CN101538236B - Method for synthesizing lauroyl-prolyl amino acid methyl ester as novel transdermal enhancer and applications thereof - Google Patents

Abstract

Synthetic method and its application the present invention relates to lauroyl prolyl amino acid methyl ester as novel transdermal enhancer, it is as follows that the novel skin promotees saturating compound.

Description

The lauroyl prolyl amino acid methyl ester is as the synthetic method and the application thereof of novel permeate promotor

Technical field

The present invention relates to a kind of preparation method of novel percutaneous dosing penetration enhancer, with and to lipophilic drugs nimodipine and the salicylic promotion infiltration applications of hydrophilic medicament.Belong to biomedicine field.

Background technology

Over past ten years, be the important research and development object of field of medicaments through the skin absorption system always.Such preparation has been created a kind of simple and conveniently for the treatment of some chronicity diseases and chronic disease and prevention, is the effective administering mode of a class.It avoids first pass effect, improves compliance of patients, reduces administration number of times, improves advantages such as security.

Because percutaneous dosing has plurality of advantages, the research of percutaneous dosing penetration enhancer becomes the focus of pharmacy worker research, as far back as 1970, there is the investigator successfully to use the own phthalein amine of dimethyl alum and dimethyl and promotes the steroid drugs topical and obtained patent; U.S. Nelson company developed azone (Azone) from a series of N-alkyl nitrogen heterocyclic ketone compounds in 1976, and patented in the U.S., and the patent No. is: US3989816, US4301481 and US4316893; In Chinese Pharmacopoeia was listed it in 1987 by China.Have and discover that the N-lauroyl-L-amino acid methyl ester that constitutes with lipid acid and these two kinds of human body endogenous material of amino acid has short preferably saturating performance; wherein; the compound that contains proline(Pro) during as permeate promotor the accumulated concentrations of medicine the highest; the this patent author analyzes because its rigidity five-ring of proline(Pro) combines with hydrophobic lipid acid; structure and azone are similar, so effect is better than other compounds.Also have the investigator to synthesize the compound of two series, the compound of these two series all contains a lactan five-ring and a lactan seven-membered ring, and be combined with hydrophobic afterbody, hydrophobic tail is respectively by ester bond and amido linkage combination, its lactan structure and hydrophobic tail structure are all similar with azone, test-results confirms, the series compound of the hydrophobic tail that connects with ester bond is short functional thoroughly.At present, transdermal enhancer kind commonly used is more and more, as sulfoxide and analogue thereof, azone, lipid acid, alcohols, tensio-active agent, urea, terpene etc.

Summary of the invention

One of purpose of the present invention provides a kind of pentacyclic skin permeate promotor of rigidity and preparation method thereof that contains;

Two of purpose of the present invention provides a kind of preparation method who contains fat-soluble medicine nimodipine and the salicylic salve of water soluble drug;

Three of purpose of the present invention provides and a kind ofly replaces skin to carry out the method for permeate promotor performance evaluation with the synthetic membrane.

Above-mentioned purpose of the present invention is achieved through the following technical solutions:

A kind of structure as shown in the formula have amphipathic and contain the short saturating compound of novel skin of proline(Pro),

Wherein, AA represents 12 kinds of different amino acid, is selected from Ala, Phe, Gly, Ile, Leu, Met, Pro, Ser, Thr, Val, Trp or Tyr.

Lauric acid and proline(Pro) are obtained intermediate C with the method condensation of active ester ₁₂-Pro-OH, this intermediate carry out condensation with 12 kinds of different aminoacids methyl esters again and promptly prepare above-mentioned series compound.

The preparation method of The compounds of this invention may further comprise the steps:

(1) lauric acid and proline(Pro) are with the method synthetic intermediate C of active ester ₁₂-Pro-OH;

(2) with intermediate C ₁₂-Pro-OH respectively with Ala-OMe, Phe-OMe, Gly-OMe, Ile-OMe, Leu-OMe, Met-OMe, Pro-OMe, Ser-OMe, Thr-OMe, Val-OMe, Trp-OMe or Tyr-OMe condensation promptly get respective compound.

The present invention includes the pharmaceutical composition that contains above-claimed cpd.Wherein contain medicine, above-claimed cpd and medicine acceptable carrier.Pharmaceutical composition of the present invention wherein contains above-mentioned compound 0.01%-99.99%, and the drug weight percentage composition is 0.1%-5%, and all the other are the medicine acceptable carrier.

Pharmaceutical composition of the present invention is selected from ointment, Eye ointments, gelifying agent or suppository.Its Chinese traditional medicine is selected from externally applied medicine.Described externally applied medicine is selected from fat-soluble medicine or water soluble drug.Wherein fat-soluble medicine is selected from nimodipine, and water soluble drug is selected from Whitfield's ointment.

