CA1284994C - Cephalosporin salts and injectable compositions - Google Patents

Abstract

Cephalosporin Salts and Injectable Compositions Abstract of the Disclosure Crystalline sulfuric, di-nitric, mono-hydrochloric, di-hydrochloric, and di- and sesqui-orthosphosphoric acid addition salts of 7-[.alpha.-(2-aminothiazol-4-yl)-.alpha.-(Z)-methoxyimino-acetamido]-3-[(1-methyl-1-pyrrolidinio)-methyl]-3-cephem-4-carboxylate are stable even at elevated temperatures. The crystalline sulfuric acid addition salt is made by forming an admixture of (a) at least one molar equivalent of sulfuric acid with (b) zwitterion in an amount so as to be present in the admixture at a concentration of greater than 25 milligrams/ml, causing crystallization, separating the crystals, washing and drying. The crystalline monohydrochloride, dihydrochloride, and orthophosphate salts are prepared by dissolving the zwitterion in the appropriate amount of acid, causing crystallization by adding acetone and isolating the crystal. Physical admixtures of the salts with certain bases in proportions to give a pH ranging from about 3 5 to about 7 on dilution with water provide injectable compositions on dilution.

Brief Description of the Drawing Figure 1 is a graphical representation of the infra red absorption spectrum of crystalline 7-[.alpha.-(2-aminothiazol-4-yl)-.alpha.-(Z)-methoxyiminoacetamido]-3-[(1-methyl-1-pyrrolidinio)methyl]-3-cephem-4-carboxylate sulfate salt measured on a KBr dilution thereof. BMY-28142.H2SO4 - IR Spectrum.

Description

~21~34994 ~echnieal rield ~hit ~nvent$on i~ direeted to temperature table emi-~ynthetic cephalo~porin calt~ whoee preparation ba~ not been de~cribed in the literature, to the preparation of ~uch salts, and to admixtures containing thece calte Backqround Of ~he lnvention Aburaki et al ~ S Patent No. 4,406,899 di~closes 7-I~-(2-aminothiazol-4-Yl)~ 2)-nethoximinoacetamido~-3-~ methyl-1-pyrrolidinio)-methyl]-3-cephem-4-carboxyl~te in the zwitterion form and mentionc corresponding cid addition ~alts (which are pre~ent in the 2witterion form in injectable composi-tions) and chows t~Dt the zwitterion form has broader spectru~
activity than ceftazidime nd cefotaxime.

However, the aforementioned A~uraki et al cephalosporins re table only for a few hour~ a~ ~njectable composition~ and the zwitterion form even as a dry powder i~ un6table at room temperature and 106e~ 30~ or more of it~ activity on ~torage at clevated temperature~ (e g. 45 deg C nd above) for even one week and therefore reguires ~pecial in-ulated packaging and/or refrigeration nd i~ t a p-ckag~ng and torage di-advantaqe comparcd to ceftazidime ~nd cefot~xime.

While Aburaki et al. mentions acid addition calts, the patent does not ctate how to make tbe~e or rtate which if any of these ~alts have good rtability in dry powder form ~Xessler et al., ~Compari~on of a New Cephalo~porin, BMY 28142, with ~ther Bro~d-Spectrum ~-Lactam A~tibiotic~, Antimicrob~al Aqents and Chemothera~v, Vol 27, No. 2, pp. 207-216, February l9B5 mentions the ~ulfatc ~alt, but doe- not di~clo~e how to prepare ruch or th~t thi~ ~alt has room temperaturc tability nd good elevated temperature rta~illty in ~ry powder form ~ ` lZ84994 ~umm~rv Of The Inventlon It ha- been d~-covered herein that certain eryctalline acid ddition zalt~ of ~-[~-~2-aminothiazol-4-yl)-o-~z)-methox~1m1no~cet~1do~-3-l~l~methYl-l-pyrrolidinio~-mcthyll-3-cephem-~-carboxylate in dry powder form have excellent room temperature ~tability and bave euperior elevatea temperature etability eompared to the zwitterion form The term ~dry powder form~ a5 used herein means a moisture content of leee than 5~ by weight The~e acid addition calt- are the cry~talline eolt~ of 7-l~-(2-aminothiazol-4-yl~-~-(Z)-methox~iminoacetamido~-3-t~l-metbyl-l-pyrrolidinio)~methyll-3-cephem-4-carboxylate eelected from the groùp consi-ting of the ulfuric, di-nitric, mono-bydrochloric, ~nd di-hydrocbloric acid addition ~alt~ and _; orthophosphoric cid ddition ealte (l 5-2 molee of orthophospboric cid per mole of alt, e g range of from the ~c~qui- to the di-orthophosphoric acid ralte), or ~olvate~
thereof The term ~cry~talline~ is ueed berein to mean at least come characterizing rrangement of molecule~ While the :D rulfuric, di-nitric, di-bydrochloric and ortbopho~pboric acid ddition ealte herein are prepared in clearly cryetalline form ~-e evidenced by birefringence under polarizinq micro~cope) with precire rrangement of molccules, the mono-hydrochloric acid daition ealt hae been prepared only with ome regularity in tbe ~~ arrangement of ite moleeules ~ae evidenced by poor birefringence under polarizing microecope) ana not preci~e predictable arrangement and thue ie ~poorly~ cryetalline The term ~cryetalline~ ie ueed herein to embrace not only the clearly cryetalline ralt- but leo the ~poorly~ appearing cry~talline _D ~ono-hydrochloric cid addition ~alt ~ he acid ddition altc herein when formed into queou~
injectable compo~ition- provide the zwitterion in ~olution The zwitterion bae the rtrueture C~ :~ S
N ~ ~ C~2-N

