Abstract:
Peptide fragments of the alpha thymosin molecule, ranging from four to eighteen peptides in length, and salts thereof, are disclosed. These fragments possess immunostimulatory and immunoregulatory properties similar to the alpha thymosin molecule, and are useful in the same fashion as in this molecule.

Description:
This is a continuation-in-part of application Ser. No. 133,708, filed Mar. 25, 1980, which is now U.S. Pat. No. 4,470,926, which in turn is a continuation-in-part of Ser. No. 490,210, filed May 6, 1983, now abandoned, which in turn is a continuation-in-part of Ser. No. 337,961, filed Jan. 8, 1982. 
    
    
     DESCRIPTION 
     The invention relates to medicaments with immunostimulating or immunoregulating action, which can be used for the treatment of immunity deficiency diseases, of virus infections, treatment for probable tumor formation and especially the treatment of cancer, and also thymosin α 1  fragments which are contained as active substance in this medicament. 
     The influencing of the immune apparatus of the body by polypeptides of the thymus gland has recently been studied with great interest and growing success. This is a consequence of the discovery that cell-free protein extracts of the thymus gland of calves, such as the standard preparation of what is knonw as &#34;thymosin fraction No. 5,&#34; suppress to various extents immune deficiency diseases such as a poor rejection of transplants, an increased sensitivity to infection, and increased probability of the occurrence of tumors. Recently it was reported that the clinical use of thymosin fraction No. 5 on patients suffering from leukemia and other types of cancer has resulted in healing effects, especially in the case of lung cancer (P. B. Chretien et al, J.D. Cancer Treat. Rep. 62 (1978) 1787-1790). 
     In 1977, A. L. Goldstein et al. (J. Proc. Natl. Acad. Sci, USA, 74 (1977) 725) succeeded in separating an acid component in pure form from the thymosin polypeptide mixture, and called it thymosin α 1  ; they also gave its peptide sequence. With 28 amino acids, thymosin α 1  has a molecular It has now been found surprisingly that even fragments of α 1  -thymosin exercise the same immunoregulating or immunostimulating actions as α 1  -thymosin, although to a lesser extent. 
     Despite the somewhat weaker action, the use of α 1  -thymosin fragments instead of α 1  -thymosin in immunotherapy offers advantages: 
     1. With a molecular weight of 3107 and 28 amino acids, α 1  -thymosin is a relatively large polypeptide whose synthesis presents difficulties. The α 1  -thymosin fragments in accordance with the invention, however, are very much smaller and therefore they are substantially easier to prepare, in higher yields and in better purity. 
     2. Among the α 1  -thymosin fragments are found both peptides having an immunostimulating action and peptides having an immunosuppressive action. In the case of the fragments of immunostimulating action, they are mainly C-terminal fragments, while the N-terminal fragments are considered rather to have an immunosuppressive action. On the basis of their different pharmacological properties, these fragments can be used individually or in combination for controlled immunotherapy. 
     Therefore, α 1  -fragments, which are the subject matter of the present invention, can be used for the treatment of immunodeficiency diseases, such as T-cell deficiency states, the elevated probability of tumor formation, and especially cancer. 
     The subject matter of the invention, therefore, is medicaments having an immunostimulating action, which contain as active substance at least one thymosin α 1  fragment and/or at least one derivative thereof in free form or in the form of a pharmacologically acceptable salt. This medicament of the invention can also contain two or more of the thymosin α 1  fragments defined hereinafter, and contains, in addition to the active substances, conventional, pharmacologically acceptable binding agents, carrier materials and/or adjuvants suitable for the selected method of administration. 
     The thymosin α 1  fragments which are contained as active substances in the medicament of the invention, and which are also subject matter of the present invention, are the following: ##STR1## 
     The above-given fragments I to IX in accordance with the invention can bear at their N-terminus an acyl group having 1 to 6 carbon atoms, especially an acetyl group or an acyl glycine group whose acyl residue can have 1 to 6 carbon atoms. At the same time, in the corresponding thymosin α 1  fragments the glutamic acid residues 10, 21, 24, 25 and 27 as well as the aspartic acid residue 15 and the asparagine residue 28 can be present as amide or alkyl amide with 1 to 6 carbon atoms in the alkyl group or as diamide or as dialkyl amide with 1 to 6 carbon atoms in the alkyl groups. 
     The acyl groups with 1 to 6 carbon atoms are especially the acetyl group, the propionyl group and the butyryl group, while the preferred alkyl groups of 1 to 6 carbon atoms are the methyl group, the ethyl group, the propyl group and the various butyl groups. Also, in the corresponding thymosin α 1  fragments position 1 can be varied by a lysine residue instead of serine, and positions 24 and 25 by gamma-carboxy-glutamic acid (Gla) instead of glutamic acid residues. 
