CN106458857A - Crystalline free acid, hemicalcium salt and alfa-phenylethylamine salt of ahu-377 as well as preparation method therefor and application thereof - Google Patents

Abstract

Disclosed is a crystalline form of NEP inhibitor AHU-377, including crystalline free acid, hemi-calcium salt, α-phenethylamine salt and its preparation method and application. The physical and chemical properties of AHU‑377 are improved, the process is mature, the operability is strong, the quality of the obtained product is high, uniform and stable, the chemical stability is improved, and it is convenient for storage.

Description

AHU-377 crystal types free acid, half calcium salt, α ﹣ phenyl ethylamine salt and its preparation method and application Technical field

The invention belongs to technical field of pharmaceuticals, and in particular to a kind of nep inhibitor AHU-377 crystal types free acid, half calcium salt, α ﹣ phenyl ethylamine salt and its preparation method and application.

Background technology

Neutral endopeptidase (EC 3.4.24.11；Enkephalinase；Must peptase；NEP it is) a kind of the metalloproteinases for including zinc of various peptide substrates can be cracked on the amino terminal of aromatic amino acid.The substrate of this enzyme includes atrial natriuretic peptide (ANF, also referred to as ANP), brain natriuretic peptide (BNP), met and leu enkephalins, bradykinin, neurokinin A and Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH2 without limitation.

ANF is family's vasodilation, diuresis and antihypertensive peptides, a kind of form --- ANF 99-126 are the circulation peptide hormones discharged in cardiac enlargement situation by heart.ANF function is the homeostasis and regulation blood pressure for maintaining salt and water.ANF is in the circulating cycle by least two process rapid inactivations：The removing of acceptor-mediation and the enzyme-deactivating carried out in NEP.Nep inhibitor enhances experimental animal and carries out the low blood pressure after pharmacology ANF injections, diuresis, promotees natruresis and plasma ANF response.The ANF carried out by two kinds of specific nep inhibitors enhancing, 1988, generically discloses and can strengthen ANF with NEP in United States Patent (USP) US4749688.Thiorphan (thiorphan) and kelatorphan are disclosed in the same year, United States Patent (USP) US4740499 can be used for strengthening the application of atrial natriuretic peptide.In addition, nep inhibitor can reduce blood pressure and play the effect of diuresis and increase cyclic guanosine 3 ', 5 '-monophosphate (cGMP) excretion of the ANF- samples effect such as in some form of experimental hypertension.Because ANF antibody will offset the reduction of blood pressure, the antihypertensive function of nep inhibitor is mediated by ANF.Most cause for a long time and at last the various pathological changes of the target organ such as heart and kidney without the hypertensive vascular disease of control.Lasting hypertension can result in the incidence increase of apoplexy.Therefore, need to be estimated effect of antihypertensive therapy strongly, i.e., to being reduced except blood pressure in addition to other cardiovascular endpoints events checked to be further discovered that the benefit of therapeutic alliance.

1993, Gary Ksander etc. disclose the 4-Aminobutanoicacid derivative of a class biaryl substituted in United States Patent (USP) US5217996, such compound is found to have obvious NEP inhibitory activity, wherein a most representational compound is N- (3- carboxyl -1- oxopropyls)-(4S)-p- phenylphenylmethyl) -4- amino -2R- methylbutanoic acid ethyl esters (also known as AHU-377).

AHU-377 generally occurs with grease in actual production preparation process, is unfavorable for preserving and Subsequent pharmacological exploitation.AHU-377 crude products are esterified through the tert-butyl alcohol, the AHU-377 sodium salts obtained after sodium hydroxide basic hydrolysis coherent condition after dichloromethane/n-hexane recrystallization is failed to understand, and find that AHU-377 sodium salts hygroscopicity is quite big through inventor's further investigation, material is unstable and is unfavorable for storage, therefore, AHU-377 is used as active constituents of medicine, its product quality, material stability, there is very big problem in storage or amount etc., its sodium salt does not also solve the above problems, although United States Patent (USP) US5217996 and WO2007056546, which disclose AHU-377, can also be prepared into various salt type compounds, such as alkali metal salt, alkali salt, amine salt, alkylamine salt, hydroxyalkyl amine salt etc., but, infer mainly or from the chemical constitution that AHU-377 contains a carboxyl and draw, various other salt type compounds in addition to sodium salt are not actually obtained, AHU-377 poor product qualities can not be solved, material is unstable, the problems such as being difficult to store and weigh.

Therefore, in view of the problem of prior art is present, it is necessary to a kind of free acid coherent condition of stabilization is developed to AHU-377, the need for meeting drug development.

In addition, the application of the medicine is the patient for having hypertension and heart failure class angiocardiopathy, the intake of sodium can also increase cardiovascular burden, therefore extremely be necessary sodium salt being replaced with the salt form without sodium ion, or other classes to cardiovascular harmless or beneficial salt.Therefore, in view of the problem of prior art is present, it is necessary to a kind of more stable free acid coherent condition is developed for AHU-377 or its sodium salt, the need for meeting drug development.

Calcium ion is accepted extensively to cardiovascular beneficial effect, and the effect of calcium ion hypotensive may be caused by following mechanism：1st, the membrane stabilizing action of calcium, calcium is combined on cell membrane, it is possible to decrease permeability of cell membrane, is improved threshold of excitability, can be made vascular smooth muscle relaxation；2nd, calcium itself can block calcium channel, prevent extracellular calcium ion intracellular from entering；3rd, blood calcium height can resist row's potassium increase of caused by high natrium, and potassium ion plays an important role to stabilizing cell membrane.People maintains enough high calcium intakes, can resist the illeffects of high sodium；4th, scholar thinks, 40% blood pressure rise is relevant with parathyroid gland.Parathyroid gland can produce a kind of high heat-proof peptide material, and this is the arch-criminal for causing hypertension, be referred to as " the hypertension induced factor ".The generation of " the hypertension induced factor " is stimulated by low calcium diet, and high calcium diet can suppress it and produce " the hypertension induced factor ".Modern study proves that the elderly, and the food for eating rich calcium can be hardened with prevention of arterial, and hypertensive patient coordinates while depressor is taken to replenish the calcium, moreover it is possible to too high blood pressure is dropped to normally.So, for the people of calcium deficiency, supplement calcium in large quantities, makes blood calcium keep normal level, the balance of calcium metabolism inside and outside smooth muscle cell is maintained, it is to avoid the spasm of vascular smooth muscle, so that the purpose for reaching effective prevention of arterial vascular sclerosis, reducing blood pressure for a long time.Therefore, will AHU-377 sodium salt changes calcium salt into, can not only reduce the side effect that sodium salt is brought, and can also bring the extra beneficial effect to angiocardiopathy.

Although WO2008031567 also discloses that AHU-377 free acids are prepared into the synthetic route of calcium salt, as shown below：

But, those skilled in the art will be seen that, it is above-mentioned common to reflect that technology path is the general technology route into salt based on the compound containing carboxyl and calcium salt into salt, whether but the technology path can not be adapted to all carboxyl compounds, AHU-377 calcium salt can be prepared by this reflection by also having no way of finding out about it.The technology path is supported without embodiment, so whether the more unknowable technology path can produce the calcium salt of crystal formation.In addition, the technology path is set up only under specific condition, especially second step in synthetic route is AHU-377 sodium salts and CaCl₂The ion-exchange reactions of generation, if the specific condition that AHU-377 calcium salts or sodium chloride are separated out from system can not be created, said synthesis route is then invalid.Secondly, from the general technology path, it is impossible to which whether supposition can prepare the AHU-377 calcium salts of solid form, and whether gained calcium salt is crystal formation, is which kind of crystal formation.Therefore, the general technology path can not obtain the AHU-337 crystals of calcium salts of AHU-377 calcium salts or crystal form.

The content of the invention

In order to solve the problem of prior art is present, improve AHU-377 physicochemical properties, such as improved crystallinity increases dissolubility, reduces hygroscopicity, strengthens process operability, improves chemical stability etc..Inventor have developed a kind of AHU-377 crystal types free acid, substantially improve the physicochemical property of AHU-377 free acids, such as product quality, dissolubility, hygroscopicity, chemical stability on the basis of existing technology by further investigation.Meanwhile, inventor has prepared the calcium salt crystal formations of AHU-377 half and AHU-377 α-phenylethylamine salt crystal formation on the basis of further investigation, and (α-phenylethylamine, molecular formula is C₈H₁₁N, α-phenylethylamine has two stereoisomers：(R)-(+)-α-phenylethylamine and (S)-(-)-α-phenylethylamine).Change the physicochemical property of AHU-377 free acids or its sodium salt by the way that AHU-377 is prepared into calcium salt or α-phenylethylamine salt, such as crystallinity, dissolubility, hygroscopicity.And solve sodium salt bring cardiovascular side effects the problem of, especially treatment heart failure with depression complication, have very wide prospect.

First aspect present invention provides a kind of AHU-377 crystal types free acid (being appointed as crystal formation I), its x-ray powder diagram includes being located at 23.70 ± 0.2 °, 15.84 ± 0.2 °, 20.84 ± 0.2 ° and 8.56 ± 0.2 °, the angle of diffraction (2 θ) place peak.

As further preferred scheme, preferably its X-ray powder diffraction figure also includes the peak positioned at 17.06 ± 0.2 °, 25.98 ± 0.2 °, 18.01 ± 0.2 °, 9.42 ± 0.2 ° and 26.91 ± 0.2 ° angle of diffraction (2 θ) place.

It is used as scheme still more preferably, it is preferred that its X-ray powder diffraction figure further comprises positioned at 34.70 ± 0.2 °, 21.83 ± 0.2 °, 25.69 ± 0.2 °, the peak at 25.33 ± 0.2 °, 11.67 ± 0.2 ° and 26.24 ± 0.2 ° angle of diffraction (2 θ) place.

As scheme still more preferably, the peak at the most preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 1 is substantially the same, and its X-ray powder diffraction data is as shown in table 1：

Table 1

2θ(°)	Intensity %	2θ(°)	Intensity %
				8.56	44.7	20.84	66.5
9.42	13.6	21.83	8.5
				11.42	6.6	23.70	100
11.67	6.8	25.33	7.9
				15.43	6.1	25.69	8.1
15.84	70.7	25.98	31.9
				17.06	41.8	26.24	6.8
18.01	18.6	26.91	9.6
				18.69	5.4	29.47	5.9
19.12	6.5	34.70	9

Term " substantially the same " used herein on X-ray diffraction peak position means to consider typical peak position and intensity variable.For example, it will be understood by those skilled in the art that peak position (2 θ) will be different due to XRPD instruments, and causing measured value to be varied from, this change is up to up to 0.2 ° sometimes sometimes.Further, it will be understood by those skilled in the art that XRPD sample method for making sample, XRPD instruments, sample crystallinity, the factor such as amount of samples and crystal preferred orientation will cause the change of relative peak intensities in sample XRPD diffraction patterns.