~ O-C~,~ C~De C~

The bro-d cpee~rum utility gainst variou~ organicms of t~e zwitterion form, nd thu- ~f queouc compo6ition~ made up from t~c ~alt~ ~ere~n, is ihown by the data in Abura~i et al ~ S
4,~06,899 S Aqueous compocitions made up from the acid addition fialts hercin cimply by the addition of cterile water provide acidic ~olutione whic~ provoke unacceptable irritati~n on intravenous dministration to rabbitc ~nd unacceptably painful eensation on intr muscular administration to rabbits ~he culfuric acid and di-nitric acid addition calts bave reduced colubilities which are insufficient for typical injectable compositions It bas been found herein that these objectionable characteri~tics are overcome by utilizing t~e ~alts herein in pbysical admixture (that ic as an admixture of colids) with a pharmaceutically cceptable n~n-toxic organic or inorganic base in proportions to provide a pH of about 3 5 to abDut 7 on dilution with water to a rwitterion activity of from 1 mg/nl to 400 mg/ml, normally 250 ~g/nl (as determined by high pe~formance liquid chromatography, ~ereinafter HPLC) A preferred Eialt herein is the crystalline culfuric aeid ddition calt It is preferred because its lo~ colubility in water ~25 mg/ml) allows high recovery from aqueous medium on cryctallization ~he crystalline ~ulfurie acid addition calt is readily preparcd by a process compri~ing the teps of la) forming an aqueous admixture of ~i) at least 1 molar equivalent of culfuric acid and (ii) zwitterion in an a~ount co as to be present in t~e adnuxture ~t a concentration greater tban 25 mgtml, (b) causing cryctallization of the culfuric acid addition calt to occur, and (c) ~colating erystalline culfuric acid addition calt `_ "~,!

Detailed DeccriDtion The cryctallihe c~ltc herein Ihereinafter referred to cimply c the salt~ ~erein) have excellent etability at room temperature ~nd bave ~ potency loss las determined by HPLC), of le~s than 1 on ztorage for a mont~ at room temperature ~hese ealt~ also bave excell nt stability ot elevated temperatures ~nd have a potency loss (a~ determined by HPLC) of less than 15~ on stor~ge for a month at ~5-56 deg C

The culfuric acid additiDn alt is a preferred ~alt herein lD It has a potency loss of less t~an 10~ on ~torage for a month a ~5-56 deg C Very importantly, it has a low olubility in water, i e ~bout 25 mg/ml, and therefore i5 crystallized from water with minimized residual loss.