     
                       TABLE A______________________________________α.sub.1 -Thymosin Fragments Having an Immunoregulating ActionFragmentNo.______________________________________   Fragments of the formula R.sub.1 --Val--Val--R.sub.110      22-24 Val--Val--Glu11      21-23 Glu--Val--Val12      21-24 Glu--Val--Val--Glu13      19-24 Lys--Lys--Glu--Val--Val--Glu14      18-24 Glu--Lys--Lys--Glu--Val--Val--Glu15      17-24 Lys--Glu--Lys--Lys--Glu--Val--Val--Glu16      13-24 Thr--Lys--Asp--Leu--Lys--Glu--Lys--Lys--   Glu--Val--Val--Glu   R.sub.1 --Lys--Glu--R.sub.217      17-18 Lys--Glu18      20-22 Lys--Glu--Val19      20-23 Lys--Glu--Val--Val20      20-25 Lys--Glu--Val--Val--Glu--Glu21      20-26 Lys--glu--Val--Val--Glu--Glu--Ala22      20-27 Lys--Glu--Val--Val--Glu--Glu--Ala--Glu23      19-22 Lys--Lys--Glu--Val24      18-22 Glu--Lys--Lys--Glu--Val25      17-22 Lys--Glu--Lys--Lys--Glu--Val26      16-22 Leu--Lys--Glu--Lys--Lys--Glu--Val27      16-18 Leu--Lys--Glu28      15- 18 Asp--Leu--Lys--Glu29      14-18 Lys--Asp--Leu--Lys--Glu   R.sub.1 --Glu--Lys--R.sub.230      18-19 Glu--Lys31      17-19 Lys--Glu--Lys32      16-19 Leu--Lys--Glu--Lys33      15-19 Asp--Leu--Lys--Glu--Lys34      14-19 Lys--Asp--Leu--Lys--Glu--Lys35      7-19 Thr--Ser--Ser--Glu--Ile--Thr--Thr--Lys--   Leu--Lys--Glu--Lys   R.sub.1 --Glu--Ala--R.sub.236      25-26 Glu--Ala37      25-27 Glu--Ala--Glu38      24-26 Glu--Glu--Ala39      23-26 Val--Glu--Ala   R--Glu--Ile--Thr40      10-12 Glu--Ile--Thr41      9-12 Ser--Glu--Ile--Thr42      8-12 Ser--Ser--Glu--Ile--Thr   R--Ala--Val--Asp43      4-6 Ala--Val--Asp44      3-6 Ala--Ala--Val--Asp45      2-6 Asp--Ala--Ala--Val--Asp______________________________________ 
    
     Also subject matter of the invention are the pharmaceutically acceptable salts of the above-specified fragments I to IX and 10 to 45. 
     The thymosin α 1  fragments of the invention can be prepared by the application of conventional methods of peptide synthesis, such as conventional synthesis in solution with maximum protection or minimum protection of the side chains, by solid-phase synthesis according to Merrifield wherein the peptide is built up on the polymeric carrier, or by the polymer reagent method wherein the peptide is passed over the polymer-bound activated amino acid to lengthen the chain. 
     In an especially preferred manner, however, the thymosin α 1  fragments of the invention are formed by the process of the Applicant&#39;s U.S. Pat. No. 4,353,821. 
     In this process the thymosin α 1  fragments of the invention are built up by a step-wise sequential lengthening by the mixed anhydride method, wherein especially protective groups are used which have a tertiary carbon atom in the urethane grouping, such as the tertbutyloxycarbonyl group (abbreviated Boc), the α,α-dimethyl-3,5-dimethoxybenzyloxycarbonyl group (Ddz) and the 2-(p-biphenyl)-propyl-2-oxycarbonyl group (Bpoc), which suppress secondary reactions by steric interference, and make it possible to use repeatedly great excesses of the mixed anhydride. 
     The invention will now be explained further with reference to the appended FIG. 2, which is a schematic representation of the thymosin α 1  fragments of the invention with the protective groups used in their preparation. The abbreviations used therein and in the examples correspond to the proposals of the IUPAC-IUP (J. Biol. Chem. 247 (1972) 977-983). These are: 
     Ddz: α,α-dimethyl-3,5-dimethoxybenzyloxycarbonyl, 
     Z: Benzyloxycarbonyl, 
     Ac: Acetyl, 
     OBu t  : tertbutyl ester 
     MbH: 4,4&#39;-dimethyloxybenzohydryl, 
     Me: methyl, 
     Obzl: benzyl ester, and 
     Bu t  : tertbutyl. 
     The following examples will elucidate the preparation of the thymosin α 1  fragments of the invention and their pharmacological effectiveness. 
     The methods of preparation described in the following examples terminate in the protected thymosin α 1  fragments, whose protective groups can be split off in the following manner: 
     SPLITTING OFF THE PROTECTIVE GROUPS 
     An eight-fold to ten-fold excess of anhydrous trifluoroacetic acid in the form of a 5% solution in methylene chloride (absolute) is added to the Ddz-protected peptide ester and stirred for 15 to 30 minutes at room temperature. In the case of the longer peptides the time is to be extended. If the acid-labile 4,4&#39;-dimethoxybenzohydryl group is contained in the sequence, the Ddz-protective group is split off with 2.5% or 1% trifluoroacetic acid for 30 to 60 minutes. To stop the reaction the solution is chilled to -15° C. and brought to a pH of 7.0 to 7.5 with the calculated amount of N-methylmorpholine (+10%). 