Second aspect of the present invention provides a kind of preparation method of AHU-377 crystal types free acid, comprises the following steps,

1) AHU-377 free acids are dissolved in suitable organic solvent；

2) solution system temperature is reduced, and/or adds crystal seed, and/or adds appropriate anti-solvent to solution and muddy, crystallization occurs；

3) separation of solid and liquid obtains AHU-377 crystal type free acids.

As further preferred scheme, comprise the following steps,

1) AHU-377 free acids are dissolved under normal temperature or heated condition in suitable organic solvent；

2) reduce step 1) in solution system temperature, and/or add crystal seed, crystallization；

3) separation of solid and liquid obtains AHU-377 crystal type free acids.

As further preferred scheme, comprise the following steps,

1) AHU-377 free acids are dissolved in suitable organic solvent；

2) by step 1) in solution mixed with anti-solvent；

3) separation of solid and liquid obtains AHU-377 crystal type free acids.

It is preferred that, step 2) in mixed with anti-solvent before AHU-377 crystal type free acid crystal seeds are previously added in anti-solvent.

Described " mixing " refers to step 1) in solution be added drop-wise among anti-solvent or anti-solvent be added drop-wise to step 1) in solution among.

As scheme still more preferably, step 1 in above-mentioned preparation method) the suitable organic solvent is selected from ethers, cyclic ethers class, esters, halogenated alkane, benzene class organic solvent or its mixture, including but not limited to following solvent：From ethyl acetate, isopropyl acetate, dichloromethane, trichloroethanes, carbon tetrachloride, methyl tertiary butyl ether(MTBE), isopropyl ether, benzene,toluene,xylene or its composition, isopropyl acetate, dichloromethane, methyl tertiary butyl ether(MTBE), isopropyl ether or its composition preferably are selected from.

As scheme still more preferably, step 2 in above-mentioned preparation method) anti-solvent refers to the less organic solvent of polarity, including but not limited to following solvent：Normal heptane, n-hexane, isooctane, pentane, hexamethylene, pentamethylene, ether or its composition；It preferably is selected from normal heptane, n-hexane, hexamethylene, ether or its composition.

It is used as further preferred scheme, " dissolving " refers to the operation of one of ordinary skill in the art typically in above-mentioned preparation method, generally can appropriate heating make AHU-377 free acids dissolution of raw material or dissolved clarification, or the consumptions of (20~25 DEG C) increasing solvents makes AHU-377 free acids dissolution of raw material or dissolved clarification at normal temperatures.

As scheme still more preferably, solvent is preferably heated to 40~80 DEG C by the appropriate heating makes AHU-377 free acids dissolution of raw material or dissolved clarification.

As scheme still more preferably, the addition of the preferred recrystallisation solvent of consumption of the increasing solvent is 5-20 times of volume mass ratio of AHU-377 free acids, makes AHU-377 free acids dissolution of raw material or dissolved clarification.

It is used as further preferred scheme, step 2 in above-mentioned preparation method) it is described addition crystal seed refer in AHU-377 free acid solutions add homology crystalline solid, add crystal seed amount to be preferred with the 0.1-20.0% of AHU-377 free acid mass ratioes, the 0.5-5.0% using mass ratio is optimal.It is appreciated that those skilled in the art can also further increase the addition of crystal seed, the technique effect that this technology can reach can be equally reached as added in batches, same crystal is obtained.Therefore, the technical scheme for adding crystal seed is modified or equivalent substitution, it all should cover within the spirit of the present invention.

It is used as further preferred scheme, step 3 of the present invention) obtained by AHU-377 crystal types free acid its x-ray diffractogram of powder include being located at 23.70 ± 0.2 °, 15.84 ± 0.2 °, 20.84 ± 0.2 ° and 8.56 ± 0.2 °, the angle of diffraction (2 θ) place peak.

Third aspect present invention provides a kind of calcium salt crystal formations of AHU-377 half, and its X-ray powder diffraction figure includes the peak positioned at 12.70 ± 0.2 °, 7.32 ± 0.2 °, 15.90 ± 0.2 ° and 18.56 ± 0.2 ° angle of diffraction (2 θ) place, or Its X-ray powder diffraction figure includes the peak positioned at 4.02 ± 0.2 °, 3.62 ± 0.2 ° and 17.82 ± 0.2 ° angle of diffraction (2 θ) place.

Fourth aspect present invention provides a kind of calcium salt trihydrate crystal formations (being appointed as crystal formation I) of AHU-377 half, its X-ray powder diffraction figure includes being located at 12.70 ± 0.2 °, the peak at 7.32 ± 0.2 °, 15.90 ± 0.2 ° and 18.56 ± 0.2 ° angle of diffraction (2 θ) place.

As further preferred scheme, preferably its X-ray powder diffraction figure also includes the peak positioned at 14.74 ± 0.2 °, 15.42 ± 0.2 °, 7.82 ± 0.2 °, 19.42 ± 0.2 °, 16.64 ± 0.2 ° and 17.54 ± 0.2 ° angle of diffraction (2 θ) place.

As scheme still more preferably, the peak at the most preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 4 is substantially the same, and its X-ray powder diffraction data is as shown in table 2：

Table 2

2θ(°)	Intensity %	2θ(°)	Intensity %
				7.32	39	15.90	37.3
7.82	15.1	16.64	13.4
				12.70	100	17.54	11.6
14.74	27.3	18.56	33.5
				15.42	18.4	19.42	13.5

Fifth aspect present invention provides a kind of calcium salt anhydrous crystal forms (being appointed as crystal formation II) of AHU-377 half, and its X-ray powder diffraction figure includes the peak positioned at 4.02 ± 0.2 °, 3.62 ± 0.2 ° and 17.82 ± 0.2 ° angle of diffraction (2 θ) place.

As further preferred scheme, preferably its X-ray powder diffraction figure also includes the peak positioned at 20.48 ± 0.2 °, 14.50 ± 0.2 ° and 19.66 ± 0.2 ° angle of diffraction (2 θ) place.

As scheme still more preferably, the peak at the most preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 6 is substantially the same, and its X-ray powder diffraction data is as shown in table 3：

Table 3

2θ(°)	Intensity %	2θ(°)	Intensity %
				3.62	58.2	17.82	36.8
4.02	100	19.66	13.8
				14.50	25.8	20.48	29

Sixth aspect present invention provides a kind of preparation method of the calcium salt trihydrate crystal formations of AHU-377 half, including,

1) AHU-377 free acids are dissolved or are suspended in a suitable solvent, mixed the corresponding salt of preparation generation AHU-377 with equivalent or the excessive caustic alkali or alkalescent sodium salt or sylvite for dissolving in the system, preferably mix alkalescent sodium salt or sylvite；

2) above-mentioned AHU-377 salt systems and water-soluble Ca salt are mixed in water or aqueous systems containing organic solvent, produces AHU-377 calcium precipitations；

3) product obtains the calcium salt trihydrate crystal formations of AHU-377 half during collecting above-mentioned salt-forming reaction.

It is used as further preferred scheme, step 1) the suitable solvent is recrystallisation solvent, including water, water-soluble solvent or its mixture.

It is used as scheme still more preferably, the water-soluble solvent is selected from alcohols, ketone, cyclic ethers class, amide-type, sulfoxide type organic solvent or its mixture, it preferably is selected from methanol, ethanol, normal propyl alcohol, isopropanol, n-butanol, acetonitrile, acetone, MEK, tetrahydrofuran, dioxane, DMF, dimethyl sulfoxide (DMSO) or its mixture；

Be used as further preferred scheme, step 1) caustic alkali be sodium hydroxide or potassium hydroxide.

Be used as further preferred scheme, step 1) the alkalescent sodium salt be selected from sodium carbonate, sodium acid carbonate, sodium acetate, sodium formate, sodium propionate, PAA, sodium benzoate or its mixture；It is preferred that sodium acid carbonate, sodium acetate or its mixture；Step 1) the alkalescent sylvite be selected from potassium carbonate, saleratus, potassium acetate, potassium formate, potassium propionate, potassium acrylate, Potassium Benzoate or its mixture；Potassium bicarbonate, potassium acetate or its mixture；

Be used as further preferred scheme, step 2) water-soluble Ca salt be selected from calcium chloride, calcium bromide, calcium iodide, calcium nitrate, calcium chlorate, calcium hypochlorite, Calcium perchlorate, calcium lactate, calcium gluconae or its mixture；

Be used as scheme still more preferably, step 2) water-soluble Ca salt be selected from calcium chloride, calcium chlorate or its mixture.

It is used as further preferred scheme, the dissolving refers to the operation of one of ordinary skill in the art typically, generally can appropriate heating make AHU-377 free acids dissolution of raw material or dissolved clarification, or increase the consumption of solvent to make AHU-377 free acids dissolution of raw material or dissolved clarification.

As scheme still more preferably, the positive solvent of crystallization is preferably heated to 30~80 DEG C and makes AHU-377 free acids dissolution of raw material or dissolved clarification by the appropriate heating.

As scheme still more preferably, the addition of the preferred recrystallisation solvent of consumption of the increasing solvent is 2-20 times of volume mass ratio of AHU-377 free acids, makes AHU-377 free acids dissolution of raw material or dissolved clarification.

It is used as further preferred scheme, step 3) described in the above-mentioned salt-forming reaction of collection during product the step of can be specifically directly to collect the solid product that is separated out in the above-mentioned course of reaction into calcium salt or by solvent flashing or add anti-solvent or directly cooling crystallization or add crystal seed, obtain AHU-377 calcium salt polymorphics.

As scheme still more preferably, the anti-solvent refers to the less organic solvent of polarity, including but not limited to following solvent：Normal heptane, n-hexane, isooctane, pentane, hexamethylene, pentamethylene, ether or its composition.The amount for adding anti-solvent is 1-10 times of volume ratio of recrystallisation solvent.

It is used as further preferred scheme, step 3 of the present invention) obtained by AHU-377 calcium salt crystal formation I its x-ray diffractogram of powder include being located at 12.70 ± 0.2 °, the peak at 7.32 ± 0.2 °, 15.90 ± 0.2 ° and 18.56 ± 0.2 ° angle of diffraction (2 θ) place.

Seventh aspect present invention provides a kind of preparation method of the calcium salt anhydrous crystal forms of AHU-377 half, and the calcium salt trihydrates of AHU-377 half are heated into more than 50 DEG C, its dehydration is transformed into anhydrous crystal forms and obtains the calcium salt anhydrous crystal forms of AHU-377 half.