~ he di-nitric acid addition ~alt herein alco has a 1DW
colubility in water, i e about 60 mg/ml, ~nd therefore also provides low residual loss on crystallization from water The mono-hydrochloric, di-hydrochloric ~nd sesqui- or di-orthophotphoric acid addition calts have water colubilities greater than 200 mg/ml , ~nd therefore are preferably crystallized from orqanic ~olvents, rather than from water, in order to obtain goDd yieldc We turn now to the preparation of the sslts herein As previously indicated the ~ulfuric acid addition calt berein is prepared by a process comprising the teps of (a)forming an aqueous admixture of (i) at least one molar equivalent of zulfuric acid and (ii) zwitterion corresponding to caid calt in an ~mount ~o ac to be present in the admixture at ~ concentration of greater than 25 mg/nl, (b) causing erystallization to occur, ~nd (c) ~solating crystalline culfuric acid ~ddition salt ~21~9~4 .
Preferably the ~witterion is used ~n step ~a) ~n an um~unt eD a~
to be pre-ent in the dmixture t concentration ranging from ab~ut 100 mg/ml to b9ut 200 mg/ml, nd tep (b) ls carried out ~n an aqueou~ medium free of organic solvent. Normally no more than 2 molar cquivalent6 of eulfuric ac~d are utilized in ctep la). Normally rwitterion ic u5ed in etep (a) in an amount so as to be present in the admixture at a concentration less than 500 mg/ml.

Step (a) ie readily c~rried out either by addin~ solid zwitterion to ~ulfuric acid solution (e.g. lN H2S~4) with rapid etirring to form a ~olution. Alternatively step (a) can be carried by diseolving solid zwitterion in water and slowly adding ~ulfuric acid with stirring to form a ~olution.

Step (b) ie carried out by inducing crystallization, preferably by ~eeding, and then slurrying, preferably for 15 minutes to 2 ho~rs. It is preferred that this crystallization etep be carried out in aqueous medium, free of organic ~olvent, ~nd in such ea~e purities greater than 98~ are normally obtained.
While the pre6ence of organic olvent, euch as acetsne, fosters crystalliiation and increases yield by lowering the ~olubility of the formed sulfurie acid addition ealt in the cryst~llization medium, it also can foeter precipitation of impurities resulting in decreased purity. When the zwitterion i~ used in step (a) in an amount o as to be present in the admixture in an amount less than 25 mg/ml, orqanic solvent, preferably acetone, must be included in the cry~tsllization medium to provide reasonable recovery. When acetone is used, it i5 appropriately used in amounte of 0.5 to 10 volumes per volume of ague~us cry~tallization medium.

Step (c) i~ carried out by eeparating the cry~tals from the crystallization ~edium, preferably by vacuum filtration, then ~ashing e.g. with acetone/water followed by acetone alone or O.lN
~ulfuric acid (e.g. 1/10 volume) followed by acetone le.g. 1/4 ` - ~2~ 3i9~
volume), ~nd then drying, g by v-cuu~ ~rying t 30-50 de~ C
for ~-20 hour~

The ~et~Dd herein ~or forming the ~ulfuric ~cid ddition ~lt rc~ults in the purificati~n of the ~itterion form because S of the limited ~olubility ~ the au~furic acid eddition ~lt compared to the zwitterion form nd can be used to purify ~witterion without icolating it ~c ~olid If it i~ desired to obtain ~ubstantially pure zwitterion (free-~ase) from the formed ~ulfuric acid addi~ion calt, thi~ can be carried out by di~o~ving the ~alt in water, edaing ~a~H)2 8H2O in an am~unt of 90-100~ of theory ~t a p~ of le-~ than 6 5 to precipitate ~aS~4, filtering to remove the BaSO4 nd recovering the iltrate containing the rwitterion di~solvea therein and utilizing it as a rolution or i~olating ~olid zwitterion (free-base) by lyophilizinq it or by dding ~cetone to precipitate amorphous z~itterion followed by i~olating ~olid zwitterion by vacuum filtration, washing e g wit~ aeetone, and vacuum dryinq _ Alternatively, the ~ulfuric acid ~daition alt i- converted to the free-base ùtilizing ion exchange re~ins, g Dowex WGR (a weak ba~e anion exc~ange re~in) and Dowex XU-40090 01 (a ~trong acid cation exchange resin) with ~ub~eguent lyophilization Turning now to the preparation of thc cryttalline di-nitric cid addition alt herein, thic ~ obtained by admixinq ~i) at lea~t two molar equivalen~ of nitric cid and lii) zwitterion corre~ponding to taid ~alt ~o to be pre~ent in the admixture at a concentration greater than 100 mg/ml, and then indueinq cry~talli~ation by ~eeding or rub~ing with a glass rod, dilutinq with 2-propanol ~nd eooling The cry~talline di-nitric acid ddition ~alt i~ recovered e g by filtering, wa~ing requentially, e g with 2-propanol-h2O ISD~ v/v), 2-propan~l, and ether, and then vacuum drying ~t 50 deg C for 2 hour~