     With trifluoroacetic acid in the form of a 50% solution in methylene chloride with the addition of anisole, the tertbutyl ester residues as well as the tertbutyl ether groups are primarily split off, and then, finally, the 4,4&#39;-dimethoxybenzhydryl protective group is dissolved out with 95% trifluoroacetic acid/anisole. If the fragments contain benzyloxycarbonyl protective groups and/or benzyl esters, these are split off before the trifluoroacetic acid treatment by hydrogenation in alcoholic solution in the presence of palladium on charcoal in a conventional manner. 
     The α 1  thymosin fragments of the invention can be transformed in a conventional manner, by the use of preferably pharmacologically acceptable acids or bases, to the corresponding salts. 
     EXAMPLE 1 
     α 1  Thymosin Fragment V 
     Preparation of Ddz-(25-28)-Obzl, Ddz-(28)-OBzl, Ddz-Asn(Mbh)-OBzl 
     3 ml of triethylamine (10% excess) and 3.48 g (=2.43 ml) of benzyl bromide are added to 11.6 g (20 mmol) of Ddz-Asn(MbH)-OH in 10 ml of dimethylformamide at 0° C. The solution is stirred overnight at room temperature and poured into 200 ml of ice water. The suspension is immediately neutralized with NaHCO 3  solution, the solid substance is dissolved with ethyl acetate, and the organic phase is washed with KHSO 4  solution (cold) and NaHCO 3  solution. If free acid is still found in 93:7 [sic], a silica gel column with CHCl 3  /ethanol 94:6 as eluent is connected for the final purification. Yield 8.2 g=61%. For further data see Table I. 
     Ddz-(27-28)-OBzl, Ddz-Glu(OBu t )-Asn(MbH)-OBzl 
     7 mmol of Ddz-Glu(OBu t )-OH is reacted with 10 mmol of Asn(Mbh)-OBzl to form the dipeptide in accordance with the general working procedure for mixed anhydride synthesis. After the conventional shake-out procedure, a chromatography is performed on a silica gel column with CHCl 3  /ethanol 93:7.  The dipeptide is then still contaminated with Ddz cleavage product, which does not, however, interfere with further synthesis operations. For information see Tables I and IX. 
     Ddz-(26-28)-OBzl, Ddz-Ala-Glu(OBu t )-Asn(Mbh)-OBzl 
     15 mmol of Ddz-Ala-OH is reacted as usual with 9.3 mmol of Ddz-(27-28)-OBzl to form the tripeptide, which is purified chromatographically on Sephadex LH20 with methanol and on silica gel with CHCl 3  /ethanol 93:7. Yield 50%. For further data see Tables I and IX. 
     Ddz-(25-28)-OBzl, Ddz-Glu(OBu t )-Ala-Glu(OBu t )-Asn(Mbh)-IBzl (protected α 1  thymosin fragment V) 
     The tetrapeptide is obtained pure by a standard method after chromatography on Sephadex LH20 with methanol. Yield 78%. The peptide is slightly soluble in pure methanol but after the addition of dimethylformamide it is easily soluble. The solubility in CHCl 3  is slight, but is good in a mixture of CHCl 3  and methanol. For further data see Tables I and IX. 
     The splitting off of the protective groups in the manner described above leads to the desired α 1  thymosin fragment V. 
     EXAMPLE 2 
     α 1  Thymosin Fragment IV 
     Preparation of Ddz-(20-24)-OMe 
     (a) Ddz-(24)-OMe, Ddz-Glu(OBu t )-OMe: 20 mmol of Ddz-Glu(OBu t )-OH is dissolved in acetone and the equivalent amount of diazomethane (ether solution) is added. 100% yield of a yellowish oil, chromatographically uniform. For additional data see Table II. 
     (b) Ddz-(23-24)-OMe, Ddz-Val-Glu(OBu t )-OMe: 7.5 mmol of Ddz-Val-OH is reacted with 5 mmol of Ddz-Glu(OBu t )-OMe to the dipeptide. Final purification on silica gel with CHCl 3  /ethanol 93:7. The peptide still contains Ddz cleavage fragments which do not affect further synthesis steps. For information see Tables II and IX. 
     (c) Ddz-(22-24)-OMe, Ddz-Val-Val-Glu(OBu t )-OMe: The tripeptide is obtained pure by a conventional method after chromatography through Sephadex LH 20 with methanol. It is easily soluble in methanol and CHCl 3 . 60% yield of a colorless oil. For further data see Tables II and IX. 
     (d) Ddz-(21-24)-OMe, Ddz-Glu(OBu t )-Val-Val-Glu(OBu t )-OMe: 6 mmol of Ddz-Glu(OBu t )-OH is reacted to the tetrapeptide as usual with 2.98 mmol of Ddz-(22-24)-OMe. It is placed on a Sephadex LH 20/methanol column to obtain the product as a colorless glass in a 94% yield. The tetrapeptide dissolves easily in CHCl 3 , methanol, dimethylformamide and dioxane. It is uniform in thin-layer chromatography in 93:7. 7:1 and 85:10:5, and it has the usual violet color with ninhydrin. For further data see Tables II and IX. 
     (e) Ddz-(20-24)-OMe, Ddz-Lys(Z)-Glu(OBu t )-Val-Val-Glu(OBu t )-OMe (protected fragment IV): 9 mmol of Ddz-Lys(Z)-OH is reacted to the pentapeptide as usual with 2.43 mmol of Ddz-(21-24)-OMe. After the shake-out procedure it is purified chromatographically on a methanol/Sephadex LH 20 column, to obtain the product as a colorless glass in a 99% yield. For further information see Tables II and IX. 