The present invention also comprises the application of compound of the present invention in preparation medicine, healthcare products, agricultural chemicals and makeup.

Mainly use animal skin and synthetic skin medium at present in the research of Transdermal absorption as simulated skin, though it is more extensive that animal skin is used, but reason owing to animal individual difference, judgement criteria for the short performance of permeate promotor can not well be unified, and the permeability of animal skin has been compared certain gap with fell; At present, the synthetic skin has obtained researchist's concern more and more, the employed synthetic membrane of this patent---polysulfone membrane (Pall, 66221) has been carried out research to the ointment that contains the hydrogenation cortisone and has been used on the collaborative project of Hanson company and U.S. FDA.

Compound of the present invention is compared with prior art and is had the following advantages:

All compounds constitute by endogenous material (amino acid and lipid acid), and are almost non-toxic, and nonirritant is suitable for the medium-term and long-term use of percutaneous drug administration preparation and makeup.

Description of drawings:

Fig. 1 is nimodipine group Q-t figure

Fig. 2 is Whitfield's ointment group Q-t figure

Embodiment

In order further to set forth the present invention, provide a series of embodiment below.These embodiment are illustrative fully, and they only are used for the present invention is specifically described, and not should be understood to limitation of the present invention.

Embodiment 1 C ₁₂The preparation of-Pro-OH

Under the condition of ice bath, 1.49g (10mmol) N-hydroxy-succinamide (HOSu) and 2.0g (10mmol) lauric acid stir in 20ml anhydrous tetrahydro furan (THF); 2.67g (13mmol) being dissolved in the anhydrous tetrahydro furan of dicyclohexylcarbodiimide (DCC) is added dropwise in the reaction solution.React filtration under diminished pressure removal dicyclohexylurea (DCU) (DCU) back ethyl acetate (EA) dissolving after 6 hours, use saturated NaHCO behind the filtration under diminished pressure DCU ₃Solution is washed 3 times, adds anhydrous Na after saturated NaCl solution is washed 3 times ₂SO ₄Drying is spent the night.Filter and remove Na ₂SO ₄After be evaporated to dried.This product dissolves with anhydrous THF, adds Pro, regulates the pH value to 8-9 with N-methylmorpholine (NMM), reacts 6 hours.Behind the reaction solution concentrating under reduced pressure, use saturated KHSO ₄Solution dissolving, and use the EA extracting twice is behind the merging EA layer, with saturated NaCl solution extraction 1 time.EA layer anhydrous Na ₂SO ₄Dried overnight is filtered and is removed NaSO ₄, solution decompression is concentrated into dried, obtains intermediate C ₁₂-Pro-OH, it is standby to put into moisture eliminator.

Embodiment 2 C ₁₂The preparation of-Pro-Gly-OMe

Under the condition of ice bath, 2.97g intermediate lauroyl proline (C12-Pro-OH) is dissolved with anhydrous THF, add 1.35g t (10mmol) I-hydroxybenzotriazole (HOBt), 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, add 1.25g (10mmol) Gly-OMe, regulate pH value to 9 with NMM, reacted 10 hours, be evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is respectively washed 3 times with the saturated NaHCO3 aqueous solution, saturated NaCl solution, saturated KHSO4 solution and saturated NaCl solution successively.The EA layer with anhydrous Na 2SO4 drying, filtration, filtrate decompression concentrate target compound C12-Pro-Gly-OMe crude product.With leaving standstill crystallization behind the petroleum ether dissolution, obtain the white powder solid.ESI-MS(m/z)369[M+H]+，737[2M+H]+， $759[2M+\mathrm{Na}]+{\left[\mathrm{\α}\right]}_D^{20}=-49.8333$ mp.45.4-45.6℃

Embodiment 3 C ₁₂The preparation of-Pro-Phe-OMe

Under the condition of ice bath, with 2.97g intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 2.15 (10mmol) Phe-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Phe-OMe crude product.Column chromatography for separation obtains the white powder title compound.ESI-MS(m/z)459[M+H] ⁺，481[M+Na] ⁺， $940{[2M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=-34.1667$ mp.44.4-45-0℃

Embodiment 4 C ₁₂The preparation of-Pro-Ile-OMe

Under the condition of ice bath, with 2.97g intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 1.81g (10mmol) Ile-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Ile-OMe crude product.Column chromatography for separation obtains water white transparency oily title compound.ESI-MS(m/z)425[M+H] ⁺， $447{[M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=-50.7333$

