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Stability Constants of Palladium, Copper, Nickel, Zinc and Manganese Chelates of N-Benzoyl-N-phenylhydroxylamine

J . P . C . J A I M N I a n d N . C . S O G A N I

Chemistry Department, University of Rajasthan, Jaipur, India

(Z . Naturforschg. 22 b , 922—924 [1967] ; eingegangen am 20. Januar 1967)

Stabilities of palladium, copper, nickel, zinc and manganese chelates of A-benzoyl-TV-phenyl-hydroxylamine have been determined in 10% v/v dioxan-water mixture by employing B j e r r u m -C a l v i n pn titration technique. The titration medium was maintained at a constant ionic strengih (0.1 M KCl) and at a temperature, 25 + 0.5°. Titrations were carried out in duplicate using 40 : 1 ratio of chelating agent to metal ion concentration.

/V-Benzoyl-./V-phenylhydroxylamine (BPHA) is one of the most interesting of the many organic reagents which have been reported in recent years. A number of procedures have been developed for estimating various metals by using this reagent. Since 1960 three reviews 1 - 3 have been published on this reagent and its analogues. A survey of the literature revealed that stabilities of metal chelates of BPHA have not been studied so far. The present communication deals with the determination of sta-bilities of bivalent metal chelates of BPHA in 70% v/v dioxan-water mixture, by employing B J E R R U M -

C A L V I N 5 pn titration technique.

Experimental

Ligand solution: TV-Benzoyl-A-phenylhydroxylamine was prepared by employing the method given by S H O M E 5. A weighed amount of this compound was dis-solved in 70:30 v/v dioxan-water mixture and diluted to give a 0.04 M solution

Standard palladium, copper, nickel, zinc, and man-ganese solutions: Analytical grades of palladium chloride, copper chloride, nickel sulphate, zinc oxide, manganese chloride were used for preparing standard solutions. After standardising these solutions by usual classical methods, these were diluted to give 0.002 M solutions.

Sodium hydroxide: Approximately 0.1 M carbonate free sodium hydroxide was prepared and standardised with analytical grade potassium hydrogen phthalate. It was diluted to give 0.02 M solution.

Dioxan: The B.D.H. dioxan was purified by W E I S S -

B E R G E R ' S method 6. Potentiometrie method: The PH titrations of the

ligands with standard alkali solution in absence and

1 I. P. ALIMARIN, F. P. SUDAKOV, and B. G . GOLOVKIN, Russian Chemical Reviews 31, 466 [1962].

2 A . M . G . MACDONALD, Ind. Chemist 36. 512 [I960]. 3 A . M . G . MACDONALD, Ind. Chemist 37, 30 [1961].

in the presence of different metal ions were carried out by using Cambridge bench type PH meter which gives values accurate up to 0.02 PH unit. The pn meter was standardised before, and checked after, each titration with buffer solutions of PH 4.00 and 9.27. Electrode system consisted of glass electrode (PH range 1 — 13) and reference electrode as saturated calomel electrode.

10 ml. of 0.04 M ligand solution in 70% v/c dioxan were pipetted in a titrating vessel, requisite amounts of 0.02 M nitric acid was added to it, to lower the PH to about 2. The final ionic concentration was maintained at 0.1 M by adding 5 ml. of 1 M KCl. When titrating in presence of metal ions, 5 ml. of 2 x l O _ 3 M metal ion solution were added at this stage. The total volume of the contents was made 50 ml. by adding varying amounts of dioxan and distilled water in such propor-tions that it finally became 70% v/v dioxan-water mixture. It was titrated against 0.02 M NaOH. The temperature of the solution was maintained at

10

U PH

8

7

6

5

3

2

1

4 M . CALVIN and N . C . MELCHIOR, J . Amer. chem. Soc. 7 0 , 3270 [1948].

5 S . C . SHOME, Analyst 75, 27 [1950]. 6 A. WEISSBERGER and E. S. PROSKAUER, Organic Solvents, Ox-

ford 1935, p. 139.

Ml. 0.02 M NaOH »-

Fig. 1. Titration curves.

25 + 0.5°. The pn meter readings obtained at a given alkali addition in duplicate titrations were reproducible with a maximum variation of ± 0.02 pn unit. The titra-tion curves are shown in Fig. 1.

Calculations: The horizontal distance between the reference titration curve and the curve obtained in presence of a metal ion gives the amount of metal bound ligand. This value divided by the total metal ion concentration gives n. In this way the values of h at different PH values were calculated.

