning, A. Chana, C. Dardonville. Synthesis and structure - activity analysis of new phosphonium salts with potent activity against african trypanosomes. J. Med. Chem. 2012, 55, 2606-2622]

Phosphonium derivatives inhibited the proliferation of parasites at low micromolar concentrations with little toxicity to human cells. Under the influence of these compounds directed to complex II of the respiratory chain of parasites, the cytoplasmic ATP rapidly decreased and the electrochemical mitochondrial potential decreased.

Phosphonium lipocations based on phthalimides and 1,4-naphthoquinones showed in vitro high antiprotozoal activity against Plasmodium falciparum and Trypanosoma cruzi [T.E. Long, Xiao Lu, M. Galizzi, R. Docampo, J. Gut, P.J. Rosenthal. Phosphonium lipocations as antiparasitic agents. Bioorganic & Medicinal Chemistry Letters. 2012.22. 2976-2979].

Recently, we have obtained new triphenylphosphonium derivatives of lupane triterpenoids. The new compounds showed a significantly higher cytotoxic effect in vitro than betulinic acid on the cancer cell lines of Ehrlich carcinoma and mast cell P-815 [A.Yu. Spivak, D.A. Nedopyokina, E.R. Shakurova, R.R. Halitova, R.R. Gubaidullin, V.N. Odinokov, U.M. Dzhemilev, Yu.P. Velsky, N.V. Velskaya, S.A. Stankevich, E.V. Short, V.A. Khazanov. Synthesis of lupane triterpenoids with triphenylphosphonium fragments and study of their antitumor activity. News of the Academy of Sciences. Ser. chemical. 2013, 62, No. 1, 189-199].

It is known that the outer cover of trematodes is a tegument. It consists of a layer of cells that merge with each other and form the total mass of protoplasm (syncytium) [J.J. Van Hellemond, K. Retra, J. F.H.M. Brouwers, B.W.M. van Balkom, M. Yazdanbakhsh, C. B. Shoemaker, A.G.M. Tielens. Functions of the tegument of schistosomes: Clues from the proteome and lipidome. International Journal for Parasitology. 2006, 36, 691-699]. The outer layer of the tegument is a nuclear-free cytoplasm containing a large number of mitochondria. The biological target of betulinic acid are mitochondria. Betulinic acid promotes the accumulation of active oxygen-containing particles in the mitochondria, which leads to serious dysfunctions in these organelles [S. Fulda, G. Kroemer, Drug Discovery Today, 2009, 14, 885]. We suggested that covalent binding of the lipophilic triphenylphosphonium cation to betulinic acid or other derivatives of the lupano group will enhance their orientation towards mitochondria located on the surface of the tegument. Destruction of the surface of the tegument will contribute to the death of parasites, since the tegument performs vital functions for the body of schistosomes, such as absorption of nutrients and secretion, protection against the humoral and cellular immunity of the host organism.

In connection with the stated facts, the object of the present invention is to obtain triphenylphosphonium derivatives of lupanic and ursanic triterpenoids, which can be used as new schistosomicidal drugs. The problem is solved by obtaining triphenylphosphonium salts 1-11 by the interaction of triphenylphosphine with C (2) - or C (30) -bromo (iodine) -substituted C (3) - and C (28) -ether derivatives of betulin 12, betulinic acid 13, dihydrobetulin acids 14, dihydro-3-epibetulinic acid 15 and ursolic acid 16, as well as the in vitro detection of the schistosomicidal effect of the obtained phosphonium salts on schistosomules, and adult Schistosoma mansoni worms.

The claimed compounds 1-6, which are conjugates of the C (3) - and C (28) ester derivatives of triterpenic acids 13-15 with the triphenylphosphonium cation linked to the triterpenoid molecule at the C (2) position by the propyl bridge, were synthesized as follows: dihydrobetulonates 17 18 and transformed into 3β-dihydrobetulinic acid epimers 19 and 20 by stereoselective reduction of 3-keto group under the action of NaBH _4, CeCl modified ₃ · 7H ₂ O. Digidrobetulonat 17 also subjected to reduction in the α-epimer 21 under the action of tri-sec-butylborohydride whether Ia (L-Selectride) (chemical reaction I).

Chemical reactions I

Reagents and conditions: (1a) NaBH ₄ , CeCl ₃ · 7H ₂ O, MeOH-THF, -30 ° C → 20 ° C, Ar; (1b) L-selectride, THF, -78 ° C → 20 ° C, Ar; (2) Ac ₂ O or AcCl; (3) BH ₃ · THF or BH ₃ Me ₂ S, THF 20 ° C, Ar; (4) I ₂ , PPh ₃ , imidazole, THF, 0 ° C; (5) MsCl, Py, CH ₂ Cl ₂ , DMAP, 20 ° C; (6) LiBr, (CH ₃ ) ₂ CO, boiling, Ar; (7) Pd / C, -Et ₂ O; (8) PPh ₃ , CH ₃ C ₆ H ₅ or CH ₃ CN, boiling, Ar.

When salts 1, 2, 5, and 6 were obtained, alcohols 19-21 were transformed into 3-O-acetylated dihydrobetulonates 22-24 by the standard procedure under the action of Ac ₂ O in pyridine or in AcCl in THF in the presence of catalytic amounts of pyridine and 4-dimethylaminopyridine ( DMAP). Then, compounds 19, 21-24 were subjected to successive transformations, including hydroboration of a double bond in an allyl substituent using the complex BH ₃ · THF or BH ₃ · Me ₂ S followed by oxidation of the organoboron compound to alcohols 25-29 under the influence of 30% H ₂ O ₂ and 10% aqueous NaOH solution. This was followed by the preparation of iodides 30-32 by nucleophilic substitution of the primary hydroxyl group in alcohols 25, 26, 29 using crystalline iodine in the presence of imidazole and triphenylphosphine. Bromides 36-38 were obtained from alcohols 27-29 in two stages through intermediate mesylates 33-35 by reacting alcohols with methanesulfonyl chloride (MsCl) in the presence of pyridine and DMAP, and then nucleophilic substitution of the mesyl group by LiBr in acetone.

In the preparation of salt 5, benzyldihydrobetulinate 38 was transformed into the corresponding acid 39 by hydrogenolysis of the ether bond using 10% Pd / C as a catalyst. The final stage of obtaining the desired triphenylphosphonium salts 1-6 was carried out by reacting halides 30-32, 36, 37, 39 with an excess of triphenylphosphine while boiling in toluene or in acetonitrile for 16-48 hours in an argon atmosphere.

In the preparation of allyl salt 8, betulin 12 was acetylated into ether 40 according to the standard procedure under the action of Ac ₂ O in pyridine or AcCl in THF in the presence of catalytic amounts of pyridine and 4-dimethylaminopyridine (DMAP) or ether 40 was obtained directly from birch birch by boiling it in acetic acid acid for 12 hours according to the method of [Kuznetsova S.A., Kuznetsov B.N., Redkina E.S., Sokolenko V.A., Skvortsova G.P. RF patent №2324700]. Compound 40 was then transformed into allylic bromide 41 under the influence of N-bromosuccinimide (NBS) in CCl ₄ according to the method of [NV Uzenkova, NI Petrenko, MM Shakirov, EES hul`ts, GA Tolstikov. Chemistry of Natural Compounds. 2005, 41, 692-700]. The bromide 41 was boiled with an excess of triphenylphosphine in CH ₃ CN in an argon atmosphere (chemical reactions II). When CH ₃ CN was replaced with toluene, the reaction proceeded with skeletal rearrangement of the lupane backbone to give a mixture of the compound.

Chemical Reactions II

Reagents and conditions: a. Ac ₂ O, Py, DMAP or AcCl, THF, Py, DMAP, 20 ° C; b. NBS, CCl ₄ ; c. PPh ₃ , CH ₃ CN, boiling, Ar; d. CH ₂ N ₂ , Et ₂ O.

Allyl salts 7 and 9 were obtained from betulinic acid 13 through intermediate 3β-OAc-betulinic acid 42, its methyl ester 42a and intermediate bromides 43 and 43a, as described for salt 8.

The synthesis of compound 11 was carried out according to the method described by us for the preparation of salts 1-6. Ursolic acid 16 was transformed into methylursonoate 44 under the action of CH ₂ N ₂ in Et ₂ O, oxidized to methyl ursonic acid 45 under the action of PCC (pyridinium chlorochromate) in CH ₂ Cl ₂ and transformed into 2β-allyl substituted ether 46 by reaction with KN (SiMe ₃ ) ₂ , Et ₃ B and allyl bromide in dimethoxyethane. Compound 46 was transformed into 3β-OH-epimer 47 by stereoselective reduction of the 3-keto group under the influence of NaBH ₄ -CeCl ₃ · 7H ₂ O. Compound 47 was acetylated into ether 48, as described for ethers 40 and 42. Then, ether 48 was subjected to successive transformations including the hydroboration of a double bond in an allyl substituent using a complex of BH ₃ · THF or BH ₃ · Me2S followed by oxidation of the organoboron compound to alcohol 49 under the influence of 30% H ₂ O ₂ and 10% aqueous NaOH. This was followed by obtaining iodide 50 by nucleophilic substitution of the primary hydroxyl group in alcohol 49 using crystalline iodine in the presence of imidazole and triphenylphosphine and obtaining the target salt 11 by reacting iodide 50 with an excess of triphenylphosphine when boiling in toluene or acetonitrile under argon atmosphere (chemical reactions III).