As further preferred scheme, the heating is the operation of one of ordinary skill in the art typically, can generally be heated up under nitrogen protection or under vacuum condition.

As further preferred scheme, the heating temperature range is at 50-100 DEG C.

It is used as further preferred scheme, its X-ray powder diffraction figure of AHU-377 calcium salt anhydrous crystal forms (crystal formation II) obtained by the present invention includes the peak at the angle of diffraction (2 θ) place positioned at 4.02 ± 0.2 °, 3.62 ± 0.2 ° and 17.82 ± 0.2 °.

As further preferred scheme, preferably its X-ray powder diffraction figure also includes the peak positioned at 20.48 ± 0.2 °, 14.50 ± 0.2 ° and 19.66 ± 0.2 ° angle of diffraction (2 θ) place.

Eighth aspect present invention provides a kind of AHU-377 α-phenylethylamines salt, its X-ray powder diffraction figure includes being located at 20.58 ± 0.2 °, 24.28 ± 0.2 °, the peak at 8.38 ± 0.2 ° and 23.20 ± 0.2 ° angle of diffraction (2 θ) place, or including positioned at 23.28 ± 0.2 °, the peak at 18.9 ± 0.2 °, 13.7 ± 0.2 ° and 14.72 ± 0.2 ° angle of diffraction (2 θ) place.

Ninth aspect present invention provides a kind of AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation (being appointed as crystal formation I), its X-ray powder diffraction figure includes being located at 20.58 ± 0.2 °, the peak at 24.28 ± 0.2 °, 8.38 ± 0.2 ° and 23.20 ± 0.2 ° angle of diffraction (2 θ) place.

As further preferred scheme, its X-ray powder diffraction figure also includes the peak positioned at 19.36 ± 0.2 °, 15.16 ± 0.2 °, 16.78 ± 0.2 °, 18.84 ± 0.2 ° and 22.06 ± 0.2 ° angle of diffraction (2 θ) place.

As scheme still more preferably, its X-ray powder diffraction figure also includes the peak positioned at 17.34 ± 0.2 °, 7.52 ± 0.2 °, 28.10 ± 0.2 °, 29.66 ± 0.2 °, 28.94 ± 0.2 ° and 10.96 ± 0.2 ° angle of diffraction (2 θ) place.

As scheme still more preferably, the peak at the most preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 7 is substantially the same, and its X-ray powder diffraction data is as shown in table 4：

Table 4

2θ(°)	Intensity %	2θ(°)	Intensity %
				7.52	7.4	19.4	22.8
8.38	41.7	20.6	100
				10.64	2.7	22.1	13.5
10.96	3.9	23.2	25.8
				13.64	3.9	24.3	52.1
15.16	22.2	27.2	3.8
				16.78	16.8	28.1	5.9
17.34	11.4	28.9	4.2
				17.74	3.3	29.7	4.4
18.84	14.9	31.8	3

Tenth aspect present invention provides a kind of AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation (being appointed as crystal formation II), its X-ray powder diffraction figure includes being located at 23.28 ± 0.2 °, the peak at 18.9 ± 0.2 °, 13.7 ± 0.2 ° and 14.72 ± 0.2 ° angle of diffraction (2 θ) place.

As further preferred scheme, preferably its X-ray powder diffraction figure also includes the peak positioned at 3.13 ± 0.2 °, 15.98 ± 0.2 °, 18.36 ± 0.2 °, 9.12 ± 0.2 ° and 32.38 ± 0.2 ° angle of diffraction (2 θ) place.

It is used as scheme still more preferably, it is preferred that its X-ray powder diffraction figure also includes being located at 10.88 ± 0.2 °, 22.18 ± 0.2 °, 21.92 ± 0.2 °, the peak at 27.66 ± 0.2 °, 20.34 ± 0.2 ° and 27.24 ± 0.2 ° angle of diffraction (2 θ) place.

As scheme still more preferably, the peak at the most preferably angle of diffraction (2 θ) place of its x-ray diffractogram of powder with being shown in Fig. 9 is substantially the same, and its X-ray powder diffraction data is as shown in table 5：

Table 5

2θ(°)	Intensity %	2θ(°)	Intensity %
				3.13	23.7	22.18	10.1
7.06	6.6	23.28	100
				9.12	16.3	24.06	5.5
10.88	10.2	27.24	6.7
				13.70	32.3	27.66	7
14.72	29.5	29.74	4.3
				15.98	20.7	31.14	3.8
18.36	17.7	32.38	11.6
				18.90	68.9	32.98	3.3
20.34	6.8	35.42	3.4
				21.92	9.5

Tenth one side of the invention provides a kind of preparation method of AHU-377 α-phenylethylamines salt crystal formation, including,

1) AHU-377 free acids are dissolved or are suspended in a suitable solvent, preparation generation AHU-377 α-phenylethylamine salt is mixed with equivalent or excessive α-phenylethylamine；

2) product obtains AHU-377 α-phenylethylamine salt crystal formations during collecting above-mentioned salt-forming reaction.

The α-phenylethylamine includes (R)-(+)-α-phenylethylamine, (S)-(-)-α-phenylethylamine.

It is used as further preferred scheme, step 1) the suitable solvent is recrystallisation solvent, including water, water-soluble solvent or water-insoluble solvent.

As scheme still more preferably, the water-soluble solvent is selected from alcohols, ketone, cyclic ethers class, amide-type, sulfoxide type organic solvent or its mixture, including but not limited to following solvent：Methanol, ethanol, normal propyl alcohol, isopropanol, n-butanol, acetonitrile, acetone, MEK, tetrahydrofuran, dioxane, DMF, dimethyl sulfoxide (DMSO) or its mixture；The water-insoluble solvent is selected from chloralkane, ethers, esters, alkanes, cycloalkane, benzene class organic solvent or its mixture, including but not limited to following solvent：Ethyl acetate, isopropyl acetate, dichloromethane, trichloroethanes, carbon tetrachloride, methyl tertiary butyl ether(MTBE), isopropyl ether, benzene,toluene,xylene or its composition.

It is used as further preferred scheme, the dissolving refers to the operation of one of ordinary skill in the art typically, generally can appropriate heating make AHU-377 free acids dissolution of raw material or dissolved clarification, or increase the consumption of solvent to make AHU-377 free acids dissolution of raw material or dissolved clarification.

As scheme still more preferably, recrystallisation solvent is preferably heated to 30~80 DEG C by the appropriate heating makes AHU-377 free acids dissolution of raw material or dissolved clarification.

As scheme still more preferably, the addition of the preferred recrystallisation solvent of consumption of the increasing solvent is 1-200 times of volume mass ratio of AHU-377 free acids, makes AHU-377 free acids dissolution of raw material or dissolved clarification.

It is used as further preferred scheme, step 2) described in the above-mentioned salt-forming reaction of collection during product the step of can be specifically directly to collect the solid product that is separated out during the above-mentioned reactant salt into α-phenylethylamine or by solvent flashing or add anti-solvent or directly cooling crystallization or add crystal seed, obtain AHU-377 α-phenylethylamine salt crystal formations；It is preferred that directly collecting the solid product separated out during the above-mentioned reactant salt into α-phenylethylamine obtains AHU-377 α-phenylethylamine salt crystal formations.

As scheme still more preferably, the anti-solvent refers to the less organic solvent of polarity, including but not limited to following solvent：Normal heptane, n-hexane, isooctane, pentane, hexamethylene, pentamethylene, ether or its composition.

It is used as further preferred scheme, step 2 of the present invention) obtained by its X-ray powder diffraction figure of AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation include being located at 20.58 ± 0.2 °, 24.28 ± 0.2 °, the peak at 8.38 ± 0.2 ° and 23.20 ± 0.2 ° angle of diffraction (2 θ) place, or resulting AHU-377 (S)-(-)-its X-ray powder diffraction figure of α-phenylethylamine salt is including being located at 23.28 ± 0.2 °, the peak at 18.9 ± 0.2 °, 13.7 ± 0.2 ° and 14.72 ± 0.2 ° angle of diffraction (2 θ) place.

The twelfth aspect of the present invention provides a kind of pharmaceutical composition, foregoing AHU-377 crystal type free acid of the described pharmaceutical composition comprising treatment effective dose, the calcium salt crystal formations of AHU-377 half, the calcium salt trihydrate crystal formations of AHU-377 half, the calcium salt anhydrous crystal forms of AHU-377 half, AHU-377 α-phenylethylamine salt crystal formation, AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation, AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation and pharmaceutically acceptable carrier or excipient.

As further preferred scheme, described pharmaceutical acceptable carrier or excipient are selected from diluent or filler, disintegrant, adhesive, glidant, lubricant, colouring agent or its combination.

It is used as scheme still more preferably, the diluent or filler are selected from Icing Sugar, sompressible sugar, glucose, sucrose, lactose, dextrin, mannitol, microcrystalline cellulose, sorbierite, starch or its combination, the consumption of diluent or filler is the 4.0%~60.0% of composition weight, preferably 10.0%~40.0%；The disintegrant is selected from starch, clay, cellulose, alginates, natural gum, cross-linked polymer, soybean polyoses, guar gum or its combination, the cross-linked polymer preferably is selected from PVPP, Ac-Di-Sol, cross-linked carboxymethyl cellulose calcium or its combination, the consumption of disintegrant is the 0%~65.0% of composition weight, preferably 1.0%~40.0%；Described adhesive is selected from starch, cellulose and its derivates, sucrose, glucose, corn syrup, gelatin, PVP or its combination, and the cellulose and its derivates preferably are selected from microcrystalline cellulose, hydroxypropyl cellulose, hydroxyethyl cellulose, hydroxypropyl methyl cellulose or its combination；More preferably hydroxypropyl cellulose；Most preferably low-substituted hydroxypropyl cellulose, the consumption of adhesive is the 1.0%~60.0% of composition weight, preferably 5.0%~40.0%, more preferably 10.0%~30.0%；The glidant or lubricant are selected from colloidal silica, magnesium trisilicate, starch, talcum powder, tricalcium orthophosphate, magnesium stearate, aluminum stearate, calcium stearate, calcium carbonate, magnesia, polyethylene glycol, powdered cellulose, Compritol 888 ATO, stearic acid, rilanit special, glycerin monostearate, sodium stearyl fumarate or its combination, the consumption of glidant or lubricant is the 0%~10.0% of composition weight, preferably 0.5%~5.0%.

As scheme still more preferably, described pharmaceutical composition further comprises diuretics；The diuretics is selected from frusemide, ethacrynic acid, bumetanide, Torasemide, Hydrochioro, chlorthalidone, bendroflumethiazide, Cyclopenthiazide, polythiazide, metolazone, indapamide or its combination；The consumption of diuretics is the 0.1%~10.0% of composition weight, preferably 0.5%~5.0%.