The mono-hydrochlorie cid ddition ~alt ~erein i~ prepared ~y di~solvinq ~witterion in ~pproximately one molar eguivalent of _?_ *Trade Mark !

hydrocbloric cid nd c~using cry~tallization by ddinq cetone witb stirring and eontinuing to ctir, followed by i-31ating cry-talc, e.g. by vacuum filtration followed by ~a-hinq with cetone nd vacuum drying. Alternatively the mono-hydrochloric cid ddition salt is formed from the di-hy~rocblorlc cid ddition salt by slurrying the di-hydrochloric cid adaition salt in methylene chloride ~n~ ~dding 1 mole eguivalent of triethylamine followed by slurrying to form the mono-~ydrochloric cid addition talt ~hich is isolated, e.g. by vacuum filtration, followed by washing wit~ met~ylene cbloride and vacuum drying.

The crystalline di-hydrochloric acid eddition c~lt herein is prepared by dissolvins zwitterion in at least t~o molar equivalents of hydrochloric acid, then causing crystallization by adding acetone, then isolating crystal~ e.g. by vacuum filtration, wa~hing with ~cetone and vacuum dryinq.

~ he crystalline di-orthophospboric acid addition salt herein is prepared by dissolving the zwitterion in at least 2 molar equivalents of phosphoric acid, causing crystallization by ~dding acetone, ~nd isolating crystals by e.g. by vacuum filtration followed by washing first with acetone and tben ~itb ether and then vacuum drying. The crystalline eesqui-orthopbosphoric acid addition salt i5 formed by this same procedure except that about 1.5 molar equivalents of phosphoric acid is used.

~he salts berein are formed into injectable compositions by diluting witb sterile water ~nd buffering to a pH of 3.5-7 to form an injectable concentration of 1 mg/ml up to 400 mg/ml of zwitterion. Suitable buffering agentC include, for example, tri~odium ortbopbosp~ate, sodium bicarbonate, sodium eitrate, N-metbylglucamine, L~) lysine ~nd L(~) arginine. For intramuscular or intravenous administration to ~n sdult human, a total dossge of from ~bout 750 to about 3000 mg per day in divided dose~ ic norm~lly sufficient.

9a~
~ he c~lt6 herein re not desirably formed into ln~cctable compo~itions simply by the addition of terile wates becau-e the sulfuric ~nd di-nitric acid addition altc are not ~ufficiently soluble to form compositions of normal concentration for admini~-tr-tion and because the caltc herein when dit~olved provide very low pH compositions (1.8-2.5) which provide painful eensation on injection. As indicated above, it ~a8 been found herein that these chortcomings are overcome by forming the calts herein into a phy~ical, i.e. colid, admixture with pharm~ceutically acceptable, norm3lly colid non-toxic organic or inorganic bases in proportions to provide a pH ranging from a~out 3.5 to ab~ut ~, preferably from about 4 to about 6, on dilution of the admixture with water to injectable concentration of 1 mg/ml up to ~00 mg/ml of zwitterion, e.g. zwitterion activity of 250 mg/ml as determined by HPLC assay.

~ he exact proportior.s of ingredients in the physical admixture vary from lot to lot of the salt since the purity of the calt varies from lot to lot. The proportions of ingredients are established for a particular lot by pretitrating in respect to a cample to obtain a selected pH within the aforementioned range.

The physical admixture is readily stored and shipped in colid form thereby taking advantage of the ~tability of the salts herein and i~ readily converted into an injectable composition simply by addition of water, e.g. by a nurse or doctor,just prior to use.

~ he physical admixture is prepared by blending the calt and the base into a uniform blend, e.g. utilizing a ~tandard blender $n a dry atmosphere, ~nd is then preferably filled into a vial or other container, all under aseptic conditions.

The bases for u~e in the admixture include, for example, tri~odium orthophosphate, codium bicarbonate, codium citrate, N-methylglucamine, Ll+~ lysine and L(+) arginine. L(+) lysine _9_ 9~

and L(~) a~ginine are preferred ince admixture~ eontaining t~ese re reconstituted to provide injectable composition6 which on ~njection provide ~es~ pain in animals than compo~itions derived from admixture~ containing other ba~e~. ~he L(~) ly~ine i6 very prcfer~bly utili2ed in a proportion to provide pH of 3.5-6 on dilution of the admixture with water to provide a composition with a zwitterion activity of 250 ns/ml (as determined by HPLC
assay).