Chemical Reactions III

Reagents and conditions: a. CH ₂ N ₂ , Et ₂ O, 20 ° C; b. PCC, CH ₂ Cl ₂ , 20 ° C; c. KN (SiMe ₃ ) ₂ , BEt ₃ , C3H ₅ Br, dimethoxyethane, 20 ° C, Ar; d. NaBH ₄ , CeCl ₃ · 7H ₂ O, MeOH-THF, 30 ° C → 20 ° C, Ar; e. Ac ₂ O, Py, DMAP or AcCl, THF, Py, DMAP, 20 ° C; f. BH ₃ · THF or BH3 · Me ₂ S, THF 20 ° C, Ar; g. I ₂ , PPh ₃ , imidazole, THF, 0 ° C; h. PPh ₃ , CH ₃ C ₆ H ₅ , boiling, Ar.

Triphenylphosphonium salt 10 was obtained from 2β-allylbetulonate 51, which was transformed into ether 51b in two stages. Compound 51b was hydroborated under the action of the BH ₃ · THF complex in THF to obtain diol 52. Diol 52 was iodinated with crystalline iodine in the presence of imidazole and triphenylphosphine. The resulting diiodide 53 was reacted with an excess of triphenylphosphine by boiling in toluene or acetonitrile under argon atmosphere to give compound 10 (chemical reactions IV).

Chemical Reactions IV

The starting compounds in the synthesis of phosphonium salts 1-11 were prepared as follows. Ursolic acid was obtained from chokeberry according to the method of [L.P. Kozlova, E.V. Malykhin, SM. Obut, S.P. Ponov, O.P. Sheremet, RF patent 2329048]. Betulin was isolated from birch bark according to the method [G.A. Tolstikov, M.I., Goryaev, Kh.O. Kim, R.A. Come on, Zhurn. Adj Chemistry, 1967, 40, 920]. Betulonic acid 54 was obtained by oxidation of betulin 12 under the influence of Jones reagent in acetone. Betulinic acid 13 in the form of a mixture of 3α- and 3β-epimers (3α: 3β = 5: 95) was obtained by reduction of betulonic acid NaBH ₄ in a solvent mixture of CHCl ₃ : MeOH = 1: 1. Crystallization of a mixture of products from MeOH gave an individual 3β-epimer [DSHL Kim, Z. Chen, T. Nguyen, JM Pezzuto, S. Qiu, Z.-Z.Lu. Synthetic Communications. 1997, 27, 1607-1612] (chemical reactions V). Hydrogenation of betulononic acid 54 to dihydrobetulonic acid 55 was carried out according to the method of [L. Pohjala, S. Alakurtti, T. Ahola, J. Yli-Kauhaluoma, P. Tammela. Journal of Natural Products. 2009, 72, 1917-1926].

Chemical reactions V

Reagents and conditions: (1) CrO ₃ , H ₂ SO ₄ , CH ₃ COCH ₃ , 0 ° C; (2) NaBH ₄ , CHCl ₃ -MeOH (50/50), 20 ° C; (3) H ₂ , 10% Pd / C, MeOH-THF (50/50), 20 ° C; (4) CH ₂ N ₂ , Et ₂ O, 20 ° C; BnCl, DMF, K ₂ CO ₃ , 55 ° C; (5) KN (SiMe ₃ ) ₂ , BEt ₃ , C ₃ H ₅ Br, dimethoxyethane, 20 ° C, Ar; (6) Ac ₂ O or AcCl, 20 ° C.

The protection of 28-COOH and 3-OH groups in betulinic, betulonic and dihydrobetulonic acids was carried out according to standard methods. Methyl betulonate 51 and dihydrobetulonates 17 and 18 were obtained by the interaction of esters of betulonic and dihydrobetulonic acids 56-58 with KN (SiMe ₃ ) ₂ , Et ₃ B and allyl bromide in dimethoxyethane according to our previously published method [A.Yu. Spivak, E.R. Shakurova, D.A. Nedopekina, R.R. Halitova, L.M. Khalilov, V.N. Odinokov, Yu.P. Belsky, A.N. Ivanova, N.V. Belskaya, M.G. Danilets, A.A. Ligacheva. News of the Academy of Sciences. - Ser. chemical. 2011, No. 4, 681-688].

The schistosomicidal effect of synthesized salts 1-11 (11 compounds) was studied in vitro against schistosomules and adults S. mansoni and was classified in the following categories with respect to control: 3 - “all are alive”; 0 - "everyone is killed." For comparison, betulin 12, betulinic acid 13, ursolic acid 16, and the commercially available triphenylphosphonium salt of bromobutanoic acid 59 were tested 59. The compounds were initially tested against S. mansoni schistosomula at concentrations of 12.5 μg / ml and 6.25 μg / ml. The action of all phosphonium salts in these concentrations led to the complete death of schistosomules after 72 hours. Betulinic and ursolic acids showed moderate activity under these conditions, betulin and synthetic salt 59 did not show activity. Triphenylphosphonium salts 1-6 and 11 were additionally tested against schistosomules in the following six concentrations: 0.39 μg / ml, 0.78 μg / ml, 1.56 μg / ml, 3.13 μg / ml, 6.25 μg / ml, 12.5 μg / ml. The highest schistosomicidal activity against schistosomules in the series of compounds 1-6 and 11 was demonstrated by phosphonium salts 1 (IC ₅₀ 0.56 μg / ml) and 2 (IC ₅₀ 0.52 μg / ml) (Table 1).

The selected five phosphonium salts 1, 2, 7, 8 and 10, as well as betulinic and ursolic acids, were tested against adult S. mansoni at a concentration of 25 μg / ml. The action of all phosphonium salts led to the complete death of parasites after 24 hours and 72 hours. Betulinic and ursolic acids did not show activity under these conditions. Next, five phosphonium salts were tested at the following concentrations: 0.31 μg / ml, 0.93 μg / ml, 2.78 μg / ml, 8.33 μg / ml. Antiparasitic activity of all salts began to appear at a concentration of 2.78 μg / ml, then dose-dependently increased, and at a concentration of 8.33 μg / ml after 24 hours or 72 hours, 100% death of adult schistosomes was observed (Table 2; Appendix. Fig. 1).

The highest antiparasitic activity against adult S. mansoni was shown by allylic phosphonium salts of 7 (IC ₅₀ 0.76 μg / ml) and 8 (IC ₅₀ 0.64 μg / ml).

The advantages of the proposed method:

1. For the first time, new ionic derivatives of the triterpenoids of the lupanic and ursanic groups — triphenylphosphonium salts of betulin betulinic and ursolic acids — were investigated as potential schistosomicidal drug agents. In in vitro trials, triphenylphosphonium derivatives of triterpenoids showed high schistosomicidal activity against schistosomules and adult Manson schistosome worms.

2. Among the tested triphenylphosphonium derivatives of lupane and ursane triterpenoids, allyl phosphonium salts 7 and 8 with IC ₅₀ values of 0.76 and 0.64 μg / ml, respectively, showed the highest anti-schistosomal activity. These compounds are obtained in stages 2-4 from an available plant metabolite of betulin using simple and cheap reagents. Simple preparative synthesis allows you to get compounds 7 and 8 in any quantities necessary for scientific research and medical testing.

3. The high antishistosomal activity of triphenylphosphonium salts 1-11 in comparison with the activity of betulin, betulinic, ursolic acid and commercially available salt 59 indicates the mutual synergistic effect of the triphenylphosphonium cation and fragments of betulinic or ursolic acid on the antishistosomal activity of hybrid molecules 1-11.

The specific structure of the skin-muscle membrane of trematodes with a high content of mitochondrial tegument on the surface makes the outer cover of parasites a vulnerable place for potential mitochondria-targeted drugs, which include triphenylphosphonium derivatives of betulinic and ursolic acids. Betulinic acid and triphenylphosphonium cation act as prooxidants [J.S. Modica-Napolitano, J.R. Aprille, Adv Drug Delivery Rev, 2001, 49, 63; S. Fulda, G. Kroemer, Drug Discovery Today, 2009, 14, 885]. The presence of these compounds in mitochondria induces the formation of aggressive oxygen-containing radicals that destroy the membranes of mitochondria. Schistosomes exist in an aerobic environment and under conditions of oxidative stress provoked by the presence in the body of phosphonium salts of betulinic and ursolic acids, schistosomes should give an adequate antioxidant response. At the same time, the reserve of the redox system of schistosomes is much weaker than the potential of the human biological redox system [Hsin-Hung Huang, C. Rigouin, D.L. Williams. The redox biology of schistosome parasites and applications for drug development. Curr Pharm Des. 2012. 18. 3595-3611]. So, schistosomes lack an endogenous antioxidant catalase, and schistosomal glutathione peroxidase proteins exhibit low activity with respect to hydrogen peroxide. Schistosomules exhibit low expression of cytosolic and peptide-containing superoxide dismutase.