Embodiment 5 C ₁₂The preparation of-Pro-Leu-OMe

Under the condition of ice bath, with 2.97g intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 1.81g (10mmol) Leu-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Leu-OMe crude product.Column chromatography for separation obtains white wax shape title compound.ESI-MS(m/z)425[M+H] ⁺，849[2M+H] ⁺， $869{[2M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=-57.8788$ mp.41.8-42.2℃

Embodiment 6 C ₁₂The preparation of-Pro-Pro-OMe

Under the condition of ice bath, with 2.97g intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 1.61g (10mmol) Pro-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Pro-OMe crude product.Column chromatography for separation obtains white wax shape title compound.ESI-MS(m/z)409[M+H] ⁺， $431{{[M+\mathrm{Na}]}^{+}\left[\mathrm{\α}\right]}_D^{20}=71.1111$ mp.26.1-26.6.℃

Embodiment 7 C ₁₂The preparation of-Pro-Met-OMe

Under the condition of ice bath, with 2.97g intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 1.99g (10mmol) Met-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Met-OMe crude product.Column chromatography for separation obtains white wax shape title compound.ESI-MS(m/z)443[M+H] ⁺，462[M+Na] ⁺，? $907{[2M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=90.2222$ mp.21.2-22.2℃

Embodiment 8 C ₁₂The preparation of-Pro-Ser-OMe

Under the condition of ice bath, with 2.97g (10mmol) intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 1.99g (10mmol) Ser-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Ser-OMe crude product.Column chromatography for separation obtains white wax shape title compound.ESI-MS(m/z)339[M+H] ⁺，421[M+Na] ⁺， ${820{[2M+\mathrm{Na}]}^{+}\left[\mathrm{\α}\right]}_D^{20}=-20.7576$ mp.30-31℃

Embodiment 9 C ₁₂The preparation of-Pro-Thr-OMe

Under the condition of ice bath, with 2.97g (10mmol) intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 1.69g (10mmol) Thr-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Thr-OMe crude product.Column chromatography for separation obtains the white powder title compound.ESI-MS(m/z)413[M+H] ⁺，435[M+Na] ⁺， $848{[2M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=-52.5926$ mp.83.4-84.3℃

Embodiment 10 C ₁₂The preparation of-Pro-Tyr-OMe

Under the condition of ice bath, with 2.97g (10mmol) intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 2.31g (10mmol) Tyr-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Tyr-OMe crude product.Column chromatography for separation obtains the white powder title compound.ESI-MS(m/z)475[M+H] ⁺，497[M+Na] ⁺?， $972{[2M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=-40.4016$ mp.131.3-132.2℃

Embodiment 11 C ₁₂The preparation of-Pro-Trp-OMe

Under the condition of ice bath, with 2.97g (10mmol) intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 2.54g (10mmol) Trp-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Trp-OMe crude product.Crude product leaves standstill after with acetone solution, and the white needle-like crystals of separating out is title compound. $\mathrm{ESI}-\mathrm{MS}(m/z)520{[M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=7.2727$ mp.44.6-45.4℃

Embodiment 12C ₁₂The preparation of-Pro-Val-OMe

Under the condition of ice bath, with 2.97g (10mmol) intermediate lauroyl proline (C ₁₂-Pro-OH) with anhydrous THF dissolving, adding 1.35g (10mmol) HOBt, 2.67g (13mmol) DCC reacted after 6 hours, filtered DCU, added 1.67g (10mmol) Val-OMe, regulated pH value to 9 with NMM, reacted 10 hours, was evaporated to dried.With the EA dissolving, filter and remove DCU, the solution that obtains is used saturated NaHCO successively ₃The aqueous solution, saturated NaCl solution, saturated KHSO ₄Solution and saturated NaCl solution are respectively washed 3 times.EA layer anhydrous Na ₂SO ₄Drying, filtration, filtrate decompression concentrate target compound C ₁₂-Pro-Val-OMe crude product.Column chromatography for separation obtains white title compound.ESI-MS(m/z)410[M+Na] ⁺， $797{[2M+\mathrm{Na}]}^{+}{\left[\mathrm{\α}\right]}_D^{20}=-28.3951$ mp.47.3-47.7℃

Embodiment 14 contain the nimodipine salve preparation

Accurately take by weighing nimodipine powder 200mg, add white vaseline 10g, be heated to 60 ℃, the Vaseline dissolving stirs, and obtains the blank ointment of nimodipine.Take by weighing the blank ointment 3g of nimodipine, add the disclosed transdermal enhancer of 0.1067mmol this patent, stir, obtain containing the nimodipine ointment of permeate promotor.