As scheme still more preferably, described pharmaceutical composition can prepare piece agent, capsule, granule.

As most preferred scheme, the capsule component proportion is as follows：

As most preferred scheme, the capsule component proportion is as follows：

The aspect of the present invention the 13rd provides a kind of foregoing AHU-377 crystal types free acid, the calcium salt crystal formations of AHU-377 half, the calcium salt trihydrate crystal formations of AHU-377 half, the calcium salt anhydrous crystal forms of AHU-377 half, AHU-377 α-phenylethylamine salt crystal formations, AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation, AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation or foregoing pharmaceutical composition are preparing the treatment or prevention disease relevant with neutral endopeptidase, it is cardiovascular, purposes in antihypertensive medicine.

It is used as scheme still more preferably, the antihypertensive medicine refers to by improving effect and the medicine of more effective antihypertensive therapy, including but not limited to anti-accelerated hypertension, essential hypertension, renovascular hypertension, diabetic hypertension, simple systolic hypertension or other secondary hypertension medicines being generated with higher corresponding rate.

Fourteenth aspect of the present invention provides a kind of foregoing AHU-377 crystal types free acid, the calcium salt crystal formations of AHU-377 half, the calcium salt trihydrate crystal formations of AHU-377 half, the calcium salt anhydrous crystal forms of AHU-377 half, AHU-377 α-phenylethylamine salt crystal formations, AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation, AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation or foregoing pharmaceutical composition are preparing treatment or prevention acute and chronic heart failure such as, congestive heart failure, left ventricle dysfunction, hypertrophic cardiomyopathy, diabetes cardiomyopathy, supraventricular and heart ventricle arrhythmia, atrial fibrillation, purposes in auricular flutter or harmful vascular remodeling medicine.

The fifteenth aspect of the present invention provides a kind of pharmaceutical composition, described pharmaceutical composition includes foregoing AHU-377 crystal types free acid, the calcium salt crystal formations of AHU-377 half, the calcium salt trihydrate crystal formations of AHU-377 half, the calcium salt anhydrous crystal forms of AHU-377 half, AHU-377 α-phenylethylamine salt crystal formation, AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation, AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation and angiotensinⅡ (AT1) receptor antagonist for the treatment of effective dose, and pharmaceutically acceptable carrier or excipient.

As further preferred scheme, the angiotensinⅡantagonist is selected from Losartan, Irbesartan, Olmesartan, Telmisartan, Valsartan, Azilsartan, Candesartan, Eprosartan, Losartan, smooth husky Puli's sand, Elisartan, Tasosartan, Ai Lishatan or its officinal salt.

As scheme still more preferably, the angiotensinⅡantagonist is selected from Valsartan, Azilsartan or its officinal salt.

The aspect of the present invention the 16th provides a kind of foregoing pharmaceutical composition and is preparing treatment or prevention miocardial infarction and its sequelae, atherosclerosis, angina pectoris, diabetic keratopathy or non-diabetic renal insufficiency, secondary aldosteronism, primary or secondary pulmonary hypertension, nephrosis, glomerulonephritis, chorionitis, glomerulosclerosis, the albuminuria of Primary Nephrosis, renovascular hypertension, diabetic retinopathy, antimigraine, peripheral vascular disease, Raynaud's disease, the hyperplasia of chamber, cognitive dysfunction, purposes in glaucoma or apoplexy medicine.

" pharmaceutical composition " is represented containing one or more compounds described herein or its physiologically/pharmaceutically acceptable salt or pro-drug and the mixture of other chemical constituents, with or other components such as physiology/pharmaceutically acceptable carrier and excipient.The purpose of pharmaceutical composition is to promote the administration to organism, the absorption beneficial to active component and then performance bioactivity.

Pharmaceutical composition containing active component can apply to oral form, such as tablet, dragee, lozenge, water or oil suspension, dispersible powder or particle, emulsion, hard or soft capsule, or syrup or elixir.Orally administered composition can be prepared according to any known method for preparing Pharmaceutical composition in this area, such composition can be selected from following composition containing one or more：Sweetener, flavouring, colouring agent and preservative, to provide pleasing and tasty pharmaceutical formulation.Tablet contains active component and the suitable nontoxic pharmaceutically acceptable excipient for preparing tablet for mixing.These excipient can be inert excipient, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate；Granulating agent and disintegrant, such as microcrystalline cellulose, Ac-Di-Sol, cornstarch or alginic acid；Adhesive, such as starch, gelatin, polyvinylpyrrolidone or Arabic gum and lubricant, such as magnesium stearate, stearic acid or talcum powder.These tablets can not be coated or can be by covering the taste of medicine or delay disintegration and absorption in the gastrointestinal tract, thus provide the known of slow releasing function in a long time Technology is coated.For example, water soluble taste can be used to shelter material, such as hydroxypropyl methyl cellulose or hydroxypropyl cellulose, or extension time material such as ethyl cellulose, acetylbutyrylcellulose.

Also wherein active component and the inert solid diluent hard gelatin capsule that for example calcium carbonate, calcium phosphate or kaolin are mixed are can use, or wherein active component provides oral formulations with water-solubility carrier such as polyethylene glycol or the oily solvent Perle that for example peanut oil, atoleine or olive oil are mixed.

Water slurry contains active material and the suitable excipient for preparing water slurry for mixing.Such excipient is suspending agent, for example sodium carboxy methyl cellulose, methylcellulose, hydroxypropyl methyl cellulose, mosanom, polyvinylpyrrolidone and Arabic gum；Dispersant or wetting agent can be naturally-produced phosphatide such as lecithin, or the condensation product of alkylene oxide and aliphatic acid such as Myrj 45, or the condensation product of oxirane and long-chain fatty alcohol, such as 17 carbon ethyleneoxy group cetanols (heptadecaethyleneoxy cetanol), or the condensation product of oxirane and the part ester as derived from aliphatic acid and hexitol, such as polyoxyethylene sorbitol monoleate, or the condensation product of oxirane and the partial ester as derived from aliphatic acid and hexitan, such as PEO Arlacel-80.Aqueous suspension can also contain one or more preservatives such as ethylparaben or nipalgin n-propyl, one or more colouring agents, one or more flavourings and one or more sweeteners, such as sucrose, saccharin or aspartame.

Oil suspension can be formulated by making active component be suspended in vegetable oil such as peanut oil, olive oil, sesame oil or coconut oil, or in mineral oil such as atoleine.Oil suspension can contain thickener, such as beeswax, hard paraffin or cetanol.Above-mentioned sweetener and flavouring can be added, to provide tasty preparation.These compositions can be preserved by adding antioxidant such as Butylated Hydroxyanisole or alpha-tocopherol.

It can make to be applied to prepare dispersible powder and particle offer active component and dispersant or wetting agent for mixing, suspending agent or one or more preservatives that water is suspended also by adding water.Above-mentioned example can be explained in suitable dispersant or wetting agent and suspending agent.Also other excipient such as sweetener, flavouring and colouring agent can be added.These compositions are preserved by adding antioxidant such as ascorbic acid.

The pharmaceutical composition of the present invention can also be the form of oil in water emulsion.Oil phase can be vegetable oil such as olive oil or peanut oil, or mineral oil such as atoleine or its mixture.Suitable emulsifying agent can be naturally-produced phosphatide, such as soybean lecithin and ester or partial ester such as sorbitan monooleate as derived from aliphatic acid and hexitan, and the partial ester and oxirane condensation product, such as polyoxyethylene sorbitol monoleate.Emulsion can also contain sweetener, flavouring, preservative and antioxidant.Syrup and elixir can be prepared with Sweetening agents such as glycerine, propane diols, sorbierite or sucrose.Such preparation can also contain moderator, preservative, colouring agent and antioxidant.

Pharmaceutical composition can be sterile injectable aqueous form.Can there are water, ringer's solution and isotonic sodium chlorrde solution in the acceptable solvent and solvent used.Aseptic injection preparation can be the aseptic injection oil-in-water microemulsion that wherein active component is dissolved in oil phase.For example active component is dissolved in the mixture of soybean oil and lecithin.Then oil solution is added to processing in the mixture of water and glycerine and forms micro emulsion.Parenteral solution or micro emulsion can be injected in the blood flow of patient by local a large amount of injections.Or, preferably give solution and micro emulsion in the way of it can keep the compounds of this invention constant circulating concentration.To keep this constant density, continuous intravenous delivery device can be used.

Pharmaceutical composition can be aseptic injection water or the form of oil suspension for intramuscular and subcutaneous administration.By known technology the suspension can be prepared with the suitable dispersant of those described above or wetting agent and suspending agent.Aseptic injection preparation can also be the solution prepared in the aseptic injectable solution or suspension prepared in the acceptable diluent of nontoxic parenteral or solvent, such as 1,3-BDO.In addition, it is convenient to be used as solvent or suspension media with sterile fixed oil.For this purpose, any mediation fixing oil including synthetic glycerine list or diester can be used.In addition, aliphatic acid such as oleic acid can also prepare injection.

The compounds of this invention can be given by the suppository form for rectally.Can be by the way that by medicine, with being at normal temperatures solid but being liquid in the rectum, thus the suitable nonirritant excipient that can be dissolved in the rectum and discharge medicine mixes to prepare these pharmaceutical compositions.Such material includes the mixture of the fatty acid ester of cocoa butter, glycerin gelatine, hydrogenated vegetable oil, the polyethylene glycol of various molecular weight and polyethylene glycol.

Well-known to those skilled in the art, the dosage of medicine depends on many factors, including but and non-limiting following factor：The activity of specific compound used, the age of patient, the body weight of patient, the health status of patient, the row quilt of patient, the diet of patient, administration time, administering mode, speed, the combination of medicine of excretion etc.；In addition, optimal therapeutic modality can be verified such as the species of the pattern treated, AHU-377 or its salt consumption per day or pharmaceutically acceptable salt according to traditional therapeutic scheme.

Compared with prior art, the present invention has the advantage that：

1) present invention provides a kind of new AHU-377 free acid crystal formations.AHU-377 free acids are oily compound, and pure compound is hardly resulted in technique, because dissolvent residual is difficult to remove, and AHU-377 crystallizations can then be obtained into the AHU-377 of high-purity, preparation technology is simple.Oily compound is extremely unfavorable for using and producing for oral solid formulation, and solid, which is made, then very can easily realize the use of oral solid formulation.Crystal is better than grease in stability.

2) present invention changes the physicochemical property of AHU-377 free acids by the way that AHU-377 is prepared into α-phenylethylamine salt, such as crystallinity, dissolubility, hygroscopicity, present invention process is ripe, it is workable, products obtained therefrom quality height and stable homogeneous, chemical stability, beneficial to storage and the AHU-377 raising isolated and purified with chiral purity.