In connection with the stated facts, triphenylphosphonium derivatives of the triterpenoids of the lupanic and ursanovian groups are of interest as new potential antishistosomal drug agents, which, apparently, play the role of prooxidants that destroy the mitochondria of the parasite tegument.

The invention is illustrated by examples.

Example 1

(с 0.35, CHCl₃). ИК- и УФ-спектры, спектры ЯМР 1H и ¹³C соединения 17 приведены в работе [А.Ю. Спивак, Э.Р. Шакурова, Д.А. Недопекина, P.P. Халитова, Л.М. Халилов, В.Н. Одиноков, Ю.П. Вельский, А.Н. Иванова, Н.В. Вельская, М.Г. Данилец, А.А. Лигачева. Известия АН, Сер. химическая, 2011, №4, 680-687].Methyl 3β-acetoxy-2β- (3-triphenylphosphoniopropyl) -20.29-dihydrobetulinate bromide (1): To a solution of compound 56 (0.42 mmol) in DME (2.50 ml) at room temperature under argon atmosphere with stirring KN (SiMe ₃ ) ₂ (1 M solution in THF) (0.55 ml, 0.55 mmol) was added. After 15 minutes, Et ₃ B (1 M solution in THF) (0.55 ml, 0.55 mmol) was added to the solution and stirred for 1 h. Then a solution of allyl bromide in 1 ml DME (0.55-0.84) was added. mmol, mainly 0.84). The reaction mixture was stirred for a control time (3-4 h, mainly 3 h, TLC control), neutralized with 3 M HCl, diluted with water (1.5 ml) and extracted with EtOAc. The combined extracts were dried with MgSO ₄ . The residue was evaporated and chromatographed on a SiO ₂ column (eluent: hexane: EtOAc, 30: 1 → 1: 1), methyl 2β-allyl-3-oxolup-20,29-dihydrobetulonate was obtained (17). Yield 77%, white crystals, mp = 84-86 ° C (EtOH), $${[\alpha ]}_D^{\mathrm{twenty}}-36.0$$

(c 0.35, CHCl ₃ ). IR and UV spectra, 1H and ¹³ C NMR spectra of compound 17 are given in [A.Yu. Spivak, E.R. Shakurova, D.A. Nedopekina, PP Halitova, L.M. Khalilov, V.N. Odinokov, Yu.P. Velsky, A.N. Ivanova, N.V. Velskaya, M.G. Danilets, A.A. Ligacheva. Izvestia AN, Ser. Chemical, 2011, No. 4, 680-687].

Compound 17 (0.3 mmol) was dissolved in a solvent mixture of THF-methanol (1: 2, 7.5 ml) in an Ar atmosphere and cooled to -30 ° C. To this solution was added dropwise a solution of CeCl ₃ · 7H ₂ O (0.15 g, 0.4 mmol) in a solvent mixture of THF-methanol (1: 1, 1 ml). Then NaBH ₄ (0.023 g, 0.6 mmol) was added in small portions over 5 minutes, the temperature was raised to room temperature and stirred at this temperature for 2 hours (the reaction progress was monitored by TLC). After completion, a 5% HCl solution was added and the mixture was extracted with ethyl acetate (25 ml). The extract was washed with saturated aqueous NaHCO ₃ , water, dried (MgSO ₄ ) and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent hexane: EtOAc, 30: 1 → 1: 1), and methyl 2β-allyl-3β-hydroxy-20,29-dihydrobetulinate was obtained (19). Yield 74%. Physical characteristics, IR and UV spectra, 1H and ¹³ C NMR spectra of compound 19, as well as salts 1-6 and intermediates 20-39 described below (Scheme 1) are given in [A.Yu. Spivak, D.A. Nedopekina, E.R. Shakurova, PP Halitova, PP Gubaidullin, V.N. Odinokov, U.M. Dzhemilev, Yu.P. Velsky, N.V. Velskaya, S.A. Stankevich, E.V. Short, V.A. Khazanov. Izvestia AN, Ser. Chemical, 2013, No. 1, 189-199].

To a solution of compound 19 (0.47 mmol) in 4 ml of dry pyridine, cooled to 0 ° C with stirring, was added acetic anhydride (0.07 ml, 0.69 mmol), 4-dimethylaminopyridine (DMAP) (0.029 g, 0.24 mmol) and stirred at room a temperature of 10-16 hours, mainly 16 hours (TLC control). Then a 10% HCl solution was added and the mixture was extracted with ethyl acetate (4 × 15 ml). The extract was washed with brine, dried (MgSO ₄ ) and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent - hexane: EtOAc, 15: 1). Methyl 2β-allyl-3β-acetoxy-20,29-dihydrobetulinate was obtained (22). Yield 95%.

The BH ₃ · THF complex (1 M solution in THF) (0.86 ml, 0.86 mmol) was added in an argon atmosphere at room temperature to a stirred solution of compound 22 (0.24 g, 0.43 mmol) in dry THF (5 ml). After 3 hours, the reaction mixture was cooled to 0 ° C. and 10% NaOH (1 ml) was carefully added dropwise, followed by 30% H ₂ O ₂ (1 ml). The reaction mixture was stirred at room temperature for 1 h, neutralized with 3M HCl and extracted with ethyl acetate (3 × 20 ml). The combined organic phases were washed with brine, dried (MgSO ₄ ) and the solvent was removed in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent: hexane: EtOAc, 30: 1 → 1: 1) to obtain methyl 3β-hydroxy-2β- (3-hydroxypropyl) -20,29-dihydrobetulinate (27). Yield: 76%.

Compound 27 (0.44 mmol), pyridine (0.06 g, 0.77 mmol) and DMAP (0.03 g, 0.25 mmol) were dissolved in CH ₂ Cl ₂ (2 ml) and cooled to 0 ° C. To this solution was added dropwise a solution of methanesulfonyl chloride (0.07 g, 0.65 mmol) in CH ₂ Cl ₂ (1 ml) and stirred at room temperature for 20-24 hours, preferably 24 hours (TLC control). After completion of the reaction, cold 5% HCl solution was added and the mixture was extracted with ethyl acetate (25 ml). The extract was washed with saturated NaHSO ₃ solution, water, dried (MgSO ₄ ) and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent hexane - EtOAc, 10: 1 → 1: 1), methyl 3β-acetoxy-2β- (3-mesyloxypropyl) -20,29-dihydrobetulinate was obtained (33). Yield 93%.

Mesylate 33 (0.42 mmol) was dissolved in dry acetone (12 ml) and LiBr (0.06-0.07 g, 0.7-0.8 mmol, mainly 0.8 mmol) was added. The mixture was boiled for 3-4 hours, mainly 3 hours, and then cooled, the precipitate was filtered off and washed with acetone (2 ml). The filtrate and washes were combined and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent - hexane: EtOAc, 30: 1 → 1: 1). Methyl 3β-acetoxy-2β- (3-bromopropyl) -20,29-dihydrobetulinate was obtained (36). Yield 79%.

A mixture of bromide 36 (0.24 mmol) and triphenylphosphine (0.72 mmol-1.2 mmol, mainly 0.72 mmol) was boiled in toluene (8 ml) for 28-32 h, mainly 32 h (TLC control). After completion of the reaction, the solution was cooled and evaporated in vacuo, the resulting solid was washed with hot hexane (7 ml × 2), dissolved in a minimum volume of EtOAc (2 ml) and precipitated with hexane (8 ml). The precipitate was filtered off, and methyl-3β-acetoxy-2β- (3-triphenylphosphoniopropyl) -20,29-dihydrobetulinate bromide was obtained (1). Yield 94%.

Example 2

Methyl 3α-acetoxy-2β- (3-triphenylphosphoniopropyl) -20,29-dihydrobetulinate bromide (2): Compound 17 (0.8 mmol) was dissolved in dry THF (15 ml) in an Ar atmosphere, cooled to -78 ° C and a 1 M solution of L-selectride in THF (2.4 ml, 2.4 mmol) was added. Then, the solution was stirred at room temperature for 2 hours, and a 2M NaOH solution (18 ml) and H ₂ O ₂ (30%, 4 ml) were added, stirred for 1 hour. The mixture was evaporated to a small volume and extracted with ethyl acetate. The organic phase was washed with water, dried (MgSO ₄ ) and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent hexane: EtOAc, 30: 1 → 1: 1). Methyl 2β-allyl-3α-hydroxy-20,29-dihydrobetulinate was obtained (21). Yield 68%.