     By conventional splitting off of the protective groups the desired α 1  thymosin fragment IV is obtained. 
     EXAMPLE 3 
     α 1  Thymosin Fragment III 
     Preparation of Ddz-(13-19)-OMe 
     (a) Ddz-(19)-OMe, Ddz-Lys(Z)-OMe: 20 mmol of Ddz-Lys(Z)-OH is dissolved in tetrahydrofuran and the equivalent amount of ethereal diazomethane solution is added. The amino acid residue is obtained in a 95% yield as a yellowish oil after shaking with NaHCO 3  solution. Crystallization from the common solvents was unsuccessful. For further data see Table III. 
     (b) Ddz-(18-19)-OMe, Ddz-Glu(OBu t )-Lys(Z)-OMe: The dipeptide is prepared by conventional method. The peptide is obtained as a yellowish oil contaminated with small amounts of Ddz cleavage fragments. For data see Tables III and X. 
     (c) Ddz-(17-19)-OMe, Ddz-Lys(Z)-Glu(OBu t )-Lys(Z)-OMe: The synthesis of the tripeptide is accomplished in a normal manner by the conventional method. 20 mmol of Ddz-Lys(Z)-OH is reacted with 10 mmol of Ddz-(18-19)-OMe. After the acid and alkaline extraction, the chromatography is performed first on silica gel with CHCl 3  /ethanol 97:3, then on Sephadex LH 20 with methanol. Yield: 86% of a colorless, deliquescent foam. For further data see Tables III and X. 
     (d) Ddz-(16-19)-OMe, Ddz-Leu-Lys(Z)-Glu(OBu t )-Lys(Z)-OMe: The tetrapeptide is prepared by a conventional method. After chromatography on LH 20/methanol, 4.1 g of a white, amorphous solid can be isolated in a 77% yield. The peptide is easily soluble in methanol and CHCl 3 . For further data see Tables III and X. 
     (e) Ddz-(15-19)-OMe, Ddz-Asp(OBu t )-Leu-Lys(Z)-Glu(OBu t )-Lys(Z)-OMe: It is necessary to modify the general working procedure for the synthesis of the pentapeptide. The mixed anhydride synthesis is performed in the usual stoichometry (9.5 mmol of Ddz-Asp(OBu t )-OH and 3.7 mmol of tetrapeptide) in a three-necked flask with a powerful stirrer. After the carboxyl component and the amine component are combined, the methylene chloride solution gels spontaneously at -15° C. Liquefaction can be achieved by the addition of 40 to 60 ml of dimethylformamide. After 4 to 6 hours of reaction time, another 100 to 200 ml of dimethylformamide is added and the methyl chloride is cautiously removed by evaporation. The remaining dimethylformamide solution is slowly stirred into five times the volume of ice cold NaCl solution, and the white, flocculent precipitate is removed by suction filtering. After washing with water, repeating the precipitation and drying, the peptide is obtained in an 84% yield. Ddz cleavage residues can be washed out with ether. 
     The peptide is very slightly soluble in methanol, CH 2  Cl 2 , CHCl 3  and dioxane. A suspension can be prepared in 1,1-dichloroethane. The peptide is soluble without problems in 5% trifluoroacetic acid/CH 2  Cl 2 . For further data see Tables III and X. 
     (f) Ddz-(14-19)-OMe, Ddz-Lys(Z)-Asp(OBu 6 )-Leu-Lys(Z)-Glu(OBu t )-Lys(Z)-OMe: 3.5 mmol of Ddz-Lys(Z)-OH and 1.74 mmol of Ddz-(15-19)-OMe can be reacted to the hexapeptide without further solubility problems. After chromatography on Sephadex LH 20/methanol, a glassy solid is obtained in a yield of about 100%. The product is easily soluble in methanol, especially after the addition of di-methylformamide. For further data see Tables III and X. 
     (g) Ddz-(13-19)-OMe, Ddz-Thr(Bu t )-Lys(Z)-Asp(OBu t )-Leu-Lys(Z)-Glu(OBu t )-Lys(Z)-OMe (protected fragment III): 11.4 mmol of Ddz-Thr(BU t )-OH is reacted with 5 mmol of Ddz-(14-19)-OMe, being careful that the heptapeptide remains dissolved for the acid/alkaline extraction in CH 2  Cl 2 , that is, that the usual change of solvent is eliminated. The peptide is only slightly soluble in ethyl acetate, methanol and dimethylformamide, but easily soluble in CH 2  Cl 2  and CHCl 3 . Chromatographic purification is performed on silica gel and CHCl 3  /ethanol 98:2 as eluent, after which the heptapeptide is obtained in a 56% yield. It strikingly turns yellowish red with ninhydrin. For further data see Tables III and X. 
     (h) By splitting off the protective groups the desired α 1  thymosin fragment III is obtained. 