3) present invention provides a kind of new AHU-377 calcium salt trihydrates, and purity is high, and steady quality is reliable, improves AHU-377 or the hygroscopicity of its sodium salt, improves chemical stability etc., beneficial to storage, meets industrialized production needs.

4) the present invention gained suitable clinical medicine applications of AHU-377 calcium salt crystal formations I, prepared capsule can meet medical science or pharmacy needs, suitable industrialization.

5) method for crystallising of the present invention is workable, technical maturity, it is easy to control.

6) present invention gained AHU-377 calcium salts are compared with sodium salt, in hypertension and heart failure therapeutic process, it is possible to reduce the intake of sodium, alleviate patient's angiocarpy burden.And calcium effect of releiving to blood vessel, is conducive to the treatment of angiocardiopathy.

Therefore, AHU-377 is made into calcium salt on the drug safety and validity in hypertension and heart failure disease treatment has obvious advantage compared with sodium salt.

Brief description of the drawings

1st, Fig. 1 is AHU-377 free acid crystal formations I X-ray powder diffraction figure, and abscissa is the θ (°) of angle 2, and ordinate is intensity.

2nd, Fig. 2 is AHU-377 free acid crystal formations I differential calorimetric scan figure (DSC) figure, and abscissa is temperature (DEG C), ordinate position hot-fluid (W/g).

3rd, Fig. 3 is AHU-377 free acid crystal formations I thermogravimetric analysis (TGA) figure, and abscissa is temperature (DEG C), and ordinate position is weightless ratio (%).

4th, Fig. 4 is the X-ray powder diffraction figure (XRPD figures) of AHU-377 calcium salt trihydrate crystal formations.

5th, Fig. 5 is the thermogravimetric analysis figure (TGA figures) of AHU-377 calcium salt trihydrate crystal formations.

6th, Fig. 6 is the X-ray powder diffraction figure (XRPD figures) of AHU-377 calcium salt anhydrous crystal forms.

7th, Fig. 7 schemes for the XRPD of AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation, and abscissa is the θ (°) of angle 2, and ordinate is intensity.

8th, Fig. 8 schemes for the DSC of AHU-377 (R)-(+)-α-benzene second salt crystal formation, and abscissa is temperature (DEG C), and ordinate is hot-fluid (w/g).

9th, Fig. 9 schemes for the XRPD of AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation, and abscissa is the θ (°) of angle 2, and ordinate is intensity.

10th, Figure 10 schemes for the DSC of AHU-377 (S)-(-)-α-phenylethylamine salt form, and abscissa is temperature (DEG C), and ordinate is hot-fluid (w/g).

11st, Figure 11 is the chiral chromatogram figure of AHU-377 and 2- methyl S enantiomeric mixtures, and abscissa is the time (minute), and ordinate is absorbance (mAU).

12nd, Figure 12 is using AHU-377 and 2- methyl S enantiomeric mixtures as raw material, prepare the chiral chromatogram figure of AHU-377 (S)-(-)-α-phenylethylamine salt, abscissa is the time (minute), and ordinate is absorbance (mAU).

Embodiment

The particular aspects of embodiment of the present invention will be further illustrated in specific embodiment presented below and preparation method example.The scope of the following example will not limit the scope of the present invention in any way.

Method and material

AHU-377 crystal types free acid, AHU-377 calcium salts polymorphic, AHU-377 α-phenylethylamines crystal formation are characterized by their X-ray powder diffraction figure.Therefore, with use Cu K α radiationsThe X-ray powder diffraction figure of the compound is gathered on GADDS (the general area diffraction detector system) CS operated in reflection Bruker D8Discover x-ray powder diffraction instruments.Tube voltage and the magnitude of current are respectively set to 40kV and 40mA acquisition scans.In the period of sample 60 seconds being scanned in the range of 3.0 ° to 40 ° of 2 θ.The peak position represented for 2 θ, diffractometer is calibrated using corundum standard product.In all analyses of typically 20 DEG C -30 DEG C of implementation at room temperature.Using the GADDS for 4.1.14T editions WNT softwares, collection and integration data.Using the DiffracPlus softwares with 9.0.0.2 editions Eva of distribution in 2003, diffraction pattern is analyzed.The preparation of XRPD samples, by being, as on monocrystalline silicon piece, to press sample powder flat with the surface for ensuring sample with sheet glass or equivalent by sample And have appropriate height.Then sample holder is put into Bruker XRPD instruments, and X-ray powder diffraction figure is gathered using above-described instrument parameter.By producing the measurement difference related to this kind of X-ray powder diffraction analysis result including following many factors：(a) error in sample preparation thing (such as height of specimen), (b) instrument error, (c) difference is calibrated, (d) personal error (being included in those errors for occurring when determining peak position), and (e) material property (such as preferred orientation error).Calibration error and sample height errors frequently result in displacement of all peaks in equidirectional.In general, this calibration factor is consistent with expected peak position and can be in the scope of expected 2 θ value ± 0.2 ° by the peak position for making measurement.

Raw material A HU-377 free acids of the present invention are to report preparation method according to patent US5217996A and obtain, and product is grease.θ (°) value of angle 2 and intensity level (being used as the % of peak-peak) of crystal formation obtained by the embodiment of the present invention have been included in table 1-5.

Differential scanning calorimetry (DSC) is carried out on TA Instruments DSC Q2000.Respectively cell constant and thermal capacity calibration instrument are directed to using indium and sapphire.By the way that 1-3mg sample weighing is entered into aluminium dish, then cover the aluminium dish using aluminium lid and prepare sample.Use the analyze datas of Universal Analysis 2000.Start experiment in room temperature, and under nitrogen purging (flow velocity is 50ml/min), 300 DEG C are heated the sample to 10 DEG C/min.

The analysis of diastereoisomer detects that chiral chromatographic analysis condition is using HPLC：Chromatographic column：Daicel Chiralpak AD-H (4.6*250mm, 5um)；Mobile phase A:Hexamethylene：Isopropanol：Glacial acetic acid=1000:10:1；Mobile phase B：Hexamethylene：Isopropanol：Glacial acetic acid=800:200:1；Mobile phase A：Mobile phase B=50:50；Per pin run time：22min；Column temperature：30℃；Wavelength：254nm；Flow velocity：1.0ml/min.

Embodiment 1

200mg AHU-377 free acids (oily) are weighed to be placed in 20.0mL vials, then 4mL methyl tertiary butyl ether(MTBE)s are added, stir dissolved clarification, normal heptane is slowly added to occurring muddy (about 5mL), (20-25 DEG C) is continued to stir 48 hours at room temperature, separation of solid and liquid obtains AHU-377 crystal type free acids, and its X-ray powder diffraction figure is as shown in Figure 1.

There are AHU-377 crystal type free acid fusing points with differential calorimetric scan instrument (DSC, model TA Q2000) measurement.Measuring condition is to be heated to 250 DEG C from room temperature, and heating rate is 10 DEG C per minute, and heating is carried out in a nitrogen atmosphere, and nitrogen flow is that 20mL is per minute.The DSC figures of AHU-377 crystal type free acids are as shown in Figure 2.The fusing point (onset points) of AHU-377 crystal type free acids is：94.3 DEG C or so.

The thermal weight loss situation of AHU-377 crystal type free acids is measured with thermogravimetric analyzer (TGA, model TA Q500).Measuring condition is to be heated to 300 DEG C from room temperature, and heating rate is 10 DEG C per minute, and heating is carried out in a nitrogen atmosphere, and nitrogen flow is that 50mL is per minute.The TGA figures of AHU-377 crystal type free acids are as shown in Figure 3.It is hardly weightless within 100 DEG C, therefore can be determined that AHU-377 crystal types free acid is anhydrous crystal forms.

Embodiment 2

50mg AHU-377 free acids (oily) are weighed to be placed in 2.0mL vials, then 0.5mL isopropyl acetates are added, it is heated to 50 DEG C, stir dissolved clarification, 5.0mL n-hexanes are slowly added to, room temperature (20-25 DEG C) is then cooled to, continue insulated and stirred 48 hours, separation of solid and liquid obtains AHU-377 crystal type free acids, and its X-ray powder diffraction figure is consistent with Fig. 1.

Embodiment 3

50mg AHU-377 free acids (oily) are weighed to be placed in 2.0mL vials, then 0.25mL toluene is added, it is heated to 80 DEG C, stir dissolved clarification, then room temperature (20-25 DEG C) is cooled to, 2.0mL absolute ethers, insulated and stirred 48 hours is slowly added to, separation of solid and liquid obtains AHU-377 crystal type free acids, and its X-ray powder diffraction figure is consistent with Fig. 1.

Embodiment 4

50mg AHU-377 free acids (oily) are weighed to be placed in 2.0mL vials, then 0.5mL methyl tertiary butyl ether(MTBE)s are added, it is heated to 55-60 DEG C, stir dissolved clarification, then it is cooled to room temperature (20-25 DEG C), continue insulated and stirred 48 hours, separation of solid and liquid obtains AHU-377 crystal type free acids, and its X-ray powder diffraction figure is consistent with Fig. 1.

Embodiment 5

20mg AHU-377 free acids (oily) are weighed to be placed in 2.0mL vials, then 0.5mL ethyl acetate is added, stir dissolved clarification, it is slowly added to the crystal of 0.2mg embodiments 1, (20-25 DEG C) is continued to stir 24 hours at room temperature, separation of solid and liquid obtains AHU-377 crystal type free acids, and its X-ray powder diffraction figure is consistent with Fig. 1.

Embodiment 6

50mg AHU-377 free acids (oily) are weighed to be placed in 2.0mL vials, then 0.5mL isopropyl acetates are added, it is heated to 55-60 DEG C, stir dissolved clarification, then room temperature (20-25 DEG C) is cooled to, the crystal of 0.5mg embodiments 1 is slowly added to, (20-25 DEG C) is continued to stir 24 hours at room temperature, separation of solid and liquid obtains AHU-377 crystal type free acids, and its X-ray powder diffraction figure is consistent with Fig. 1.

Embodiment 7

Weigh 20mg AHU-377 free acids (oily) to be placed in 20.0mL vials, then add 2mL methyl tertiary butyl ether(MTBE)s, dissolving obtains settled solution, standby.The crystal of 2mg embodiments 1 is taken, is suspended in 10mL normal heptanes, AHU-377 t-butyl methyl ether solutions are slowly added dropwise into n-heptane solution, it is added dropwise about 1 hour, stirring while adding, separation of solid and liquid obtains AHU-377 crystal type free acids, and its X-ray powder diffraction figure is consistent with Fig. 1.