Then, compound 21 was transformed into target salt 2 (yield 89%) through intermediate acetate 24 (yield 96%), primary alcohol 28 (yield 74%), mesylate 34 (yield 87%), bromide 37 (yield 78%) according to the method, described in example 1.

Example 3

Methyl-3β-hydroxy-2β- (3-triphenylphosphoniopropyl) -20,29-dihydrobetulinate iodide (3): Compound 25 was obtained by reducing compound 17 with a NaBH ₄ -CeCl ₃ · 7H ₂ O reagent and hydroborating the allyl fragment in compound 19 by the method described in example 1.

To a solution of compound 25 (0.35 mmol) in 6 ml of dry THF at 0 ° C in an argon atmosphere was added Ph ₃ P (triphenylphosphine) (0.220 g, 0.84 mmol), imidazole (0.117 g, 1.17 mmol), crystalline I ₂ ( 0.191 g, 0.75 mmol) and stirred at 0 ° C for 0.5-1 h, mainly 1 h. After completion of the reaction, the solution was diluted with ethyl acetate (10-15 ml) and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent hexane: EtOAc, 30: 1 → 1: 1). Methyl 3β-hydroxy-2β- (3-iodopropyl) -20,29-dihydrobetulinate was obtained (30). Yield 57%.

Iodide 30 was transformed into salt 3 (yield 21%) according to the method described in example 1, but characterized in that the mixture of iodide and triphenylphosphine was boiled in toluene for 16-20 hours, mainly 16 hours

Example 4

Methyl-3α-hydroxy-2β- (3-triphenylphosphoniopropyl) -20,29-dihydrobetulinate iodide (4): Compound 21 was obtained by reduction of compound 17 using an L-selectride according to the method described in Example 2.

Compound 21 was transformed into primary alcohol 26 by the hydroboration reaction by the method described in example 1.

Compound 26 was transformed into iodide 31 (yield 73%) by reaction with crystalline iodine and imidazole according to the method described in example 3, iodide 31 was transformed into salt 4 (yield 23%) according to the method described in example 3.

Example 5

3β-acetoxy-2β- (3-triphenylphosphoniopropyl) -20,29-dihydrobetulinic bromide (5): Benzyl dihydrobetulonate 18 was reduced to alcohol 20 (yield 72%), followed by acetylation to compound 23 (87% yield). Acetate 23 was transformed into primary alcohol 29 (yield 79%), mesylate 35 (yield 90%) and bromide 38 (yield 81%) according to the method described in example 1. Benzyl protection was removed by hydrogenolysis in the presence of Pd / C in Et ₂ O with obtaining compound 39 (yield 98%), which was transformed into salt 5 (yield 81%) according to the method described in example 1.

Example 6

Benzyl-3β-acetoxy-2β- (3-triphenylphosphoniopropyl) -20,29-dihydrobetulinate iodide (6): Obtained by the method described in example 5, but characterized in that primary alcohol 29 was transformed into primary iodide 32 (yield 94% ) and then to triphenylphosphonium salt 6 (yield 78%) according to the method described in example 3.

Example 7

3β-acetoxy-30-triphenylphosphoniolup-20 (29) -en-28-oic acid methyl ester bromide (7). A mixture of compound 42a (2.09 g, 4.08 mmol) and NBS (1.45 g, 8.16 mmol) in CCl ₄ (83 ml) was stirred at 50 ° C. for 4 days. The precipitate was filtered off, the filtrate was evaporated and chromatographed on a SiO ₂ column (eluent: hexane: EtOAc, 30: 1 → 10: 1). Recrystallization of the product from hexane gave compound 43a (1.55 g, 64%). 3β-acetoxy-30-bromolup-20 (29) -en-28-oic acid methyl ester (43a): Yield 64%. White crystals mp = 216-218 ° C, 218-220 ° C lit. [NV Uzenkova, NI Petrenko, M.M. Shakirov, E.E. Shul`ts, and GA Tolstikov. Chemistry of Natural Compounds, V 41, No. 6, 2005, 692-700]. 1 H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.72 (m, 1H, H (5)); 0.83 (s, 3H, H (24)); 0.84 (s, 6H, H (23), H (25)); 0.91 (s, 3H, H (26)); 0.97 (s, 3H, H (27)); 1.00-2.30 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton); 2.05 (s, 3H, Me (OAc)); 3.04 (m, 1H, H (19)); 3.68 (s, 3H, OMe); 3.99 (br s, 2H, H (30)); 4.46 (dd, 1H, H (3), J = 10.5, J = 5.8); 5.04, 5.14 (br s, 2H, H (29)).

(с 0.46, CHCl₃). MS, m/z 773.396 [M-Br]+. ЯМР 1H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.72 (м, 1H, H(5)); 0.78, 085 (оба м, 15H, H(23)-H(27)); 1.00-2.20 (м, 23H, CH, CH₂, в пентациклическом скелете); 1.99 (с 3H, Me (OAc)); 2.75 (м, 1H, H(19)); 3.51 (с, 3H, OMe); 4.40 (д.д, 1H, H(3), J=10, J=5.2); 4.56 (д, 2H, H(30), J=15); 4.91, 5.11 (оба д, 2H, H(29), J=3.2), 7.50-7.90 (м, 15H, Ph). ЯМР ³¹P (161.98 МГц, CDCl₃): δ 20.68.A mixture of 43a bromide (0.24 g, 0.4 mmol) and triphenylphosphine (2 mmol) was boiled in acetonitrile (20 ml) for 4-5 hours (TLC control). After completion of the reaction, the solution was cooled and evaporated in vacuo, the resulting solid was washed with hot hexane (10 ml × 2), dissolved in a minimum volume of EtOAc (2-4 ml) and precipitated with hexane (12 ml). The precipitate was filtered off to give compound 7 (0.31 g, 90%). White crystals. Mp = 174-175 ° C, $${[\alpha ]}_D^{\mathrm{twenty}}=+1.09°$$

(c 0.46, CHCl ₃ ). MS, m / z 773.396 [M-Br] +. 1 H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.72 (m, 1H, H (5)); 0.78, 085 (both m, 15H, H (23) -H (27)); 1.00-2.20 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton); 1.99 (s 3H, Me (OAc)); 2.75 (m, 1H, H (19)); 3.51 (s, 3H, OMe); 4.40 (dd, 1H, H (3), J = 10, J = 5.2); 4.56 (d, 2H, H (30), J = 15); 4.91, 5.11 (both d, 2H, H (29), J = 3.2), 7.50-7.90 (m, 15H, Ph). ³¹ P NMR (161.98 MHz, CDCl ₃ ): δ 20.68.

Example 8

3β, 28-diacetoxy-30-triphenylphosphoniolup-20 (29) -ene bromide (8). A mixture of compound 40 (2.15 g, 4.08 mmol) and NBS (1.45 g, 8.16 mmol) in CCl ₄ (83 ml) was stirred at 50 ° C. for 4 days. The precipitate was filtered off, the filtrate was evaporated and chromatographed on a SiO ₂ column (eluent: hexane: EtOAc, 30: 1 → 10: 1). Recrystallization of the product from hexane gave compound 41 (1.48 g, 60%). 3β, 28-Diacetoxy-30-bromolup-20 (29) -en (41): White crystals. Mp = 183-185 ° C, lit., 185 ° C, [I-Ch. Sun, HK. Wang, Y. Kashiwada, JK. Shen Cosentino LM, Ch-HChen, LM. Yang, and KH. Lee. J. Med. Chem. 1998, 41, 4648-4657]; lit., 195-196 ° C. [NV Uzenkova, NI Petrenko, MM Shakirov, EE Shul`ts, and GA Tolstikov. Chemistry of Natural Compounds, V 41, No. 6, 2005, 692-700]. ¹ H NMR (400 MHz, CDCl 3, 5, ppm, J / Hz,): 0.78 (m, 1H, H (5)); 0.84, 0.85, 0.87 (s, 3H each, Н (23), Н (24), Н (25)); 0.99 (s, 3H, H (27)); 1.08 (s, 3H, H (26)); 0.95-2.25 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton); 2.04, 2.08 (both 3H, Me (OAc)); 2.44 (dt, 1H, H (19), J = 5.5, J = 10.8); 3.85, 4.28 (d, 2H, H (28), J = 11.2); 3.98 (s, 2H, H (30)); 4.47 (dd, 1H, H (3), J = 5.5, J = 10.5); 5.04, 5.14 (s, 2H, H (29)).