     EXAMPLE 4 
     α 1  Thymosin Fragment II 
     Preparation of Ddz-(7-12)-OBzl 
     (a) Ddz-(12)-OBzl, Ddz-Thr(Bu t )-OBzl: 20 mmol of Ddz-Thr(Bu t )-OH are esterified with benzyl bromide as specified for Ddz-Asn(Mbh)-OBzl. By extraction with KHSO 4  solution and KHCO 3  solution a chromatographically uniform oil can be obtained in a 73% yield. For further data see Table IV. 
     (b) Ddz-(11-12)-OBzl, Ddz-Ile-Thr(Bu t )-OBzl: After splitting off Ddz from 10 mmol of Ddz-Thr(Bu t )-OBzl, the dipeptide is prepared by a conventional method from 15 mmol of Ddz-Ile-OH. This is chromatographed on silica gel with chloroform/ethanol 98:2,  and crystallized from CCl 4  /n-heptane. 5.1 g of fine, white needles (=8.47 mmol, 85%) was thus obtained. For further data see Table IV. 
     (c) Ddz-(10-12)-OBzl, Ddz-Glu(OBu t )-Ile-Thr(Bu t )-OBzl: 16 mmol of Ddz-Glu(OBu t )-OH is reacted with 8 mmol of Ddz-(11-12)-OBzl in the usual manner to form the tripeptide. After acid and alkaline extraction, chromatography is performed with methanol on LH 20. Yield 89%. For further data see Table IV. 
     (d) Ddz-(9-12)-OBzl, Ddz-Ser(Bu t )-Glu(OBu t )-Ile-Thr(Bu t )-OBzl: The tripeptide is prepared conventionally from 7 mmol of Ddz-(10-12)-OBzl with 14 mmol of Ddz-Ser(Bu t )-OH. After acid and alkaline extraction, purification is performed on a silica gel column (CHCl 3  /ethanol) 98:2). 5.3 g of a white amorphous substance is obtained (5.8 mmol, 83% having a melting range from 172° to 177° C. For further data see Table IV. 
     (e) Ddz-(8-12)-OBzl, Ddz-Ser(Bu t )-Ser(Bu t )-Glu(OBu t )-Ile-Thr(Bu t )-OBzl: The pentapeptide is obtained conventionally from 11.6 mmol of Ddz-Ser(Bu t )-OH and 5.8 mmol of Ddz-(19-12)-OBzl. After acid and alkaline extraction, the product is dissolved in methanol/dimethylformamide 8:2 and placed on a Sephadex LH 20/methanol column. The peptide, which is poorly soluble in methanol, spontaneously crystallizes in the eluate. 4.7 g of white needles are obtained, which is a 78% yield. For further data see Tables IV and X. 
     (f) Ddz-(7-12)-OBzl, Ddz-Thr(Bu t )-Ser(Bu t )-Ser(Bu t )-Glu(OBu t )-Ile-Thr(Bu.sup.t)-OBzl (protected fragment II) The hexapeptide is obtained conventionally from 6.2 mmol of Ddz-Thr(Bu t )-OH and 3.5 mmol of Ddz-(8-12)-OBzl. After acid and alkaline extraction, the peptide, which is poorly soluble in methanol, is dissolved in dimethylformamide/methanol and placed on an LH 20 column and eluted with methanol. 4.0 g of a glassy, amorphous substance, which is turned yellowish red by ninhydrin, yield=97%. The peptide is soluble in benzene, CH 2  Cl 2 , CHCl 3  and dimethylformamide. For further data see Tables IV and X. 
     (g) In like manner one obtains Ddz-(7-12)-OMe, Ddz-Thr(Bu t )-Ser(Bu t )-Ser(Bu t )-Glu(OBu t )-Ile-Thr(Bu.sup.t)-OMe (protected fragment II): The synthesis is performed in a manner entirely analogous to the preparation of the Ddz-(7-12) benzyl ester. All of the properties of the intermediate products can be seen in Table V and Table I, respectively. 
     (h) By splitting off the protective groups the desired fragment II of α 1  thymosin is obtained. 
     EXAMPLE 5 
     α 1  Thymosin Fragment I 
     Preparation of Ac-(1-6)-OBzl) 
     (a) Ddz-(6)-OBzl, Ddz-Asp(OBu t )-OBzl: 20 mmol of Ddz-Asp(OBu t )-OH is esterified with benzyl bromide in a manner similar to that specified for Ddz-(28)-OBzl. By extraction with KHSO 4  solution and KHCO 3  solution, a chromatographically uniform oil can be obtained in an 81% yield. For data see Table VI. 
     (b) Ac-(1)-OH, Ac-Ser(Bu t )-OH: Ddz is split off with 5% trifluoroacetic acid from 20 mmol of Ddz-Ser(Bu t )-OH, and the solvent is withdrawn in vacuo. The product is dissolved at 0° C. with 20 ml of 1N NaOH and transferred to a three-necked flask. 200 mmol (=15.6 g) of acetyl chloride, diluted 1:1 with dioxane, is added drop by drop on the pH-stat at pH 9, while maintaining the temperature at 0° to 4° C. Thirty minutes after the addition has been completed, about 50 ml of a dioxane/H 2  O azeotrope is withdrawn in vacuo. The alkaline solution is extracted with ether, adjusted to pH 2 to 3 with KHSO 4  and washed again with ether. Finally the product is extracted from the aqueous phase with ethyl acetate. 3.6 g=96% is obtained of an amorphous substance which crystallizes in granular, colorless form from ethyl acetate in the refrigerator. For further data see Table VI. 