Palladium Copper Pn n p A : Pn n p A

2.95 0.4 10.12 2.85 0.1 10.21 3.00 0.6 10.07 2.95 0.3 10.11 3.10 0.7 9.98 3.05 0.5 10.01 3.40 0.8 9.69 3.35 0.7 9.72 3.75 1.0 9.35 3.70 0.9 9.37 3.95 1.2 9.15 3.90 1.0 9.17 4.10 1.3 9.00 4.10 1.2 8.98 4.35 1.5 8.75 4.45 1.4 8.63 4.70 1.7 8.40 4.55 1.5 8.53 5.00 1.8 8.11 4.70 1.6 8.38 5.15 1.9 7.96 4.85 1.7 8.23

5.25 1.8 7.83

Table I. Formation curve data for metal chelates of A-benzoyl-A-phenylhydroxylamine.

PH

Nickel n p A : Pn

Zinc n p A

5.25 0.1 7.81 6.80 0.1 6.26 5.75 0.3 7.31 7.00 0.2 6.06 5.95 0.4 7.11 7.15 0.3 5.91 6.15 0.5 6.90 7.30 0.5 5.76 6.55 0.7 6.51 7.50 0.8 5.56 6.70 0.8 6.36 7.75 1.1 5.31 6.85 1.0 6.22 7.85 1.3 5.22 7.20 1.2 5.87 7.90 1.4 5.17 7.45 1.4 5.62 8.25 1.7 4.82 7.85 1.6 5.22 8.45 1.8 4.62 8.20 1.8 4.87 8.75 1.9 4.32 8.80 2.0 4.27 9.25 2.0 3.82

Table II. Formation curve data for metal chelates of A-ben-zoyl-A-phenylhydroxylamine.

At any PH, the value of free ligand concentration, [^4®], was calculated from the total ligand concentra-tion, \HÄ], and its ionisation constant. This is based on the assumption that the amount of chelating agent over metal ion is so great that the removal of [HA] by chelation does not cause any significant change in the equilibrium:

H A ^ H ® + A e ,

The pKa value of A-benzoyl-A-phenylhydroxylamine was potentiometrically found to be 10.96.

In calculating the values for n and [ A e ] , the con-centrations were corrected for changes in volume, pro-duced by addition of alkali during titrations. This way

Manganese pn n p A

6.80 0.1 6.31 7.10 0.2 5.96 7.30 0.3 5.75 7.80 0.7 5.26 7.95 0.8 5.11 8.05 1.0 5.02 8.30 1.2 4.77 8.45 1.3 4.62 8.75 1.5 4.32 9.05 1.7 4.02 9.15 1.8 3.92 9.40 1.9 3.68 9.65 2.0 3.42

Table III. Formation curve data for metal chelates of A-ben-zoyl-A-phenylhydroxylamine.

Metal ion log k± log ko log ki ki

Palladium 10.11 8.74 18.85 Copper 10.01 8.53 18.54 Nickel 6.90 5,45 12.35 Zinc 5.76 5.06 10.82 Manganese 5.52 4.32 9.84

Table IV. Stability constants of metal chelates of A-benzoyl-A-phenylhydroxylamine.

a series of values for h and [ A 0 ] , corresponding to different values of PH , was obtained. The results are given in Tables I —III. The formation curves of metal chelates of A-benzoyl-A-phenylhydroxylamine are shown in Fig. 2.

Fig. 2. Formation curves of metal chelates of A-benzoyl-A-phenylhydroxylamine.

The values of log kl, log k2 and log kx k2 for various metal chelates were read directly from the formation curves and are given in Table IV.

Discussion

The consumption of alkali during the course of titration can be due to the ligand, the hydrolysis of

metal ion and the ligand protons liberated in com-plex formation.

The ligands under study is a very weak acid pK a = 10.96. Thus the ligand protons as such are not in a titrable form. Hydrolysis of these metal ions in 70% dioxan-water mixture have been studied 7 and are not likely to interfere in the stabi-lity determinations. Moreover, there was no pre-cipitation during the chelation titrations, ruling out the possibility of hydrolysis of these metal ions in presence of large excess of ligands. Thus, the con-sumption of an excess of alkali in chelation titration over the simple ligand titration is due to the ligand

protons liberated during the complex formation. This may be represented as:

M2® + HA + OHG MA® + H 2 0 , MA® + HA + OH® ^ MA2 + H 2 0 .