(с 0.64, CHCl₃). MS, m/z 787.298 [M-Br]+^. ЯМР ¹H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.69 (м, 1H, Н(5)); 0.76 (м, 9H, Н(23), H(24), H(25)); 0.84 (с, 3H, H(27)); 0.91 (с, 3H, H(26)); 0.95-2.30 (м, 23H, CH, CH₂, в пентациклическом скелете); 1.96, 1.97 (оба с по 3H, Me (ОАс)); 2.25 (м, 1H, H(19)); 3.40, 4.03 (д, 2H, H(28), J=10.8); 4.39 (д.д, 1H, H(3), J=5.6, J=10.5); 4.56, 5.70 (м, 2H, H(30)); 4.98, 5.10 (уш.с, 2H, H(29)); 7.60-7.90 (м, 15H, Ph). ЯМР ³¹P (161.98 МГц, CDCl₃): δ 19.84. A mixture of 41 bromide (0.24 g, 0.4 mmol) and triphenylphosphine (0.52 g, 2 mmol) was boiled in acetonitrile (18 ml) for 4–5 h (TLC control). After completion of the reaction, the solution was cooled and evaporated in vacuo, the resulting solid was washed with hot hexane (10 ml × 2), dissolved in a minimum volume of EtOAc (2-4 ml) and precipitated with hexane (12 ml). The precipitate was filtered off to give compound 8 (0.32 g, 92%). White crystals mp = 177-179 ° C, $${[\alpha ]}_D^{\mathrm{twenty}}+8.1°$$

(c 0.64, CHCl ₃ ). MS, m / z 787.298 [M-Br] + ^{. 1} H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.69 (m, 1H, H (5)); 0.76 (m, 9H, H (23), H (24), H (25)); 0.84 (s, 3H, H (27)); 0.91 (s, 3H, H (26)); 0.95-2.30 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton); 1.96, 1.97 (both 3H, Me (OAc)); 2.25 (m, 1H, H (19)); 3.40, 4.03 (d, 2H, H (28), J = 10.8); 4.39 (dd, 1H, H (3), J = 5.6, J = 10.5); 4.56, 5.70 (m, 2H, H (30)); 4.98, 5.10 (br.s, 2H, H (29)); 7.60-7.90 (m, 15H, Ph). ³¹ P NMR (161.98 MHz, CDCl ₃ ): δ 19.84.

Example 9

3β-Acetoxy-30-triphenylphosphoniolup-20 (29) -en-28-oic acid (9). A mixture of compound 42 (2.04 g, 4.08 mmol) and NBS (1.45 g, 8.16 mmol) in CCl ₄ (83 ml) was stirred at 50 ° C. for 4 days. The precipitate was filtered off, the filtrate was evaporated and chromatographed on a SiO ₂ column (eluent: hexane: EtOAc, 30: 1 → 10: 1). Recrystallization of the product from hexane gave compound 43 (1.32 g, 56%). 3β-Acetoxy-30-bromolup-20 (29) -en-28-oic acid (43): White crystals. ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.72 (m, 1H, H (5)); 0.84 (s, 3H, H (24)); 0.85, 0.86 (both with 3H, H (23), H (25)); 0.94 (s, 3H, H (26)); 0.99 (s, 3H, H (27)); 1.08-2.40 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton); 2.05 (s 3H, Me (OAc)); 3.05 (td, 1H, H (19), J = 10.8, J = 4.5); 4.04 (br.s, 2H, H (30)); 4.49 (dd, 1H, H (3), J = 10.4, J = 6); 5.06, 5.16 (both s, 2H, H (29)); 5.68 (m, 1H, CO ₂ H). The compound was used in further transformation into triphenylphosphonium salt 9 without further purification.

(с 0.48, CHCl₃). MS, m/z 759.394 [M-Br]+. ЯМР ¹H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.73 (м, 1H, H(5)); 0.81, 0.82 (оба м, 12H, H(23)-H(26)); 0.92 (с, 3H, H(27)); 0.90-2.28 (м, 23H, CH, CH₂, в пентациклическом скелете); 2.06 (с 3H, Me (AAc)); 2.72 (м, 1H, H(19)); 4.50 (м, 3H, H(3), H(30)); 4.96, 5.17 (оба уш.с, 2H, H(29)); 5.76 (м, 1H, CO₂H); 7.50-7.81 (м, 15H, Ph). ЯМР ³¹P (161.98 МГц, CDCl₃): δ 20.12.A mixture of 43 bromide (0.23 g, 0.4 mmol) and triphenylphosphine (0.52 g, 2 mmol) was boiled in acetonitrile (20 ml) for 4–5 h (TLC control). After completion of the reaction, the solution was cooled and evaporated in vacuo, the resulting solid was washed with hot hexane (10 ml × 2), dissolved in a minimum volume of EtOAc (2-4 ml) and precipitated with hexane (12 ml). The precipitate was filtered off to give compound 9 (0.29 g, 87%). White crystals. Mp = 221-222 ° C, $${[\alpha ]}_D^{\mathrm{twenty}}=+5.20°$$

(c 0.48, CHCl ₃ ). MS, m / z 759.394 [M-Br] +. ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.73 (m, 1H, H (5)); 0.81, 0.82 (both m, 12H, H (23) -H (26)); 0.92 (s, 3H, H (27)); 0.90-2.28 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton); 2.06 (s 3H, Me (AAc)); 2.72 (m, 1H, H (19)); 4.50 (m, 3H, H (3), H (30)); 4.96, 5.17 (both broad s, 2H, H (29)); 5.76 (m, 1H, CO ₂ H); 7.50-7.81 (m, 15H, Ph). ³¹ P NMR (161.98 MHz, CDCl ₃ ): δ 20.12.

Example 10

Methyl-3β-acetoxy-2β (3-triphenylphosphoniopropyl) -29-iodomethyl-20 (29) -dihydrobetulinate iodide (10): BH ₃ · THF complex (1 M solution in THF) (1.8 ml, 1.8 mmol) was added under argon at room temperature, to a stirred solution of compound 51 (0.24 g, 0.43 mmol) in dry THF (5 ml). After 3 hours, the reaction mixture was cooled to 0 ° C and 10% NaOH (4 ml) was carefully added dropwise, followed by 30% H ₂ O ₂ (4 ml). The reaction mixture was stirred at room temperature for 1 h, neutralized with 3M HCl and extracted with ethyl acetate. The combined organic phases were washed with brine, dried with MgSO ₄ and the solvent was removed in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent — hexane: EtOAc, 30: 1 → 1) to give compound 52 (0.14 g, 54%) as white crystals. Methyl-3β-acetoxy-2α (3-hydroxypropyl) -29-hydroxymethyl-20 (29) -dihydrobetulinate (52): ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.68 (t, 1H, H (1), J = 12); 0.80 (d, 3H, H (30), J = 6.8); 0.87, 0.91, 0.95, 0.96, 0.98 (all with 3H, H (23) -H (27)); 1.05-2.40 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton, 1H, H (20), 2H, H (1 ′), 2H, H (2 ′)); 2.08 (s, 3H, Me (OAc)); 3.42 (m, 1H, H (29)); 3.77 (dd. 1H, H (29), J = 10, J = 4); 3.58 (m, 2H, H (3 ′)); 3.60 (s, 3H, CO ₂ Me); 4.45 (d, 1H, H (3), J = 11.2). The compound was used in further transformation into iodide 53 without further purification.

To a solution of compound 52 (0.12 g, 0.20 mmol) in dry THF (6 ml) at 0 ° C under argon was added Ph ₃ P (triphenylphosphine) (0.13 g, 1.96 mmol), imidazole ( 0.13 g, 1.96 mmol), crystalline 12 (0.16 g, 0.86 mmol) and stirred at 0 ° C for 1 h. After completion of the reaction, the solution was diluted with ethyl acetate (10 ml) and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent: hexane: EtOAc, 30: 1 → 1: 1) to give compound 53 (0.11 g, 68%) as white crystals. Methyl-3β-acetoxy-2β (3-iodopropyl) -29-iodomethyl-20 (29) -dihydrobetulinate (53): ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.69 (t, 1H, H (1), J = 12.8); 0.82, 0.83, 0.87, 0.91, 0.97 (all with 3H, H (23) -H (27)); 1.07 (d, 3H, H (30), J = 6.4); 1.10-2.45 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton, 1H, H (20), 2H, H (1 ′), 2H, H (2 ′)); 2.12 (s, 3H, Me (OAc)); 2.85 (m, 1H, H (29)); 3.12 (m, 2H, H (3 ′)); 3.41 (d, 1H, H (29), J = 7.2); 3.66 (s, 3H, CO ₂ Me); 4.46 (d, 1H, H (3), J = 10.8). The compound was used in further transformation into triphenylphosphonium salt 10 without further purification.