~he salts herein and ~ubstantially dry physical admixtures containing them can be ~tored wit~out refrigeration Dr insul2ted packaging and still retain bigh potency.

In several of the preparations herein the unsta~le zwitterion i~ used as the tarting material. ~he preparation of this is described in Examples 1-3 of Aburaki et al. ~.S.
4,406,899. ~he zwitterion is referred to in Aburaki et al. as 7-l(2)-2-methoxyimino-2-(2-aminothia201-4-yl)acetamido-3-l(l-met~yl-l-pyrrolidinium)methyl]-3-cephem-4-carboxylate.

~he invention i5 illustrated in the following w~rking examples.

Example I
PreParation of the Sulfuric Acid Addition Salt 1.5 9 of zwitterion ~re added 610wly to 10 ml o~ rapidly 6tirred lN ~254 (1.59 molar equivalents) at 20-26 deg. C. A
solution is obtained. Crystalliz~tion i6 then induced by seeding with crystalline 6ulfuric ~cid addition calt and the cryctalline mass is elurried for 0.5 hours. ~he cry~tals ~re then separated by vacuum Siltration, washed with 3 ml of 50~ aeetone/water ~V/V) ~nd with two 5 nl portions of acetone, ~nd vacuum dried at 40-50 deg. C overnight.

~ ~ i A typ~l y1eld 1~ 1.3 9 of ulfuric cid addition alt.
Analys~s Calculated for ClgH2~N6O5S2 H2SO4 ~C, 39.44; ~H~
4.53; ~N, 14.52; ~S, 16.62~ ~H2O, none. Found: ~C, 38.91; ~H, ~.57s -N, 14.64; tS, 16.71s ~H2O, 1.42.

ExamDle II

Pre~aration of the Sulfuric Acid Addition Salt 1.5 9 of 2witterion ~re dissDlved in 5 ~1 of ~ter. 5 ~1 of IM 112S04 ~re slowly ~dded to this solution with stirrinq.
Crystallizntion i~ then induced by seeding Wlth crystalline acid addition salt and the crystalline mass is lurried for 0.5 hours.
The crystals are then eparated by vacuum filtration, washed with ~ ml of 50~ acetone/water (V/V) and with two 5 ~1 portions of acetone, and vacuum ~ried at 40-50 deg. C overnight.

A typical yield i~ 1.3 9 of Eulfuric acid addition salt.

lS Example III
PreParation of the (HNO~)~ Acid Addition Salt 300 mg of zwitterionare dissolved in 2N nitric acid (0.5 ml). The solution i~ ru~bed with a qlass rod, diluted with 2-propanol (0.4 ml) and cooled. The crystalline title compound is collected and i~ ~equentially washed with 0.4 ml of 2-propanol H2O ~1:1), 2-propanol ~nd then ether to afford 127 mgs of the dinitrate salt.

Analysis: Calculated for ClgH24N6O552~2H~O3 ~C, 37.62;
~H, 4.32; ~N, 18.47; ~S, 10.57. ~ound: ~C, 36.92; ~H, 4.10;
~N, 18.08; ~S, 10.67; ~H2O content 0.90~).

.

$~34 . .
Example IV

Preparation of the Monohvdrochloride Acid Addition Salt 1 g of 2witterion is dis~olved in 2.08 ml of l~ HCl (1 molar equivalent) at 20-25 deg. C. Thirty ml of aceto~e are added with rapid rtirring over a 15 minute period whereby crystal5 form.
Stirring is continued for 1 hour. ~he crystals are isolated by v~cuum filtration, washed with 10 ml of acetone and va~uum dried at 50 deg. C for 2 hour~.

A typical yield is 0.9 9 of crystalline monohydrochloride ~alt. Analysis: Calculated for ClgH24N60552.HCl: ~C, 41.37;
~H, 4.75; ~N, 15.2; ~S, 11.63; ~Cl, 12.B6. Eound: ~C, 39.32;
~H, 4.88; ~-N, 13.95; ~S, 11.28; ~Cl, 12.44; ~H20, 4.5.
(Corrected for H2~: tC, 41.17; ~, 14.61; ~S, 11.82; ~Cl, 13.03).