Embodiment 15 contains the preparation of Whitfield's ointment salve

Accurately take by weighing Whitfield's ointment powder 200mg, add white vaseline 10g, be heated to 60 ℃, the Vaseline dissolving stirs, and obtains the blank ointment of Whitfield's ointment.Take by weighing the blank ointment 3g of Whitfield's ointment, add the disclosed transdermal enhancer of 0.1067mmol this patent, stir, obtain containing the Whitfield's ointment ointment of permeate promotor.

Embodiment 16 contains the short saturating performance evaluation of nimodipine salve of different permeate promotors

The Franz diffusion cell experiment of the application choice classics of transdermal enhancer effect.At first,, be immersed in the dehydrated alcohol 15 hours, carry out pre-treatment polysulfone membrane (Pall company, 66221).Between the reception tank and supply pool of this polysulfone membrane as for the Franz diffusion cell, receiving liquid is 30% ethanol, and supply pool is nimodipine ointment or the blank nimodipine ointment that contains permeate promotor.Bath temperature is 32 ± 0.5 ℃, and stirring velocity is 600rpm, and the sampling interval is 1h, 2h, 3h, 4h, 5h, 6h, 10h, 16h, 24h.Flushing pipeline 1.5ml during sampling, sample volume 1ml.Resulting sample solution is detected quantitatively with HPLC, calculate with formula and obtain accumulative total osmotic concentration and Q-T curve.As table 1, Fig. 1.

Embodiment 17 contains the short performance evaluation of the Whitfield's ointment salve of different permeate promotors

The Franz diffusion cell experiment of the application choice classics of transdermal enhancer effect.At first,, be immersed in the dehydrated alcohol 15 hours, carry out pre-treatment polysulfone membrane (Pall company, 66221).Between the reception tank and supply pool of this polysulfone membrane as for the Franz diffusion cell, reception liquid is distilled water, and supply pool is Whitfield's ointment ointment or the blank Whitfield's ointment ointment that contains permeate promotor.Bath temperature is 32 ± 0.5 ℃, and stirring velocity is 600rpm, and the sampling interval is 1h, 2h, 3h, 4h, 5h, 6h, 10h, 16h, 24h.Flushing pipeline 1.5ml during sampling, sample volume 1ml.Resulting sample solution is detected quantitatively with HPLC, calculate with formula and obtain accumulative total osmotic concentration and Q-T curve.As table 2, Fig. 2.

Table 1 nimodipine group accumulative total osmotic concentration and and blank group ratio

^*The expression blank group of contrast (0 group) has significant difference P＜0.05

* represents to contrast blank group (0 group) utmost point significant difference P＜0.01

Table 2 Whitfield's ointment group accumulative total osmotic concentration and and blank group ratio

^*The expression blank group of contrast (0 group) has significant difference P＜0.05

* represents to contrast blank group (0 group) utmost point significant difference P＜0.01

Claims (10)

1. A structure as shown in the formula have amphipathic and contain the short saturating compound of novel skin of proline(Pro),

   

   Wherein, AA represents 12 kinds of different amino acid groups, is selected from Ala, Phe, Gly, Ile, Leu, Met, Pro, Ser, Thr, Val, Trp or Tyr group.
2. 2. the preparation method of the described compound of claim 1 may further comprise the steps:

   (1) lauric acid and proline(Pro) are with the method synthetic intermediate C of active ester ₁₂-Pro-OH;

   (2) with intermediate C ₁₂-Pro-OH respectively with Ala-OMe, Phe-OMe, Gly-OMe, Ile-OMe, Leu-OMe, Met-OMe, Pro-OMe, Ser-OMe, Thr-OMe, Val-OMe, Trp-OMe or Tyr-OMe condensation promptly get respective compound.
3. 3. the pharmaceutical composition that contains claim 1 compound.
4. 4. the pharmaceutical composition of claim 3 wherein contains medicine, described compound of claim 1 and medicine acceptable carrier.
5. 5. the pharmaceutical composition of claim 3, wherein containing the described compound weight percentage of claim 1 is 1%-10%, and the drug weight percentage composition is 0.1%-5%, and all the other are the medicine acceptable carrier.
6. 6. the pharmaceutical composition of claim 5 is selected from ointment, gelifying agent or suppository.
7. 7. the pharmaceutical composition of claim 3, its Chinese traditional medicine is selected from externally applied medicine.
8. 8. the pharmaceutical composition of claim 7, wherein externally applied medicine is selected from fat-soluble medicine or water soluble drug.
9. 9. the pharmaceutical composition of claim 8, wherein fat-soluble medicine is selected from nimodipine, and water soluble drug is selected from Whitfield's ointment.
10. 10. the application of the compound of claim 1 in preparation medicine, healthcare products, agricultural chemicals and makeup.

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