(с 0.49, CHCl₃), MS, m/z 943.312 [M-I]⁺, MS, m/z 815.349 [M-I₂]⁺. ЯМР ¹H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.50 (т, 1H, H(1), J=12); 0.72, 0.75, 0.77, 0.85, 0.87 (все с по 3H, H(23)-H(27)); 0.88 (д, 3H, H(30), J=6.8); 1.00-2.45 (м, 23H, CH, CH₂, в пентациклическом скелете, 1H, H(20), 2H, H(1′), 2H, H(2′)); 2.07 (с, 3H, Me (OAc)); 2.83 (м, 1H, H(29)); 3.40 (м, 1H, H(3′), 1H, H(29)); 3.60 (с, 3H, CO₂Me); 3.70 (м, 1H, H(3′)); 4.29 (д, 1H, H(3), J=10,4); 7.60-7.90 (м, 15H, Ph). ЯМР ³¹P (161.98 МГц, CDCl₃): δ 24.25.A mixture of iodide 53 (0.11 g, 0.14 mmol) and triphenylphosphine (0.36 g, 1.36 mmol) was boiled in acetonitrile or toluene (5 ml) for 18 h (TLC control). After completion of the reaction, the solution was cooled and evaporated in vacuo, the resulting solid was washed with hot hexane (10 ml × 2), dissolved in EtOAc (4 ml) and precipitated with hexane (12 ml). The precipitate was filtered off to give compound 10 (0.10 g, 69%). White crystals. Mp = 154-156 ° C, $${[\alpha ]}_D^{\mathrm{twenty}}=-22.2°$$

(c 0.49, CHCl ₃ ), MS, m / z 943.312 [MI] ⁺ , MS, m / z 815.349 [MI ₂ ] ⁺ . ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.50 (t, 1H, H (1), J = 12); 0.72, 0.75, 0.77, 0.85, 0.87 (all with 3H, H (23) -H (27)); 0.88 (d, 3H, H (30), J = 6.8); 1.00-2.45 (m, 23H, CH, CH ₂ , in the pentacyclic skeleton, 1H, H (20), 2H, H (1 ′), 2H, H (2 ′)); 2.07 (s, 3H, Me (OAc)); 2.83 (m, 1H, H (29)); 3.40 (m, 1H, H (3 ′), 1H, H (29)); 3.60 (s, 3H, CO ₂ Me); 3.70 (m, 1H, H (3 ′)); 4.29 (d, 1H, H (3), J = 10.4); 7.60-7.90 (m, 15H, Ph). ³¹ P NMR (161.98 MHz, CDCl ₃ ): δ 24.25.

Example 11

3β-acetoxy-2β- (3-triphenylphosphoniopropyl) urs-12en-28-oic acid methyl ester iodide (11): To a solution of compound 44 (0.50 g, 1.06 mmol) in CH ₂ Cl ₂ (13 ml) at 0 ° C and with vigorous stirring, PCC (pyridinium chlorochromate) (0.684 g, 3.18 mmol) was added. The reaction mixture was stirred at room temperature for a control time (1-2 hours, TLC control). Then silica gel (3 g) was added (Imid Silica gel, fraction 50 ± 160 μm), EtOAc (15 ml), stirred for 5 minutes and separated from the precipitate on a Schott filter, the filtrate was washed with saturated aqueous brine, water and dried with MgSO ₄ . The residue was evaporated and chromatographed on a column of SiO ₂ (eluent — hexane — EtOAc) to give compound 45 (0.40 g, 94%). Ursonic acid methyl ester (45): White crystals mp = 192-194 ° C, lit., 192-193 ° C, [SM Jainand S.K. Atal. IndianJ. Chem., Sec. B., 1986, 25B (4), 427]. Lit, 191-193 ° C (EtOH), [α] ²⁴ +87.8 (s 0.5, CHCl ₃ ) [AV Korovin and AV Tkachev. Russian Chemical Bulletin, International Edition, 2001, V 50, No. 2, 304-310]. ¹ H NMR spectrum (5, ppm, J / Hz): 0.80, 0.95, 1.05, 1.09, 1.10 (s, 3H, H (23) -H (27)); 0.84, 0.94 (both d, 3H, H (29), H (30), J = 6); 1.25-2.05 (m, 22 H, CH, CH ₂ in the skeleton); 2.25 (d, 1H, H (18), J = 11.6); 3.62 (s, 3H, OMe); 5.27 (t, 1H, H (12), J = 3.6).

(с 0.29, CHCl₃). IR (v/см^-1): 1726 (С=O). MS, m/z 531.439 [M+Na]⁺, 547.379 [M+K]⁺. Структурная формула C₃₄H₅₂O₃: C, 79.95; H, 10.26. Найдено: C, 80.26; H, 10.30. ЯМР 1H (400 МГц, CDCl₃, δ, м.д, J/Гц,): 0.80, 0.94, 1.06, 1.07, 1.19 (с, 3H, H(23)-H(27)); 0.84, 0.92 (оба д, 3H, H(29), H(30), J=6); 1.25-2.04 (м, 20 H, CH, CH₂ в скелете и 1H, H(1′); 2.25 (д, 1H, H(18), J=10.8); 2.56 (д.т, 1H, H(1′), J=14.4, J=5.5); 2.72 (м, 1H, H(2)); 3.63 (с, 3H, OMe); 5.01-5.06 (м, 2H, H(3′)); 5.27 (т, 1H, H(12), J=3.6); 5.76 (м, 1H, H(2′)).To a solution of compound 45 (1 g, 2.13 mmol) in DME (10 ml) and THF (8 ml) at room temperature in an argon atmosphere, KN (SiMe ₃ ) ₂ (1 M solution in THF) was added with stirring, (2.77 ml, 2.77 mmol). After 15 minutes, Et ₃ B (1 M solution in THF), (2.77 ml, 2.77 mmol) was added to the solution and stirred for 1 h. Then, a solution of allyl bromide in 5 ml of DME (4.69 mmol) was added. The reaction mixture was stirred for a control time (4 h, TLC control), neutralized with 3M HCl, diluted with water (17 ml) and extracted with EtOAc (10 ml × 3). The combined extracts were dried with MgSO ₄ . The residue was evaporated and chromatographed on a column of SiO ₂ (eluent was hexane-EtOAc, 30: 1 → 1: 1), compound 46 (0.87 g, 94%) was obtained. 2β-allyl-3-oxours-12en-28-oic acid methyl ester (46): White crystals mp = 109-112 ° C (EtOH), $${[\alpha ]}_D^{25}+35.17°$$

(c 0.29, CHCl ₃ ). IR (v / cm ^-1 ): 1726 (C = O). MS, m / z 531.439 [M + Na] ⁺ , 547.379 [M + K] ⁺ . Structural formula C ₃₄ H ₅₂ O ₃ : C, 79.95; H, 10.26. Found: C, 80.26; H, 10.30. 1H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.80, 0.94, 1.06, 1.07, 1.19 (s, 3H, H (23) -H (27)); 0.84, 0.92 (both d, 3H, H (29), H (30), J = 6); 1.25-2.04 (m, 20 H, CH, CH ₂ in the skeleton and 1H, H (1 ′); 2.25 (d, 1H, H (18), J = 10.8); 2.56 (d.t, 1H, H ( 1 ′), J = 14.4, J = 5.5); 2.72 (m, 1H, H (2)); 3.63 (s, 3H, OMe); 5.01-5.06 (m, 2H, H (3 ′)); 5.27 (t, 1H, H (12), J = 3.6); 5.76 (m, 1H, H (2 ′)).

(с 0.8, CHCl₃). IR (v/см^-1): 1722 (C=O), 3433 (OH). MS, m/z 533.251 [M+Na]⁺. Структурная формула C₃₄H₅₄O₃: С, 79.95; H, 11.01. Найдено: C, 79.98; H, 10.71. ЯМР ¹H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.66 (т, 1H, $$H_{eq}^a(1)$$

, J=12); 0.70, 0.82, 0.95, 1.01, 1.10 (с, 3H, H(23)-H(27)); 0.88, 0.96 (оба д, 3H, H(29), H(30), J=6); 1.26-2.06 (м, 19 H, CH, CH₂ в скелете и 2H, H(1′), $$H_{ax}^b(1)$$

); 2.25 (д, 1H, H(18), J=11); 2.49 (м, 1H, H(1′); 2.92 (д, 1H, H(3), J=10.4); 3.63 (с, 3H, OMe); 5.03-5.10 (м, 2H, H(3′)); 5.27 (т, 1H, H(12) J=3.6); 5.86 (м, 1H, H(2′)).Compound 46 (0.50 g, 0.98 mmol) was dissolved in a THF - methanol solvent mixture (1: 2, 22.5 ml) in an Ar atmosphere and cooled to -30 ° C. To this solution was added dropwise a solution of CeCl ₃ · 7H ₂ O (0.48 g, 1.27 mmol) in a mixture of THF - methanol solvents (1: 1, 1 ml). Then NaBH ₄ (0.075 g, 1.96 mmol) was added in small portions over 5 min, the temperature was raised to room temperature and stirred at this temperature for 2 h (the reaction progress was monitored by TLC). After completion, a 5% HCl solution was added and the mixture was extracted with EtOAc (10 ml × 3). The extract was washed with saturated aqueous NaHCO ₃ , water, dried with MgSO _4, and the solvent was removed in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent: hexane: EtOAc 30: 1 → 1: 1), compound 47 (0.40 g, 80%) was obtained as white crystals. 2β-allyl-3β-hydroxyurs-12en-28-oic acid methyl ester (47): White crystals. Mp = 203-206 ° C (EtOH), $${[\alpha ]}_D^{25}+55.1°$$