Exam~le V
~reparation of the Dihydrochloride Acid Addition Salt and Preparation of the Monohydrochloride Acid Addition Salt Erom It 350 mg of zwitterion are dissolved in 2 ml of IN-HCI. 10 ml of acetone are added to the resultant solution, with rapid stirrinq and over a 5 ~inute interval, whereby crystals form. Stirrinq is continued for 5 additional minutes. Then 10 additional ml of acetone are added and stirrin~ is carried out for 0.5 hours. The cry~tals are removed ~y vacuum filtration, washed with two 5 ml portions of acetone ~nd vacuum dried at 40-45 deg. C for 24 ~ours.

A typical yield iS 300 my of crystalline dihydrochloride acid addition ~alt. Analysi~ Calculated for 128499a~
C19~2~N6O5S2.2HCl: ~C, 41.38; H, 4.~5; ~N, 15.2: tS, 11.62 ~Cl, 12.B. Fo~nd: ~C, 40.78; ~H, 4.98; ~N, 14.7; ~S, 11.25;
~H20, 1. 2S. (Corrected for H2O: ~C, ~1.1; ~N, 14.8B:
~S, ~1.39; ~Cl, 11.94) 1 9 of dihydrochloride s21t prepared as aboYe is lurried in 20 ml of methylene chloride at 20-25 deg. C in a sealed flask and 0.28 ml of triethylamine is added over a 15 minute inter~ he cryst~lline mass is then slurried for 5 hours. ~he resultant monohydrochloride crystals are then isolated by vacuum filtration, washed with two 5 ml p~rtions of methylene ehloride and vacuum dried ~t 50 deg. C for 2 hours. A typical yield i6 800 m~.

ExamPle Vl Preparation of the Di-orthop~osphoric Acid Addition Salt 1 g of zwitterion is dissolved in 3.4 ml ~f 144 mg/ml PO4 ~2.2 m~lar eguivalents) at 15 deg. C. The resulting solution is suitably filtered to clarify it. 12 ml of acetone are ~dded to the clarified solution, with raDid stirrino and over a 10 minute period,whereby crystal5 form. Stirring is continued for 10 minutes. ~hen 30 ml of acetone a-eadded over a 10 minute period, and stirring is continued for an additional 15 minutes.
~he crystals are collected by vacuum filtration, washed with two 5 ml portions of acetone and two 5 ml portions of ether and dried ~nder high va~uum for 16 hours.

A typical yield for this type of preparation was 1.1 9 of cryctalline di-orthophosphoric acid ~ddition salt. Analyeis:
Calculated for ClgH24N6~5S2 3 4 ~, 12.42. ~ound: ~C, 33.43; ~H, 4.65; ~N, 12.02; ~H2O, 1.82.
(Corrected for H2O: ~C, 34.0; tN, 12.2).

~2134994 ~ e ~esqui-ort~opho-p~oric cid ddition alt ic formea as above xcept that 1.5 olar equivalents of H3P~4 are u5ed inttead of 2.2 ~olar equivalentc.

S ExamPle VII

Stabilities at Elevated TemDeratures Elevated temperature stabilities were determined by toring the preparations in dry container~ at temperatures and for time periods as denoted below and potency losses or gains were determined by ~PLC. A ~ potency qain is indicated by a plus sign in front of a figure. A less than 10~ potency loss over a 2 to 4 week period at 45-56 deg. C i~ usually indic~tive of less than 10~ potency loss over a 2-3 year period at room temperature.

9~3~

PERCENT LOSS

J5 Deg. C 56 Deg. C 100 Deo . C
(Weeks) IWeeks) (Days) Form 1 ~ _ 6 1 2 4 zwitterion 3~ ~ 71 _ 57 _ _100 ~2SO4 Salt 12.q to ~. ~ ~S 1 45 ~o ~6 ~3 0 to ~6 0-10 (NNO3)2 Salt8.8 3.4 0.6810.3 3.7 2.4 _ _ HCl Salt 4.~ ~ 6 0 6 4 6.4 _ _ lHCl)2 Salt 0 ~ 7 ~ _ 0 _ 7 2 12.4 I0 (H3P4)2 Salt j ~ ~ _ 2.7 5.C _ -' ExamPle VIII