Embodiment 8

Weigh 500mg (1.215mmol) AHU-377 free acids (oily) to be placed in 50.0mL round-bottomed flasks, add 5.0mL acetone, stir dissolved clarification, add the sodium acid carbonate that 12.2mL concentration is 0.10mmol/mL The aqueous solution, stirring reaction 12 hours, the calcium chloride water that 0.65mL concentration is 1.0mmol/mL is added, 15mL water is then added again, stirring makes it fully react in 24 hours.Separation of solid and liquid obtains AHU-377 calcium salt crystal formation I, and its X-ray powder diffraction figure (XRPD figures) is as shown in Figure 4.

AHU-377 calcium salt crystal formation I thermal weight loss situations are measured with thermogravimetric analyzer (TGA, model TA Q500).Measuring condition is to be heated to 400 DEG C from room temperature, and heating rate is 10 DEG C per minute, and heating is carried out in a nitrogen atmosphere, and nitrogen flow is that 50mL is per minute.AHU-377 calcium salt crystal formations I TGA figures are as shown in Figure 5.Weightless within 100 DEG C is 10%, (theoretical weightlessness is 11%) approximate with the theoretical weightlessness of the calcium salt trihydrates of AHU377 half therefore may determine that AHU-377 calcium salt crystal formations I is the crystal compound of half calcium of AHU377 three.

Embodiment 9

50mg (0.12mmol) AHU-377 free acids (oily) are weighed to be placed in 10.0mL round-bottomed flasks, add 1.0mL isopropanols, stir dissolved clarification, add the sodium acetate aqueous solution that 0.25mL concentration is 0.50mmol/mL, stirring reaction 12 hours, the solution of calcium bromide in water that 0.15mL concentration is 0.5mmol/mL is added, 5mL water is then added again, stirring makes it fully react in 24 hours.Separation of solid and liquid obtains AHU-377 calcium salt crystal formation I, and its X-ray powder diffraction figure (XRPD figures) is substantially consistent with such as Fig. 4.

Embodiment 10

50mg (0.12mmol) AHU-377 free acids (oily) are weighed to be placed in 10.0mL round-bottomed flasks, add 0.5mL tetrahydrofurans, stir dissolved clarification, add the potassium bicarbonate aqueous solution that 1.0mL concentration is 0.10mmol/mL, stirring reaction 12 hours, the calcium chloride water solution that 0.25mL concentration is 0.5mmol/mL is added, 5mL water is then added again, stirring makes it fully react in 24 hours.Separation of solid and liquid obtains AHU-377 calcium salt crystal formation I, and its X-ray powder diffraction figure (XRPD figures) is substantially consistent with such as Fig. 4.

Embodiment 11

50mg (0.12mmol) AHU-377 free acids (oily) are weighed to be placed in 10.0mL round-bottomed flasks, add 0.3mL N, dinethylformamide, stir dissolved clarification, the propionic acid aqueous solutions of potassium that 0.65mL concentration is 0.20mmol/mL is added, stirring reaction 12 hours adds the calcium chloride water that 0.30mL concentration is 0.5mmol/mL, then 5mL water is added again, stirring makes it fully react in 24 hours.Separation of solid and liquid obtains AHU-377 calcium salt crystal formation I, and its X-ray powder diffraction figure (XRPD figures) is substantially consistent with such as Fig. 4.

Embodiment 12

50mg (0.12mmol) AHU-377 free acids (oily) are weighed to be placed in 10.0mL round-bottomed flasks, add 1.0mL ethanol, it is heated to 60 DEG C of stirring dissolved clarifications, add the potassium bicarbonate aqueous solution that 1.4mL concentration is 0.10mmol/mL, stirring reaction 12 hours, the calcium lactate aqueous solution that 0.15mL concentration is 0.5mmol/mL is added, 5mL water is then added again, stirring makes it fully react in 24 hours.Separation of solid and liquid obtains AHU-377 calcium salt crystal formation I, and its X-ray powder diffraction figure (XRPD figures) is substantially consistent with such as Fig. 4.

Embodiment 13

50mg (0.12mmol) AHU-377 free acids (oily) are weighed to be placed in 10.0mL round-bottomed flasks, add 1.0mL acetonitriles, it is heated to 40 DEG C of stirring dissolved clarifications, add the sodium benzoate aqueous solution that 1.3mL concentration is 0.10mmol/mL, stirring reaction 12 hours, the calcium lactate aqueous solution that 0.2mL concentration is 0.5mmol/mL is added, 5mL water is then added again, stirring makes it fully react in 24 hours.Separation of solid and liquid obtains AHU-377 calcium salt crystal formation I, and its X-ray powder diffraction figure (XRPD figures) is substantially consistent with such as Fig. 4.

Embodiment 14

20mg AHU-377 calcium salt crystal formation I are weighed, in thermogravimetric analyzer, 90 DEG C, after Temperature fall are heated to, it is rapid to take out, crystal formation is detected with XRPD, its X-ray powder diffraction figure (XRPD figures) is substantially consistent with Fig. 6.

Embodiment 15

20mg (0.049mmol) AHU-377 free acids (oily) are weighed to be placed in 5.0mL vials, add 1.0mL isopropyl acetates, stir dissolved clarification, add 5.9mg (0.049mmol) (R)-(+)-α-phenylethylamine, continue stirring reaction 48 hours, its x-ray diffractogram of powder of AHU-377 (R)-(+) that collecting reaction product is obtained-α-phenylethylamine salt crystal formation is as shown in fig. 7, fusing point：126.6 DEG C (onset points) is as shown in Figure 8.

Embodiment 16

20mg (0.049mmol) AHU-377 free acids (oily) are weighed to be placed in 5.0mL vials, add 0.4mL isopropanols, stir dissolved clarification, add 6.5mg (0.053mmol) (R)-(+)-α-phenylethylamine, continue stirring reaction 48 hours, AHU-377 (R)-(+)-α-phenylethylamine crystal formation that collecting reaction product is obtained, its x-ray diffractogram of powder is substantially consistent with such as Fig. 7.

Embodiment 17

20mg (0.049mmol) AHU-377 free acids (oily) are weighed to be placed in 5.0mL vials, add 0.2mL methyl tertiary butyl ether(MTBE)s, it is heated to 50 DEG C of stirring dissolved clarifications, add 5.9mg (0.049mmol) (R)-(+)-α-phenylethylamine, it is cooled to (20~25 DEG C) of room temperature and continues stirring reaction 48 hours, AHU-377 (R)-(+) that collecting reaction product is obtained-α-phenylethylamine salt crystal formation, its x-ray diffractogram of powder is substantially consistent with such as Fig. 7.

Embodiment 18

20mg (0.049mmol) AHU-377 free acids (oily) are weighed to be placed in 5.0mL vials, add 0.2mL dichloromethane, it is heated to 40 DEG C of stirring dissolved clarifications, add 6.2mg (0.051mmol) (R)-(+)-α-phenylethylamine, it is cooled to (20~25 DEG C) of room temperature and continues stirring reaction 48 hours, AHU-377 (R)-(+) that collecting reaction product is obtained-α-phenylethylamine salt crystal formation, its x-ray diffractogram of powder is substantially consistent with such as Fig. 7.

Embodiment 19

20mg (0.049mmol) AHU-377 free acids (oily) are weighed to be placed in 5.0mL vials, add 0.2mL95% ethanol, it is heated to 60 DEG C of stirring dissolved clarifications, add 6.0mg (0.049mmol) (R)-(+)-α-phenylethylamine, it is cooled to (20~25 DEG C) of room temperature and continues stirring reaction 48 hours, AHU-377 (R)-(+) that collecting reaction product is obtained-α-phenylethylamine salt crystal formation, its x-ray diffractogram of powder is substantially consistent with such as Fig. 7.

Embodiment 20

20mg (0.049mmol) AHU-377 free acids (oily) are weighed to be placed in 5.0mL vials, add 1.0mL isopropyl acetates, stir dissolved clarification, add 5.9mg (0.049mmol) (S)-(-)-α-phenylethylamine, continue stirring reaction 48 hours, its x-ray diffractogram of powder of AHU-377 (S)-(-) that collecting reaction product is obtained-α-phenylethylamine salt crystal formation is as shown in figure 9, fusing point：140.16 DEG C (onset points) is as shown in Figure 10.

Embodiment 21

Weighing 40mg (0.096mmol) AHU-377 free acids, (ratio of AHU-377 and 2S- methyl diastereoisomers is about 50:50, as shown in figure 11, AHU-377 retention times are 10 minutes or so, 2S- methyl diastereoisomers retention time is 9 minutes or so) it is placed in 5.0mL vials, add 2.0mL isopropyl acetates, stir dissolved clarification, add 1mL 5.9mg/mL (0.049mmol) (S)-(-)-α-phenylethylamine isopropyl acetate solution, continue stirring reaction 24 hours, AHU-377 (S)-(-) that collecting reaction product is obtained-α-phenylethylamine salt crystal formation.1mg AHU-377 (S)-(-)-α-phenylethylamine salt will be taken to add in the aqueous acetic acids of 1mL 1%, add 1mL ethyl acetate, after strength is shaked, stratification, take the purity of ethyl acetate layer chiral chromatographic analysis AHU-377 diastereoisomer, HPLC results understand that AHU-377 chiral purity is greatly improved, and the ratio of AHU-377 and 2S- methyl diastereoisomers is about 99.5:0.5 as shown in spectrogram 12.

Embodiment 22

Component	The composition (mg) of per unit	Constitute (%)
			AHU-377 calcium salt crystal formations I	150	46.87
Microcrystalline cellulose	32	10.0
			PVP	70	21.88
PVPP	21.6	6.75
			Colloidal silica	3.2	1.0
Magnesium stearate	3.2	1.0
			Gross weight	320	100.0

Magnesium stearate, colloidal silica and microcrystalline cellulose are sieved by 30 mesh sieves first.Then said mixture, active components A HU-377 calcium salt crystal formations I, PVPP and PVP are mixed about 120 turns in hopper mixer.Using roller press the mixture is suppressed with 30kN pressure.After compacting, grind the mixture using grinder and sieved through 18 mesh sieves, obtain final interior phase or particle.By granule filling in capsule, capsule is made.

Embodiment 23

Component	The composition (mg) of per unit	Constitute (%)
			AHU-377 calcium salt crystal formations I	96	30.0
Microcrystalline cellulose	96	30.0
			Low-substituted hydroxypropyl cellulose	32	10.0
PVPP	89.6	28.0
			Colloidal silica	3.2	1.0
Magnesium stearate	3.2	1.0
			Gross weight	320	100

First the active components A HU-377 calcium salt crystal formations I is sieved by 40 mesh sieves.Microcrystalline cellulose and PVPP are added into active component, the mixture is sieved by 20 mesh sieves.Then the mixture is mixed to about 100 turns of rotation in hopper mixer.Then low-substituted hydroxypropyl cellulose and colloidal silica are added in hopper mixer, then it is rotated 100 turns.It is eventually adding magnesium stearate.Then powder mixture is compressed in blocks.