(c 0.8, CHCl ₃ ). IR (v / cm ^-1 ): 1722 (C = O), 3433 (OH). MS, m / z 533.251 [M + Na] ⁺ . Structural formula C ₃₄ H ₅₄ O ₃ : C, 79.95; H, 11.01. Found: C, 79.98; H, 10.71. ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.66 (t, 1H, $$H_{eq}^a(\mathrm{one})$$

, J = 12); 0.70, 0.82, 0.95, 1.01, 1.10 (s, 3H, H (23) -H (27)); 0.88, 0.96 (both d, 3H, H (29), H (30), J = 6); 1.26-2.06 (m, 19 H, CH, CH ₂ in the skeleton and 2H, H (1 ′), $$H_{ax}^b(\mathrm{one})$$

); 2.25 (d, 1H, H (18), J = 11); 2.49 (m, 1H, H (1 ′); 2.92 (d, 1H, H (3), J = 10.4); 3.63 (s, 3H, OMe); 5.03-5.10 (m, 2H, H (3 ′) ); 5.27 (t, 1H, H (12) J = 3.6); 5.86 (m, 1H, H (2 ′)).

(с 0.36, CHCl₃). IR (v/см^-1): 1735 (C=O). MS, m/z 575.204 [M+Na]⁺. Структурная формула C₃₆H₅₆O₄: C, 78.21; H, 10.21. Найдено: С, 77.97; H, 10.19. ЯМР ¹H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.76, 0.84, 0.86, 0.97, 1.09 (с, 3H, H(23)-H(27)); 0.88, 0.96 (оба д, 3H, H(29), H(30), J=6); 1.30-2.14 (м, 21H, CH, CH₂ в скелете и 2H, H(1′)); 2.09 (с, 3H, Me (OAc)); 2.25 (д, 1H, H(18), J=11); 3.62 (с, 3H, OMe); 4.49 (д, 1H, H(3), J=10.8); 4.98-5.01 (м, 2H, H(3′)); 5.26 (т, 1H, H(12) J=3.6); 5.73 (м, 1H, H(2′)).To a solution of compound 47 (0.39 g, 0.77 mmol) in 17 ml of dry THF cooled to 0 ° C, pyridine (0.16 ml, 1.92 mmol), acetyl chloride (0.13 ml, 1.54 mmol), 4-dimethylaminopyridine (DMAP) were added with stirring ) (0.047 g, 0.39 mmol) and stirred at room temperature for 24 h (TLC control). Then a 10% solution of HC1 was added and the mixture was extracted with ethyl acetate (15 ml × 4). The extract was washed with brine, dried over MgSO _4, and the solvent was evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent: hexane: EtOAc, 30: 1 → 15: 1). Compound 48 was obtained (0.35 g, 83%). 2β-allyl-3β-acetoxyurs-12en-28-oic acid methyl ester (48): 95% yield. White crystals mp = 186-188 ° C (EtOH), $${[\alpha ]}_D^{25}+14.2°$$

(c 0.36, CHCl ₃ ). IR (v / cm ^-1 ): 1735 (C = O). MS, m / z 575.204 [M + Na] ⁺ . Structural formula C ₃₆ H ₅₆ O ₄ : C, 78.21; H, 10.21. Found: C, 77.97; H, 10.19. ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.76, 0.84, 0.86, 0.97, 1.09 (s, 3H, H (23) -H (27)); 0.88, 0.96 (both d, 3H, H (29), H (30), J = 6); 1.30-2.14 (m, 21H, CH, CH ₂ in the skeleton and 2H, H (1 ′)); 2.09 (s, 3H, Me (OAc)); 2.25 (d, 1H, H (18), J = 11); 3.62 (s, 3H, OMe); 4.49 (d, 1H, H (3), J = 10.8); 4.98-5.01 (m, 2H, H (3 ′)); 5.26 (t, 1H, H (12) J = 3.6); 5.73 (m, 1H, H (2 ′)).

The BH ₃ · THF complex (1M solution in THF) (1.24 ml, 1.24 mmol) was added in an argon atmosphere at room temperature to a stirred solution of compound 48 (0.34 g, 0.62 mmol) in dry THF (7 ml). After 3 hours, the reaction mixture was cooled to 0 ° C, and a 10% NaOH solution (1.6 ml) was carefully added dropwise, followed by 30% H ₂ O ₂ (1.6 ml). The reaction mixture was stirred at room temperature for 1 h, neutralized with 3M HCl and extracted with ethyl acetate (20 ml × 3). The combined organic phases were washed with brine, dried with MgSO ₄ and the solvent was removed in vacuo.

The residue was purified by column chromatography on SiO ₂ (eluent-hexane: EtOAc, 30: 1 → 1: 1) to give compound 49 (0.21 g, 60%).

(с 0.22, CHCl₃). IR (v/см^-1): 1732 (C=O), 3446 (OH). MS, m/z 593.540 [M+Na]⁺. Структурная формула C₃₆H₅₈O₅: С, 75.75; H, 10.24. Найдено: C, 75.82; H, 10.19. ЯМР ¹H (400 МГц, CDCl₃, δ, m.д., J/Гц,): 0.76,0.84, 0.85, 0.97,1.09 (с, 3H, H(23)-H(27)); 0.88, 0.96 (оба д, 3H, H(29), H(30), J=6); 1.22-2.01 (м, 21H, CH, CH₂ в скелете, 2H, H(1′), 2H, H(2′)); 2.10 (с, 3H, Me (OAc)); 2.25 (д, 1H, H(18), J=11.6); 3.57 (м, 2H, H(3′)); 3.62 (с, 3H, OMe); 4.48 (д, 1H, H(3), J=10.8); 5.25 (т, 1H, H(12) J=3.6).3β-acetoxy-2β- (3-hydroxypropyl) urs-12en-28-oic acid methyl ester (49): White crystals, mp. 123-125 ° C (EtOH), $${[\alpha ]}_D^{25}+19.5°$$

(c 0.22, CHCl ₃ ). IR (v / cm ^-1 ): 1732 (C = O), 3446 (OH). MS, m / z 593.540 [M + Na] ⁺ . Structural formula C ₃₆ H ₅₈ O ₅ : C, 75.75; H, 10.24. Found: C, 75.82; H, 10.19. ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.76.0.84, 0.85, 0.97.1.09 (s, 3H, H (23) -H (27)); 0.88, 0.96 (both d, 3H, H (29), H (30), J = 6); 1.22-2.01 (m, 21H, CH, CH ₂ in the skeleton, 2H, H (1 ′), 2H, H (2 ′)); 2.10 (s, 3H, Me (OAc)); 2.25 (d, 1H, H (18), J = 11.6); 3.57 (m, 2H, H (3 ′)); 3.62 (s, 3H, OMe); 4.48 (d, 1H, H (3), J = 10.8); 5.25 (t, 1H, H (12) J = 3.6).

(с 0.26, CHCl₃). IR (v/см^-1): 756 (CH₂-I), 1734 (C=O). Структурная формула C₃₆H₅₇O₄I: C, 63.52; H, 8.44; I, 18.64. Найдено: C, 63.48; H, 8.37; I, 17.72. ЯМР ¹H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.76, 0.84, 0.85, 0.95, 1.09 (с, 3H, H(23)-H(27)); 0.89, 0.96 (оба д, 3H, H(29), H(30), J=6.4); 1.27-2.02 (м, 21 H, CH, CH₂ в скелете, 2H, H(1′), 2H, H(2′)); 2.13 (с, 3H, Me (OAc)); 2.25 (д, 1H, H(18), J=11.2); 3.14 (м, 2H, H(3′)); 3.62 (с, 3H, OMe); 4.47 (д, 1H, H(3), J=10.8); 5.26 (т, 1H, H(12) J=3.6).To a solution of compound 49 (0.20 g, 0.35 mmol) in 6 ml of dry THF at 0 ° C in an argon atmosphere was added Ph ₃ P (triphenylphosphine) (0.22 g, 0.84 mmol), imidazole (0.12 g, 1.72 mmol), crystalline I ₂ (0.19 g, 0.75 mmol) and stirred at 0 ° C for 1 h. After completion of the reaction, the solution was diluted with ethyl acetate (10-15 ml) and evaporated in vacuo. The residue was purified by column chromatography on SiO ₂ (eluent hexane: EtOAc, 30: 1 → 1: 1). Compound 50 was obtained (0.21 g, 88%). 3β-acetoxy-2β- (3-iodopropyl) urs-12en-28-oic acid methyl ester (50): White crystals mp = 116-118 ° C (EtOH), $${[\alpha ]}_D^{26}+3.46°$$

(c 0.26, CHCl ₃ ). IR (v / cm ^-1 ): 756 (CH ₂ -I), 1734 (C = O). Structural formula C ₃₆ H ₅₇ O ₄ I: C, 63.52; H, 8.44; I, 18.64. Found: C, 63.48; H, 8.37; I, 17.72. ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.76, 0.84, 0.85, 0.95, 1.09 (s, 3H, H (23) -H (27)); 0.89, 0.96 (both d, 3H, H (29), H (30), J = 6.4); 1.27-2.02 (m, 21 H, CH, CH ₂ in the skeleton, 2H, H (1 ′), 2H, H (2 ′)); 2.13 (s, 3H, Me (OAc)); 2.25 (d, 1H, H (18), J = 11.2); 3.14 (m, 2H, H (3 ′)); 3.62 (s, 3H, OMe); 4.47 (d, 1H, H (3), J = 10.8); 5.26 (t, 1H, H (12) J = 3.6).