Testinq of Physical Admixtures Physical admuxtures were made up of crystalline sulfuric acid ~alt with (a) trisodium orthophosphate, (b) sodium bicarbonate, Ic) L(~) lysine, and (d) L(~) arginine. The bases were added in proportions to provide pH'~ on dilution of the dmixture with water to a zwitterion activity of 250 mg/ml (as determined by NPLC acsay) as follows: trisodium orthophosphate (to provide a pH of 6.0): codium bicarbonate (to provide a pH of 6.0); L(~) lysine (to provide a pH of 6.0); L(l) arginine (to provide pH of 6.0). ln~ect-ble compo~itiont were made up by rcconttituting witb sterile water to rwitterion activity of 250 mg/ml ac determ$ned ~y BPLC scay. There were no solubility problems. Injections ~100 mg/kg) were carried out intramuscularly ~n rabbits with pain within acceptable threcholds. The least pain was with the arginine containing compo~ition.

Similar results of good ~olubility and acceptable pain on intramuseular injection are obtained on use of the other ~alts herein in the physical admixtures with the above bases.

~ igure 1 i~ the infra red absorption spectrum of the cry~talline ~ulfate ~alt prepared as described in Examples I or II pelletized in the crystalline form with potassium bromide.

The X-ray powder diffraetion pattern of the crystalline ~ulfate salt of 7-la-(2-~minothiazol-4-yl)-a-(z)-methoxyimuno-acet~mido~-3-[(1-methyl-1-pyrrolidinio)-methyl~-3-cephem-q-carboxylate prepared as de~cribed in Example I or II was determined with a Rigaku Powder Diffractometer using a copper target X-ray tube, a nickel filter, and the sample contained in a glass dich. The scan rate was 2 deg./min. over the range from 5 deg. to 40 deg. and a chart was mechanically recorded to ~h3w the ngles of maximum diffraction. ~rom this the (d) spacings and rclative intensities (I/I) were calculated. ~hey are listed bel~w.

~2~4994 d ~Pac~nq (A) I/I 1-) 9. 20 lOD
6.80 50 5.50 2B
5.09 22 q.50 38 4.41 ~4 .lg 63 3. 78 38 103 . 64 ~4 3.39 ?s 3.31 31 3.15 47 ~z1~4994 EXAMPLE IX
PREPARATION OF THE SESOUIPHOSPHATE SALT

The zwitterion, 0.70 g., is dissolved with rapid stirring in from 2.2 to 2.4 ml. of 85% phosphoric acid ~2.1 to 2.2 molar equivalents) which has been diluted 1:10 (v/v) with water. The solution is clarified by filtration through a 0.22-0.45 micron pore-size membrane filter. From 5 to 7 parts by volume (15-20 ml) of methanol is added to the filtrate with rapid stirring during a 30 to 60 min. period.
Crystals form during this operation, and rapid stirring is continued for 1.5 to 2 hours. The crystalline product is recovered by vacuum filtration. The product is washed on the filter first with 6 to 8 ml of 1:1 (v/v) methanol:acetone taking care to maintain a tightly packed filter cake, and then with acetone. The product is dried in vacuo at 50 C for 2 hours; typical yield 0.7 to 0.75 g.

Infrared Interpretation (See Figure 2) (IR, KBr pellet) Peak Position (cm 1) Functional GrouP
2800-3400 NH,NH3 ,carboxyl OH
1780 ~-lactam C=O
1680 Carboxyl C=O
1660 Amide C=O
1630 C=N,C=C
1550 Amide OH
980,1040 PO4=

~2~4~94 .

Behavior On Heatinq An exotherm is shown at 171.8C in the differential scanning calorimeter tracing.

X-Rav Diffraction Pattern The X-ray powder diffraction pattern of the foregoing sesquiphosphate salt was measured with a Rigaku Powder Diffractometer in the same fashion as described above with respect to sulfate salt with the following results.

d I/I~
l1.0~ - 32 9~2 - 16 7.89 - 24 7.02 - 42 6.7 - 32 5.5 - 26 4.64 - ]00 4.45G - S3 4.3 - 58 3 75 - ~9 3.56 - ~]
3.3] - 2G
3.05 - ~G

~8~

N ~ Interpretation ( H 90 MHz NMR, D2O solution) 2-_ .
N ~OC~, ~ C~2 ' Chemical Shift (PPm ~ vs. TSP) DescriPtion Inteqral Assiqnment 2.0-2.4 Multiplet 4 14CH2, 14'CH2 3.04 Singlet 3 12CH3 3.3-3.6 Multiplet 5 ~ 2' 13 CH2 3.94 Doublet 1 2CH
4.12 Singlet 3 20CH3 4.12 Doublet 1 llCH
4.8 Doublet 1 llCH

5.42 Doublet 1 6CH
5.88 Doublet 1 7CH
7.21 Singlet 1 18CH

~4~94 .