The stability of preparation and dissolution detection

The capsule of embodiment 22 and the tablet of embodiment 23 are put under different condition respectively, the stability of preparation is investigated, as a result it is as shown in the table：

It is demonstrated experimentally that AHU-377 calcium salt crystal formations I (AHU-377 calcium salts trihydrate) purity that the present invention is provided is high, steady quality is reliable, substantially improve AHU-377 or the hygroscopicity of its sodium salt, chemical stability etc. is improved, beneficial to the storage of product, meets industrialized production needs.The application of the suitable clinical medicines of gained AHU-377 calcium salt crystal formation I, prepared capsule can meet medical science or pharmacy needs.In hypertension and heart failure therapeutic process, it is possible to reduce the intake of sodium, patient's angiocarpy burden is alleviated, is conducive to the treatment of angiocardiopathy.Therefore AHU-377 is made into calcium salt etc. in the security and validity in hypertension and heart failure disease treatment has obvious advantage compared with sodium salt.

Finally it should be noted that, the above embodiments are merely illustrative of the technical solutions of the present invention is not intended to limit the present invention, although the present invention is described in detail with reference to preferred embodiment, it will be understood by those within the art that, the technical scheme of invention can be modified or equivalent substitution, without departing from the spirit and scope of technical solution of the present invention, it all should cover in scope of the presently claimed invention.

Claims (35)

1. A kind of AHU-377 crystal types free acid, its X-ray powder diffraction figure include be located at 23.70 ± 0.2 °, 15.84 ± 0.2 °, 20.84 ± 0.2 ° and 8.56 ± 0.2 °, the angle of diffraction (2 θ) place peak；It is preferred that its X-ray powder diffraction figure also includes the peak positioned at 17.06 ± 0.2 °, 25.98 ± 0.2 °, 18.01 ± 0.2 °, 9.42 ± 0.2 ° and 26.91 ± 0.2 ° angle of diffraction (2 θ) place；More preferably its X-ray powder diffraction figure further comprises the peak positioned at 34.70 ± 0.2 °, 21.83 ± 0.2 °, 25.69 ± 0.2 °, 25.33 ± 0.2 °, 11.67 ± 0.2 ° and 26.24 ± 0.2 ° angle of diffraction (2 θ) place；The peak at the most preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 1 is substantially the same.
2. The method for preparing AHU-377 crystal type free acids described in claim 1, it is characterised in that comprise the following steps,

   1) AHU-377 free acids are dissolved in suitable organic solvent；

   2) solution system temperature is reduced, and/or adds crystal seed, and/or adds appropriate anti-solvent to solution and muddy, crystallization occurs；

   3) separation of solid and liquid obtains AHU-377 crystal type free acids.
3. Preparation method according to claim 2, it is characterised in that comprise the following steps,

   1) AHU-377 free acids are dissolved under normal temperature or heated condition in suitable organic solvent；

   2) reduce step 1) in solution system temperature, and/or add crystal seed, crystallization；

   3) separation of solid and liquid obtains AHU-377 crystal type free acids.
4. Preparation method according to claim 2, it is characterised in that comprise the following steps,

   1) AHU-377 free acids are dissolved in suitable organic solvent；

   2) by step 1) in solution drop mixed with anti-solvent；

   3) separation of solid and liquid obtains AHU-377 crystal type free acids；

   It is preferred that, step 2) in mixed with anti-solvent before AHU-377 crystal type free acid crystal seeds are previously added in anti-solvent.
5. Preparation method according to claim any one of 2-4, it is characterised in that step 1) the suitable organic solvent be selected from halogen ethers, cyclic ethers class, esters, halogenated alkane, benzene class organic solvent or its mixture；Ethyl acetate, isopropyl acetate, dichloromethane, trichloroethanes, carbon tetrachloride, methyl tertiary butyl ether(MTBE), isopropyl ether, benzene,toluene,xylene or its composition preferably are selected from, more preferably from isopropyl acetate, dichloromethane, methyl tertiary butyl ether(MTBE), isopropyl ether or its composition.
6. Preparation method according to claim 2 or claim 4, it is characterised in that step 2) anti-solvent be selected from normal heptane, n-hexane, isooctane, pentane, hexamethylene, pentamethylene, ether or its composition；It preferably is selected from normal heptane, n-hexane, hexamethylene, ether or its composition.
7. Preparation method according to claim any one of 2-4, it is characterized in that, step 2) described in add crystal seed and refer to add homology crystalline solid in AHU-377 free acid solutions, add the 0.1-20.0% that crystal seed amount is AHU-377 free acid mass ratioes, preferably be selected from the 0.5-5.0% of mass ratio.
8. Preparation method according to claim any one of 2-4, it is characterized in that, step 3) obtained by AHU-377 crystal types free acid its X-ray powder diffraction figure include being located at 23.70 ± 0.2 °, 15.84 ± 0.2 °, 20.84 ± 0.2 ° and 8.56 ± 0.2 °, the angle of diffraction (2 θ) place peak.
9. A kind of calcium salt crystal formations of AHU-377 half, its X-ray powder diffraction figure includes being located at 12.70 ± 0.2 °, 7.32 ± 0.2 °, the peak at 15.90 ± 0.2 ° and 18.56 ± 0.2 ° angle of diffraction (2 θ) place, or its X-ray powder diffraction figure includes the peak positioned at 4.02 ± 0.2 °, 3.62 ± 0.2 ° and 17.82 ± 0.2 ° angle of diffraction (2 θ) place.
10. A kind of calcium salt trihydrate crystal formations of AHU-377 half, its X-ray powder diffraction figure includes the peak positioned at 12.70 ± 0.2 °, 7.32 ± 0.2 °, 15.90 ± 0.2 ° and 18.56 ± 0.2 ° angle of diffraction (2 θ) place；It is preferred that its X-ray powder diffraction figure also includes the peak positioned at 14.74 ± 0.2 °, 15.42 ± 0.2 °, 7.82 ± 0.2 °, 19.42 ± 0.2 °, 16.64 ± 0.2 ° and 17.54 ± 0.2 ° angle of diffraction (2 θ) place；The peak at the more preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 4 is substantially the same.
11. A kind of calcium salt anhydrous crystal forms of AHU-377 half, its X-ray powder diffraction figure includes the peak positioned at 4.02 ± 0.2 °, 3.62 ± 0.2 ° and 17.82 ± 0.2 ° angle of diffraction (2 θ) place；It is preferred that its X-ray powder diffraction figure also includes the peak positioned at 20.48 ± 0.2 °, 14.50 ± 0.2 ° and 19.66 ± 0.2 ° angle of diffraction (2 θ) place；The peak at the more preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 6 is substantially the same.
12. A kind of preparation method of the calcium salt trihydrate crystal formations of AHU-377 half, it is characterised in that comprise the following steps,

    1) AHU-377 free acids are dissolved or are suspended in a suitable solvent, the corresponding salt of preparation generation AHU-377 is mixed with equivalent or the excessive caustic alkali or alkalescent sodium salt or sylvite for dissolving in the system, it is preferred that, mixed with the alkalescent sodium salt or sylvite of equivalent or excess；

    2) above-mentioned AHU-377 salt systems are mixed with water-soluble Ca salt in the aqueous systems of water or the solvent containing machine, produces AHU-377 calcium precipitations；

    3) product obtains the calcium salt trihydrate crystal formations of AHU-377 half during collecting above-mentioned salt-forming reaction；

    Step 1) the suitable solvent is recrystallisation solvent, including water, water-soluble solvent or its mixture.
13. Preparation method according to claim 12, it is characterized in that, the water-soluble solvent is selected from alcohols, ketone, cyclic ethers class, amide-type, sulfoxide type organic solvent or its mixture, it preferably is selected from methanol, ethanol, normal propyl alcohol, isopropanol, n-butanol, acetonitrile, acetone, MEK, tetrahydrofuran, dioxane, DMF, dimethyl sulfoxide (DMSO) or its mixture.
14. Preparation method according to claim 12, it is characterized in that, step 1) caustic alkali be sodium hydroxide or potassium hydroxide, the alkalescent sodium salt is selected from sodium carbonate, sodium acid carbonate, sodium acetate, sodium formate, sodium propionate, PAA, sodium benzoate or its mixture, it is preferred that sodium acid carbonate, sodium acetate or its mixture, the alkalescent sylvite is selected from potassium carbonate, saleratus, potassium acetate, potassium formate, potassium propionate, potassium acrylate, Potassium Benzoate or its mixture, potassium bicarbonate, potassium acetate or its mixture；Step 2) water-soluble Ca salt is selected from calcium chloride, calcium bromide, calcium iodide, calcium nitrate, calcium chlorate, calcium hypochlorite, and Calcium perchlorate, calcium lactate, calcium gluconae or its mixture preferably are selected from calcium chloride, calcium chlorate or its mixture.
15. Preparation method according to claim 12, it is characterized in that, step 3) obtained by AHU-377 calcium salt crystal formation I its x-ray diffractogram of powder include being located at 12.70 ± 0.2 °, 7.32 ± 0.2 °, the peak at 15.90 ± 0.2 ° and 18.56 ± 0.2 ° angle of diffraction (2 θ) place, it is preferred that its X-ray powder diffraction figure also includes being located at 14.74 ± 0.2 °, 15.42 ± 0.2 °, 7.82 ± 0.2 °, the peak at 19.42 ± 0.2 °, 16.64 ± 0.2 ° and 17.54 ± 0.2 ° angle of diffraction (2 θ) place.
16. A kind of preparation method of the calcium salt anhydrous crystal forms of AHU-377 half, it is characterised in that the calcium salt trihydrates of AHU-377 half are heated to more than 50 DEG C, make its dehydration be transformed into anhydrous crystal forms；It is preferred that heating temperature range is at 50-100 DEG C.
17. Preparation method according to claim 16, it is characterised in that resulting its X-ray powder diffraction figure of calcium salt anhydrous crystal forms of AHU-377 half includes the peak at the angle of diffraction (2 θ) place positioned at 4.02 ± 0.2 °, 3.62 ± 0.2 ° and 17.82 ± 0.2 °；It is preferred that its X-ray powder diffraction figure also includes the peak positioned at 20.48 ± 0.2 °, 14.50 ± 0.2 ° and 19.66 ± 0.2 ° angle of diffraction (2 θ) place.
18. A kind of AHU-377 α-phenylethylamines salt, its X-ray powder diffraction figure includes being located at 20.58 ± 0.2 °, 24.28 ± 0.2 °, the peak at 8.38 ± 0.2 ° and 23.20 ± 0.2 ° angle of diffraction (2 θ) place, or including positioned at 23.28 ± 0.2 °, the peak at 18.9 ± 0.2 °, 13.7 ± 0.2 ° and 14.72 ± 0.2 ° angle of diffraction (2 θ) place.
19. A kind of AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation, its X-ray powder diffraction figure includes the peak positioned at 20.58 ± 0.2 °, 24.28 ± 0.2 °, 8.38 ± 0.2 ° and 23.20 ± 0.2 ° angle of diffraction (2 θ) place；It is preferred that its X-ray powder diffraction figure also includes the peak positioned at 19.36 ± 0.2 °, 15.16 ± 0.2 °, 16.78 ± 0.2 °, 18.84 ± 0.2 ° and 22.06 ± 0.2 ° angle of diffraction (2 θ) place；More preferably its X-ray powder diffraction figure further comprises the peak positioned at 17.34 ± 0.2 °, 7.52 ± 0.2 °, 28.10 ± 0.2 °, 29.66 ± 0.2 °, 28.94 ± 0.2 ° and 10.96 ± 0.2 ° angle of diffraction (2 θ) place；The peak at the most preferably angle of diffraction (2 θ) place of its X-ray powder diffraction figure with being shown in Fig. 7 is substantially the same.
20. A kind of AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation, its X-ray powder diffraction figure includes the peak positioned at 23.28 ± 0.2 °, 18.9 ± 0.2 °, 13.7 ± 0.2 ° and 14.72 ± 0.2 ° angle of diffraction (2 θ) place；It is preferred that also wrapping Include the peak positioned at 3.13 ± 0.2 °, 15.98 ± 0.2 °, 18.36 ± 0.2 °, 9.12 ± 0.2 ° and 32.38 ± 0.2 ° angle of diffraction (2 θ) place；More preferably its X-ray powder diffraction figure further comprises the peak positioned at 10.88 ± 0.2 °, 22.18 ± 0.2 °, 21.92 ± 0.2 °, 27.66 ± 0.2 °, 20.34 ± 0.2 ° and 27.24 ± 0.2 ° angle of diffraction (2 θ) place；The peak at the most preferably angle of diffraction (2 θ) place of its x-ray diffractogram of powder with being shown in Fig. 9 is substantially the same.
21. A kind of preparation method of AHU-377 α-phenylethylamines salt crystal formation, including,