(с 0.28, CHCl₃). IR (v/см^-1): 1726 (C=O). MS, m/z 815.611 [M-I]⁺. ЯМР ¹H (400 МГц, CDCl₃, δ, м.д., J/Гц,): 0.51 (т, 1H, $$H_{eq}^a(1)$$

, J=12.4); 0.71, 0.76, 0.79, 0.89, 1.04 (с, 3H, H(23)-H(27)); 0.88, 0.95 (оба д, 3H, H(29), H(30), J=5.6); 1.27-2.03 (м, 19 H, CH, CH₂ в скелете, 2H, H(1′), 2H, H(2′) 1H, $$H_{ax}^b(1)$$

); 2.08 (с, 3H, Me (OAc)); 2.24 (д, 1H, H(18), J=11.2); 3.55 (м, 2H, H(3′)); 3.60 (с, 3H, OMe); 4.33 (д, 1H, H(3), J=10.8); 5.23 (т, 1H, H(12) J=3.6)); 7.66-7.85 (м, 15H, Ph). ЯМР ³¹P (161.98 МГц, CDCl₃): δ 24.34.A mixture of iodide 50 (0.21 g, 0.28 mmol) and triphenylphosphine (0.37 g, 1.40 mmol) was boiled in toluene (8 ml) for 16 h (TLC control). After completion of the reaction, the solution was cooled and evaporated in vacuo, the resulting solid was washed with hot hexane (7 ml × 2), dissolved in EtOAc (2 ml) and precipitated with hexane (8 ml). The precipitate was filtered off to give compound 11 (0.24 g, 95%). Mp = 151-153 ° C (EtOH), $${[\alpha ]}_D^{24}+\mathrm{eleven}°$$

(c 0.28, CHCl ₃ ). IR (v / cm ^-1 ): 1726 (C = O). MS, m / z 815.611 [MI] ⁺ . ¹ H NMR (400 MHz, CDCl ₃ , δ, ppm, J / Hz,): 0.51 (t, 1H, $$H_{eq}^a(\mathrm{one})$$

, J = 12.4); 0.71, 0.76, 0.79, 0.89, 1.04 (s, 3H, H (23) -H (27)); 0.88, 0.95 (both d, 3H, H (29), H (30), J = 5.6); 1.27-2.03 (m, 19 H, CH, CH ₂ in the skeleton, 2H, H (1 ′), 2H, H (2 ′) 1H, $$H_{ax}^b(\mathrm{one})$$

); 2.08 (s, 3H, Me (OAc)); 2.24 (d, 1H, H (18), J = 11.2); 3.55 (m, 2H, H (3 ′)); 3.60 (s, 3H, OMe); 4.33 (d, 1H, H (3), J = 10.8); 5.23 (t, 1H, H (12) J = 3.6)); 7.66-7.85 (m, 15H, Ph). ³¹ P NMR (161.98 MHz, CDCl ₃ ): δ 24.34.

Claims (7)

1. The use of triphenylphosphonium salts of lupane and ursane triterpenoids of formula 1-11 as agents with schistosomicidal activity:

2. Triphenylphosphonium salts of lupanic and ursanic triterpenoids of the formula 8-11. 3. A method of producing triphenylphosphonium salts of lupanic and ursanic triterpenoids of the formula 8-11, characterized in that the salts are synthesized by the interaction of triphenylphosphine with betulin diacetate bromide 41, with betulinic acid bromide 43, with methylbetulonate 53 diiodide and methyluronate iodide 50:

4. The method of producing triphenylphosphonium salt 8 according to p. 3, characterized in that it includes the following stages:
a) get ether 40 by acetylation of betulin 12 under the action of Ac ₂ O in pyridine or AcCl in THF in the presence of pyridine and 4-dimethylaminopyridine (DMAP) or directly from birch birch by boiling it in acetic acid for 12 hours:

b) ether 40 is transformed into allyl bromide 41 under the influence of N-bromosuccinimide (NBS) in CCl ₄ when heated at 50 ° C for 4 days:

c) the halide 41 obtained in step 4b is reacted with an excess of triphenylphosphine by boiling in acetonitrile in an argon atmosphere for 4-5 hours to give salt 8 described in paragraph 1. 5. The method of producing triphenylphosphonium salt 9 according to p. 3, characterized in that it includes the following stages:
a) get betulonic acid 54 by oxidation of betulin 12 under the influence of Jones reagent in acetone:

b) receive betulinic acid 13 by reducing betulonic acid 54 NaBH ₄ in a mixture of solvents CHCl ₃ : MeOH = 1: 1, followed by purification of the product by recrystallization from MeOH:

c) betulinic acid 13 is converted into 3β-OAc betulinic acid 42 with Ac ₂ O in pyridine or AcCl in THF in the presence of pyridine and DMAP:

d) acetylated acid 42 is converted to bromine derivative 43 under the influence of N-bromosuccinimide (NBS) in CCl ₄ when heated at 50 ° C for 4 days:

e) the halide 43 obtained in step 5d is reacted with an excess of triphenylphosphine by boiling in acetonitrile in an argon atmosphere for 5 hours to obtain salt 9 described in paragraph 1. 6. The method of producing triphenylphosphonium salt 10 according to p. 3, characterized in that it includes the following stages:
a) get methyl ester of betulonic acid 58 from betulonic acid 54 under the action of diazomethane in ether at room temperature:

b) get 2β-allylbetulonate 51 from methylbetulonate 58 by allyl alkylation under the influence of KN (SiMe ₃ ) ₂ , Et ₃ B and allyl bromide in dimethoxyethane:

c) stereoselectively reduce methyl betulonate 51 under the influence of NaBH ₄ modified CeCl ₃ · 7H ₂ O to obtain 3β-OH-epimer 51a:

d) transform the alcohol 51a into acetate 51b using Ac ₂ O in pyridine or AcCl in THF in the presence of pyridine and DMAP:

e) hydrobonate the double bonds in compound 51b under the influence of BH ₃ · THF or BH ₃ · Me ₂ S followed by the oxidation of the organoboron compound to diol 52 under the influence of 30% H ₂ O ₂ and 10% aqueous NaOH solution:

f) diol 52 iodinated with crystalline iodine in the presence of imidazole and triphenylphosphine in an argon atmosphere in a solution of dry THF at 0 ° C to obtain diiodide 53:

g) the halide 53 obtained in step 6f is reacted with an excess of triphenylphosphine by boiling in toluene or acetonitrile under argon for 18 hours to give the salt 10 described in paragraph 1. 7. The method of producing triphenylphosphonium salt 11 according to p. 3, characterized in that it includes the following stages:
a) ursolic acid 16 is converted to methylursonoate 44 under the action of diazomethane in ether:

b) methylursonoate 44 is oxidized to ursonic acid methyl ester 45 by pyridinium chlorochromate (PCC) in CH ₂ Cl ₂ at 0 ° C:

c) ether 45 is transformed into 2β-allyl substituted ether 46 under the influence of KN (SiMe ₃ ) ₂ , Et ₃ B and allyl bromide in dimethoxyethane at 20 ° C in argon atmosphere:

B
d) ether 46 is subjected to stereoselective reduction under the influence of NaBH ₄ modified with CeCl ₃ · 7H ₂ O to give 3β-OH epimer 47:

e) the transformation of alcohol 47 into acetate 48 is carried out using Ac ₂ O in pyridine or AcCl in THF in the presence of pyridine and DMAP:

f) the double bond of the allyl substituent in compound 48 is hydroborated under the action of BH ₃ · THF or BH ₃ · Me ₂ S followed by oxidation of the organoboron compound to alcohol 49 under the influence of 30% H ₂ O ₂ and a 10% aqueous NaOH solution:

g) alcohol 49 iodinated with crystalline iodine in the presence of imidazole and triphenylphosphine to obtain iodide 50:

h) the halide 50 obtained in step 7g is reacted with an excess of triphenylphosphine by boiling in toluene in argon atmosphere for 16 hours to obtain salt 11 described in paragraph 1.

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