Stability Time-Tcmperature ~ I,oss 1 day; 100C - 10.9 3 days; 70C - o 7 days; 70C - 1.9 1 week; 56C - 1 0 2 weeks; 56C - l q 4 wecks; 56C - 0 1 week; q5C - 0 2 weeks; qSC - 1 4 4 weeks; 45C - 0 7 8 weeks; 45C - 1 6 1 month; 37C - 2 5 Elemental Analvsis ~percent bv weiqht) Found Drv Basis Theory (SesquiPhosPhate) C 35.44 36.3 ~6.4 H 4.66 4.41 . 4.7 N 12.88 13.2 13.4 H2O 2.29* -monohydrate = 2.8% H2O
H3PO4 23.06 23.6 23.6 *Karl Fischer Method

Claims (16)

1. Temperature stable crystalline salts of 7-[.alpha.-(2-aminothiazol-4-yl)-.alpha.-(Z)-methoxyiminoacetamido]-3-[(1-methyl-1-pyrrolidinio)-methyl]-3-cephem-4-carboxylate selected from the group consisting of the sulfuric, di-nitric, mono-hydrochloric, and di-hydrochloric acid addition salts, and orthophosphoric acid addition salts containing 1.5-2 molar equivalents of H3PO4, or solvates thereof.

2. The crystalline salts of claim 1 selected from the group consisting of the sulfuric, mono-hydrochloric, di-hydrochloric and orthophosphoric acid addition salts, or solvates thereof.

3. A physical admixture of the salt of claim 1 with a pharmaceutically acceptable non-toxic organic or inorganic base in proportions to provide a pH of about 3.5 to about 7 on dilution of the admixture with water to injectable concentration.

4. A physical admixture as recited in claim 3 wherein the salt and the base are present in proportions to provide a pH of about 4 to about 6 on dilution of the admixture with water to injectable concentration.

5. A physical admixture as recited in claim 4 wherein the base is L(+) lysine.

6. A physical admixture as recited in claim 4 wherein the base is L(+) arginine.

7. A method of making the sulfuric acid addition salt of claim 1 comprising the steps of (a) forming an aqueous admixture of (i) at least 1 molar equivalent of sulfuric acid and (ii) zwitterion corresponding to said salt, (b) causing crystallization of the sulfuric acid addition salt to occur, provided that when the zwitterion is present in the admixture at a concentration of less than 25 mg/ml the crystallization is carried out in the presence of an organic solvent, and (c) isolating crystalline sulfuric acid addition salt.

8. Method as recited in claim 7 where step (b) is carried out in an aqueous medium free of organic solvent.

9. The method as recited in claim 8 wherein the zwitterion is used in step (a) in an amount so as to be present in the admixture at a concentration less than 500 mg/ml.

10. The method as recited in claim 7 wherein the amount of zwitterion employed in Step (a) is such that it is present in the admixture at a concentration greater than 25 mg/ml.

11. The method as recited in claim 8 wherein the zwitterion is used in step (a) in an amount so as to be present in the admixture at a concentration ranging from about 100 mg/ml to about 200 mg/ml.

12. The crystalline sulfuric acid addition salt of claim 1.

13. Crystalline 7-[.alpha.-(2-aminothiazol-4-yl)-.alpha.-(Z)-methoxy-iminoacetamido]-3-[(1-methyl-1-pyrrolidinio)-methyl]-3-cephem-4-carboxylate sulfate salt having the following X-ray powder diffraction pattern:

14. The crystalline orthophosphoric acid addition salt of Claim 1, and hydrates thereof.

15. Crystalline 7-[.alpha.-(2-aminothiazol-4-yl)-.alpha.-(Z)-methoxyiminoacetamido]-3-[(1-methylpyrrolidinio)-methyl]-3-cephem-4-carboxylate phosphate having the following X-ray powder diffraction pattern.

16. A method as in claim 7, 9, 10 or 11 wherein the organic solvent is acetone.