     (c) Ddz-(5-6)-OBzl, Ddz-Val-Asp(OBu t )-OBzl: 15 mmol of Ddz-Val-OH is reacted in a conventional manner with 11.5 mmol of Ddz-Asp(OBu t )-OBzl. The dipeptide is chromatographed on silica gel with CHCl 3  /ethanol 99:1. For data see Tables VI and X. 
     (d) Ddz-(4-6)-OBzl, Ddz-Ala-Val-Asp(OBu 6 )-OBzl: 20 mmol of Ddz-Ala-OH is reacted with Ddz-(5-6)-OBzl to the peptide without difficulty. Chromatographic purification on Sephadex LH 20/methanol. For data see Tables VI and X. 
     (e) Ddz-(3-6)-OBzl, Ddz-Ala-Ala-Val-Asp(OBu t )-OBzl: The tetrapeptide is prepared in a conventional manner from 17 mmol of Ddz-Ala-OH and 8.8 mmol of Ddz-(4-6)-OBzl. In the acid and alkaline extraction the peptide flocculates out of ethyl acetate, but can be kept in suspension. Gel chromatography (LH 20) does not show a chromatographically uniform product. Therefore another chromatography is performed on silica gel in CHCl 3  /ethanol 99:1. 4.5 g of a colorless, amorphous product is obtained in a 70% yield. For further data see Tables VI and X. 
     (f) Ddz-(2-6)-OBzl, Ddz-Asp(OBu t )-Ala-Ala-Val-Asp(OBu t )-OBzl. In the conventional reaction of 11.6 mmol of Ddz-Asp-(OBu t )-OH with 5.8 mmol of Ddz-(3-6)-OBzl, the poor solubility of the amine component in CH 2  Cl 2  is striking. Peptide that has precipitated like a gel can be dissolved in dimethyl formamide after the Ddz removal and during the coupling. The crude pentapeptide is dissolved in methanol/dimethylformamide and chromatographed on LH 20, crystallizing from the fractions of the chromatography in fine white needles. Yield 85%. For data see Tables VI and X. 
     (g) Ac-(1∝5)-OBzl, Ac-Ser(Bu t )-Asp(OBu t )-Ala-Ala-Val-Asp-(OBu t )-OBzl (protected fragment I): 2.8 mmol of Ac-Ser(Bu t )-OH is reacted as usual with 1.4 mmol of Ddz-(2-6)-OBzl. After the reaction has ended and the CH 2  Cl 2  has been withdrawn, the product is dissolved in 150 ml of dimethyl formamide and stirred into a 5% solution of NaHCO 3  saturated with NaCl. The precipitated product is suction filtered, washed with water and dried. It is dissolved in a little CHCl 3  and applied to a MERCK ready-prepared column. The column is eluted with alcohol-free CHCl 3  and, after the run, the front is developed with a linear gradient from CHCl 3  mixed with 0 to 10% of ethanol. 950 mg of a white, amorphous peptide is obtained. It is not easily soluble in CHCl 3 , cannot be stained with ninhydrin, but reacts with &#34;Chlor I/II.&#34; For data see Tables VI and X. 
     (h) Preparation of Ddz-(1-6)-OMe: The hexapeptide is prepared in a manner similar to that used in preparing Ac-(1-6)-OBzl. 
     Ddz-Ser(Bu t )-Asp(OBu t )-Ala-Ala-Val-Asp(OBu t )OMe. The hexapeptide dissolves well in methanol/dimethylformamide. It can be chromatographed easily on Sephadex LH 20. Its solubility in CHCl 3 , tetrahydrofuran and dioxane is moderately good, but definitely better than that of the acetyl peptide. 
     The fragment Ddz-(1-6)-OBzl described hereinbelow has the same properties also. 
     (i) Preparation of Ddz-(1-6)-OBzl: The hexapeptide is prepared in a manner similar to the method used in preparing Ac-(1-6)-OBzl, using Ddz-Ser(Bu t ) instead of Ac-Ser(Bu t ). 
     (j) Preparation of Ddz-Lys(Z)-(2-6)-OBzl; (protected fragment Ia): 7 mmol of Ddz-Lys(Z)OH and 3.1 mmol of Ddz-(2-6)-OBzl are reacted to the hexapeptide as usual. After chromatography on Sephadex LH 20/methanol, the uniform end product is obtained. For data see Tables VI and XI. 
     (k) By splitting off the protective groups the desired α 1  -thymosin fragment I or Ia, Lys-(2-6)-OH, is obtained from the products designated under (i) and (j). 
     EXAMPLE 6 
     Thymosin α 1  Fragment VII 
     Synthesis of the fragment Ddz-(13-28)-OBzl 
     (a) Saponification of Ddz-(20-24)-OMe, Ddz-Lys(Z)-Glu(OBu t )-Val-Val-Glu(OBu t )-OMe: 1 mmol (1.1 g) of Ddz-(20-24)-OMe is dissolved in 20 ml of aqueous 80% dioxane (concentration 50 mmol). An ultrasonic bath is helpful for this purpose. Then 2 ml (2 mmol) of 1N NaOH is added and the mixture is stirred for one hour. The reaction time is determined by preliminary experiment. After stopping the saponification with 1 mmol of acetic acid, a part of the dioxane is withdrawn with a rotary evaporator and the residue is separated on a Sephadex LH 20/methanol column into a peptide peak and a salt peak. The peptide peak contains the chromatographically pure peptide acid in a 76% yield. For further data see Tables VII and XI. 