    1) AHU-377 free acids are dissolved or are suspended in a suitable solvent, preparation generation AHU-377 α-phenylethylamine salt is mixed with equivalent or excessive α-phenylethylamine；

    2) product obtains AHU-377 α-phenylethylamine salt crystal formations during collecting above-mentioned salt-forming reaction；

    The α-phenylethylamine includes (R)-(+)-α-phenylethylamine, (S)-(-)-α-phenylethylamine；Step 1) the suitable solvent is recrystallisation solvent, including water, water-soluble solvent or water-insoluble solvent.
22. Preparation method according to claim 21, it is characterised in that the water solubility is molten

    Agent is selected from alcohols, ketone, cyclic ethers class, amide-type, sulfoxide type organic solvent or its mixture, it preferably is selected from methanol, ethanol, normal propyl alcohol, isopropanol, n-butanol, acetonitrile, acetone, MEK, tetrahydrofuran, dioxane, DMF, dimethyl sulfoxide (DMSO) or its mixture；The water-insoluble solvent is selected from chloralkane, ethers, esters, alkanes, cycloalkane, benzene class organic solvent or its mixture, preferably is selected from ethyl acetate, isopropyl acetate, dichloromethane, trichloroethanes, carbon tetrachloride, methyl tertiary butyl ether(MTBE), isopropyl ether, benzene,toluene,xylene or its composition.
23. Preparation method according to claim 21, it is characterized in that, step 2) obtained by its X-ray powder diffraction figure of AHU-377 (R)-(+)-α-phenylethylamine salt include being located at 20.58 ± 0.2 °, 24.28 ± 0.2 °, the peak at 8.38 ± 0.2 ° and 23.20 ± 0.2 ° angle of diffraction (2 θ) place, or resulting AHU-377 (S)-(-)-its X-ray powder diffraction figure of α-phenylethylamine salt is including being located at 23.28 ± 0.2 °, the peak at 18.9 ± 0.2 °, 13.7 ± 0.2 ° and 14.72 ± 0.2 ° angle of diffraction (2 θ) place.
24. A kind of pharmaceutical composition, it is characterized in that the AHU-377 crystal type free acids described in claim 1 of the described pharmaceutical composition comprising treatment effective dose, the calcium salt crystal formations of AHU-377 half described in claim 9, the calcium salt trihydrate crystal formations of AHU-377 half described in claim 10, the calcium salt anhydrous crystal forms of AHU-377 half described in claim 11, AHU-377 α-phenylethylamine salt crystal formations described in claim 18, AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation and pharmaceutically acceptable carrier or excipient described in AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation or claim 20 described in claim 19.
25. Pharmaceutical composition according to claim 24, it is characterised in that described pharmaceutical acceptable carrier or excipient are selected from diluent or filler, disintegrant, adhesive, glidant, lubricant, colouring agent or its combination.
26. Pharmaceutical composition according to claim 24, it is characterised in that the diluent or filler Selected from Icing Sugar, sompressible sugar, glucose, sucrose, lactose, dextrin, mannitol, microcrystalline cellulose, sorbierite, starch or its combination, the consumption of diluent or filler is the 4.0%~60.0% of composition weight, preferably 10.0%~40.0%；The disintegrant is selected from starch, clay, cellulose, alginates, natural gum, cross-linked polymer, soybean polyoses, guar gum or its combination, the cross-linked polymer preferably is selected from PVPP, Ac-Di-Sol, cross-linked carboxymethyl cellulose calcium or its combination, the consumption of disintegrant is the 0%~65.0% of composition weight, preferably 1.0%~40.0%；Described adhesive is selected from starch, cellulose and its derivates, sucrose, glucose, corn syrup, gelatin, PVP or its combination, and the cellulose and its derivates preferably are selected from microcrystalline cellulose, hydroxypropyl cellulose, hydroxyethyl cellulose, hydroxypropyl methyl cellulose or its combination；More preferably hydroxypropyl cellulose；Most preferably low-substituted hydroxypropyl cellulose, the consumption of adhesive is the 1.0%~60.0% of composition weight, preferably 5.0%~40.0%, more preferably 10.0%~30.0%；The glidant or lubricant are selected from colloidal silica, magnesium trisilicate, starch, talcum powder, tricalcium orthophosphate, magnesium stearate, aluminum stearate, calcium stearate, calcium carbonate, magnesia, polyethylene glycol, powdered cellulose, Compritol 888 ATO, stearic acid, rilanit special, glycerin monostearate, sodium stearyl fumarate or its combination, the consumption of glidant or lubricant is the 0%~10.0% of composition weight, preferably 0.5%~5.0%.
27. Pharmaceutical composition according to claim 24, it is characterised in that described pharmaceutical composition further comprises diuretics；The diuretics is selected from frusemide, ethacrynic acid, bumetanide, Torasemide, Hydrochioro, chlorthalidone, bendroflumethiazide, Cyclopenthiazide, polythiazide, metolazone, indapamide or its combination；The consumption of diuretics is the 0.1%~10.0% of composition weight, preferably 0.5%~5.0%.
28. Pharmaceutical composition according to claim 24, it is characterised in that described pharmaceutical composition can prepare piece agent, capsule, granule.
29. Pharmaceutical composition according to claim 24, it is characterised in that the capsule component proportion is as follows：
30. Pharmaceutical composition according to claim 24, its spy is just that the capsule component proportion is as follows：
31. AHU-377 crystal type free acids described in claim 1, the calcium salt crystal formations of AHU-377 half described in claim 9, the calcium salt trihydrate crystal formations of AHU-377 half described in claim 10, the calcium salt anhydrous crystal forms of AHU-377 half described in claim 11, AHU-377 α-phenylethylamine salt crystal formations described in claim 18, AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation described in claim 19, the pharmaceutical composition described in AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation or claim any one of 24-30 described in claim 20 is preparing the treatment or prevention disease relevant with neutral endopeptidase, it is cardiovascular, purposes in antihypertensive medicine；The antihypertensive medicine is selected from anti-accelerated hypertension, essential hypertension, renovascular hypertension, diabetic hypertension, simple systolic hypertension or other secondary hypertension medicines.
32. AHU-377 crystal type free acids described in claim 1, the calcium salt crystal formations of AHU-377 half described in claim 9, calcium salt trihydrate crystal formations of AHU-377 half described in claim 10 or, profit requires the calcium salt anhydrous crystal forms of AHU-377 half described in 11, AHU-377 α-phenylethylamine salt crystal formations described in claim 18, AHU-377 (R)-(+)-α-phenylethylamine salt crystal formation described in claim 19, the pharmaceutical composition described in AHU-377 (S)-(-)-α-phenylethylamine salt crystal formation or claim any one of 24-30 described in claim 20 is preparing the purposes in treating or preventing acute and chronic heart failure or harmful vascular remodeling medicine, it is preferred that, the acute and chronic heart failure is selected from congestive heart failure, left ventricle dysfunction, hypertrophic cardiomyopathy, diabetes cardiomyopathy, supraventricular and heart ventricle arrhythmia, atrial fibrillation, auricular flutter.
33. Pharmaceutical composition according to claim 24, it is characterised in that described pharmaceutical composition also includes Angiotensin Ⅱ receptor antagonist.
34. Pharmaceutical composition according to claim 33, characterized in that, the angiotensinⅡantagonist is selected from Losartan, Irbesartan, Olmesartan, Telmisartan, Valsartan, Azilsartan, Candesartan, Eprosartan, Losartan, smooth husky Puli's sand, Elisartan, Tasosartan, Ai Lishatan or its officinal salt；It preferably is selected from Valsartan, Azilsartan or its officinal salt.
35. A kind of pharmaceutical composition described in any one of claim 33-34 is preparing treatment or prevention miocardial infarction and its sequelae, atherosclerosis, angina pectoris, diabetic keratopathy or non-diabetic renal insufficiency, secondary aldosteronism, primary or secondary pulmonary hypertension, nephrosis, glomerulonephritis, chorionitis, glomerulosclerosis, the albuminuria of Primary Nephrosis, renovascular hypertension, diabetic retinopathy, antimigraine, peripheral vascular disease, Raynaud's disease, the hyperplasia of chamber, cognitive dysfunction, purposes in glaucoma or apoplexy medicine.