The following order of stability of BPHA metal chelates was found. Paladium > copper > nickel > zinc > manganese.

The authors express their gratitude to Professor R. C. M E H R O T R A for providing facilities in the department. They are also grateful to the C o u n c i l o f S c i e n -t i f i c & I n d u s t r i a l R e s e a r c h for granting a research followship to one of them (J. P. C. J A I M N I ) .

7 D . N. PUROHIT and N. C. SOGANI, Naturwissenschaften 5 1 (23), 553 [1964].

Massenspektrometrie von Cysteinpeptiden E R N S T B A Y E R , G Ü N T H E R J U N G u n d W I L F R I E D K Ö N I G

Chemisches Institut der Universität Tübingen

( Z . Naturforschg. 22 b , 924—931 [ 1 9 6 7 ] ; eingegangen am 18. Februar 1967)

Zur massenspektrometrischen Sequenzbestimmung von Cysteinpeptiden eignen sich die S-Benzyl-A-Trifluoracetylpeptidäthylester. Verschiedene Di-, Tri- und Tetrapeptide mit Cystein sind neu dar-gestellt und deren Massenspektren eingehend untersucht worden. Eine Feststellung von N- und C-terminalem Cystein sowie der Position von Cystein in einer Peptidkette ist leicht möglich. Die Gas-Chromatographie einiger Dipeptide wird mitgeteilt.

Im Rahmen der Synthese von Peptiden aus der Sequenz des Ferredoxins mittels klassischer Metho-den und an fester Phase nach M E R R I F I E L D 1 war die Sequenz und die Konstitution der insbesondere Cystein enthaltenden Peptide zu überprüfen. Hierfür erschien die Massenspektrometrie der V-Trifluor-acetyl-Peptidester geeignet, die von W E Y G A N D 2 ' 3

und S T E N H A G E N 4 ausführlich untersucht worden ist. Obwohl bereits viele Peptide mit neutralen und

sauren Aminosäuren massenspektrometrisch unter-sucht worden sind, fehlen bisher eingehendere Stu-dien über Cysteinylpeptide. Bei der V-Trifluoracety-lierung und Veresterung von Cysteinylpeptiden laufen durch H2S-Eliminierung und Disulfid-bildung unerwünschte und unkontrollierbare Neben-reaktionen ab. W E Y G A N D 5 hat daher die Entschwefe-

1 G. R . MARSHALL U. R . B. MERRIFIELD, Biochemistry 4 , 2394 [1965],

2 F . WEYGAND, A. P R O X , W . KÖNIG U. H. H. FESSEL, Angew. Chem. 75, 724 [1963].

3 F . WEYGAND, A . P R O X , H . H . FESSEL U. K . KUN SUN, Z . N a -turforschg. 20 b, 1169 [1965].

4 E. STENHAGEN, Z. analyt. Chem. 181, 462 [1961].

lung der Cysteinreste mit R a n e y - Nickel vorge-schlagen, wobei die entsprechenden Alaninderivate entstehen. Die direkte massenspektrometrische Unter-scheidung von ursprünglich vorhandenem Alanin ist dann aber schwierig. Es erschien daher günstiger, die SH-Gruppe durch Verätherung zu schützen. Da-durch werden Nebenreaktionen vermieden. Als Schutzgruppe eignet sich die Benzylgruppe, die bei vielen Synthesen ohnehin bereits eingeführt wird und somit zumindest bei der Kontrolle von Synthesen am günstigsten ist. Aber auch bei der massenspektro-metrischen Sequenzanalyse ist diese Schutzgruppe vor bzw. nach der Hydrolyse leicht einzuführen und somit auch in diesen Fällen empfehlenswert.

V-Trifluoracetyl-Peptidester können nach WEY-G A N D gaschromatographisch bis zu Tetrapeptiden6a

5 F . WEYGAND, A . P R O X , E . C . JORGENSEN, R . AXEN U. P . KIRCH-NER, Z. Naturforschg. 18 b, 93 [1963].

8 F. WEYGAND et al., Chem. Ber. 9 2 , 2099 [1959]. ®a A. PROX U. F . WEYGAND, Sequenzanalyse durch Kombina-

tion von Gaschromatographie und Massenspektrometrie; VIII. Europ. Peptidsymposium, Sept. 1966, Noordwijk, North Holland Publ. Amsterdam, im Drude.