     (b) Fragment Condensation to Ddz-(20-28)-OBzl (protected fragment VI): The protective group is split off from 895 mg (=0.86 mmol, 15% excess) of Ddz-(25-28)-OBzl in a 45-minute reaction with an 8-fold excess of 2.5% trifluoroacetic acid in CH 2  Cl 2 . The stopping of the reaction and the neutralization (pH 7.5) is performed with 1.0 ml of N-methylmorpholine. The solvent is concentrated in vacuo. 
     800 mg (0.75 mmol) of Ddz-(20-24)-OH is dissolved in 15.2 ml of dimethylformamide (final concentration 50 mM); the amine component is added and the mixture is cooled to 0° C. Then 1.52 mmol (=313 mg, 2 equivalents) of dicyclohexylcarbodiimide and 349 mg (2.28 mmol, 3 equivalents) of 1-hydroxybenzotriazole. H 2  O is added. The clear solution is stirred for one hour at 0° C., and then warmed slowly to room temperature. After another hour, dicyclohexylurea suddenly precipitates in microcrystalline threads. After a total of 24 hours of reaction time, the mixture is chilled to 0° C., centrifuged free of dicyclohexylurea, and precipitated in 50 ml of 5% NaHCO 3  solution; the precipitate is washed with ice water and dried. 
     The crude substance (0.7 mmol=93%) is dissolved in CHCl 3  and placed on a MERCK prepared column K 60, Size C, which is balanced with CHCl 3 . A low gradient of CHCl 3  is applied against CHCl 3  /ethanol 90:10. 1.1 g (=70%) of pure nonapeptide is obtained. For further data see Tables VIII and XI. 
     (c) Saponification of Ddz-(13-19)-OMe, Ddz-Thr(Bu t )-Lys(Z)-Asp(OBu t )-Leu-Lys(Z)-Glu(OBu t )-Lys(Z)-OMe: 0.7 mmol of Ddz-(13-19)-OMe (=1.2 g) is dissolved with 9 ml of tetrahydrofuran and 4 ml of methanol under the action of ultrasound; then 2 ml of water is added (final concentration 47 mM). After the addition of 1.5 ml of 1N NaOH (1.4 mmol), the saponification is performed for 25 minutes. The reaction time is determined by preliminary experiment. The reaction is stopped with 0.7 mmol of acetic acid. After concentration, chromatography is performed on Sephadex LH 20/methanol. For further data see Tables VII and XI. 
     (d) Fragment condensation to Ddz-(13-28)-OBzl (protected fragment VII): The Ddz protective group is split off from 583 mg (=0.3 mmol) of Ddz-(20-28)-OBzl in a 45-minute reaction with a 10-fold excess of 2.5% trifluoroacetic acid in CH 2  Cl 2 . The stopping of the reaction and the neutralization are performed with 0.33 ml of N-methylmorpholine (pH 7.5). The solvent is withdrawn in vacuo. 
     663 mg (=0.4 mmol, 1.33 equivalents) of Ddz-(13-19)-OH is dissolved in 12 ml of dimethyl formamide (final concentration 25 mM); the amine component is added, and the mixture is chilled to 0° C. Then 165 mg (0.8 mmol) of dicyclohexylcarbodiimide and 184 mg (=1.2 mmol) of 1-hydroxybenzotriazole. H 2  O is added. The clear solution, again adjusted to pH 7.5 with N-methylmorpholine, is stirred for one hour at 0° C., then slowly warmed up to room temperature. After another 1.5 to 2 hours, dicyclohexylurea suddenly precipitates in microcrystalline threads. After a total of 66 hours (24 hours in a second batch), the temperature is lowered to 0° C., the dicyclohexylurea is centrifuged out, and chromatography is performed on a large LH 20/methanol column. From the first peptide-containing fractions a white, flocculent material precipitates, which is identified as hexadecapeptide. The second and third peaks of the separation contain unreacted carboxyl component or amine component. 
     The hexadecapeptide is obtained as a white powder which is slightly soluble in methanol, but easily soluble after the addition of dimethylformamide. Incipient decomposition appears at 230° C., which is complete at 260° C. For further data see Tables VIII and XI. 
     (e) By conventional splitting off of the protective groups, the desired fragment VII of thymosin α 1  is obtained. 
     EXAMPLE 7 
     α 1  Thymosin Fragment IX 
     (a) Hydrogenation of Ac-(1-5)-0Bzl, Ac-Ser(Bu t )-Asp(OBu t )-Ala-Ala-Val-Asp(OBu t )-OBzl: 500 mg of Ac-(1-6)-OBzl is steeped in 100 ml of isopropanol/10% acetic acid, and dissolved with 600 ml of isopropanol at 50° C. in the vibratory mixer. The hydrogenation is performed at room temperature in the vibratory mixer with 1 g of Pd/C in a stream of H 2 . After 9 hours the catalyst is removed by a suction filter and the solvent is withdrawn in vacuo with the addition of toluene. The crude product in methanol with some dimethylformamide added is placed on an LH 20 column and chromatographed. Yield 300 mg=67%. 
     The hydrogenation can also be performed in 50 ml of 2,2,2-trifluoroethanol with 0.5 g of Pd/C, and the purification can also be performed on LH 20/2,2,2-trifluoroethanol. For analysis data see Tables VII and XI. 
     (b) Fragment Condensation to Ac-(1-12)-OBzl (protected fragment IX): 1.54 g (=1.3 mmol) of Ddz-(7-12)-OBzl is dissolved in 15 ml of CH 2  Cl 2  and 0.77 ml of trifluoroacetic acid (8-fold excess, 5% solution) to split off the Ddz protective groups. After 15 minutes the mixture is neutralized (pH 7.5) with 1.4 ml of N-methylmorpholine (1.2 equivalents). 900 mg of Ac-(1-6)-OH (=1.17 mmol) is dissolved together with the amine component in 35 ml of dimethylformamide and 11 ml of N-methylmorpholine, and the mixture is cooled to 0° C. 2.34 mmol (=2 equivalents) of dicyclohexylcarbodiimide and 3.51 mmol (=3 equivalents) of 1-hydroxybenzotriazole. H 2  O are added and stirred for one hour at 0° C. During the 60 hours of reaction time at room temperature, an increasing turbidity of the solution is observed, due to dodecapeptide precipitating in the manner of a gel. The solution is centrifuged, and the precipitate is washed twice with dimethylformamide and twice with water, and dried. 1.4 g of a white powder was obtained. This corresponds to 0.72 mmol, i.e., a yield of 61% of the theory. 
     The product is largely insoluble: CH 2  Cl 2 , CHCl 3 , dioxane, tetrahydrofuran, methanol, dimethylformamide, N-methylpyrrolidone, hexamethylphosphoric acid triamide, tertbutanol, isoamyl alcohol, isopropanol, cyclohexane, cyclohexanone, and the addition of Ac-OH, ether, benzine and other solvents, all fail to dissolve the peptide. It is soluble only in 2,2,2-trifluoroethanol. For further data see Tables VIII and XI. 
     (c) Hydrogenation of Ac-(1-12)-OBzl: 566 mg of Ac-(1-12)-OBzl is hydrogenated overnight with 100 mg of Pd/C in 60 ml of distilled trifluoroethanol. 
     Purification is performed on a MERCK prepared silica gel column with a gradient of CHCl 3  /2,2,2-trifluoroethanol/acetic acid 85:10:5 after 2,2,2-trifluoroethanol/acetic acid 90:10. The substance elutes in a low peak towards the end of the gradient. For data see Tables VII and XI. 
     (d) By splitting off the protective groups the desired fragment IX of thymosin α 1  is obtained. 
     EXAMPLE 8 
     α 1  Thymosin Fragment VIII 
     (a) Saponification of Ddz-(7-12)-OMe, Ddz-Thr(Bu t )-Ser-(Bu t )-Ser(Bu t )-Glu(OBu t )-Ile-Thr(Bu.sup.t)-OMe: 1.0 g (0.95 mmol) of Ddz-(7-12)-OMe is dissolved in 19 ml of dioxane and 2.5 ml of H 2  O is added. If the substance precipitates, ultrasound treatment can be used to obtain an emulsion suitable for the saponification. The reaction is started with 0.5 ml of 1N NaOH. After 20 and 40 minutes, an additional 0.5 ml and 0.9 ml, respectively, of 1N NaOH is added. The reaction is stopped after one hour with 0.95 mmol of acetic acid (pH 7.5). 
     The solvent is withdrawn in vacuo, and the residue is dissolved in methanol and chromatographed on LH 20/methanol, unreacted peptide ester being eluted ahead of the main peak of the peptide acid. The substance (57% yield) is easily soluble in methanol, CHCl 3  and toluene/methanol. For additional data see Tables VII and XI. 
     (b) Condensation of Fragment to Ddz-(7-28)-OBzl (protected fragment VIII): In a typical batch, the Ddz protective group is split off from 120 mg of Ddz-(13-28)-OBzl (36 μmol) in a one-hour reaction with a 20-fold excess of 2.5% (or 1%) trifluoroacetic acid in CH 2  Cl 2 . The stopping of the reaction and the neutralization (pH 7.5) are performed with N-methylmorpholine (microliter syringe). The solvent is withdrawn in vacuo. 
     77 mg (72 μmol, 2 equivalents) of Ddz-(7-12)-OH is dissolved in 1.5 ml of dimethylformamide, the amine component is added, and the mixture is cooled to 0° C. Then 16 mg (78.6 μmol, 2.2 equivalents) of dicyclohexylcarbodiimide and 24 mg (157 μmol, 4.4 equivalents) of 1-hydroxybenzotriazole. H 2  O is added and the pH is adjusted to 7.5 with N-methylmorpholine (theoretically 6.4 equivalents. The clear solution is stirred overnight at 0° C. Precipitation of the dicyclohexylurea shows the progress of the reaction. The solution is kept for 10 more hours at about 20° C., chilled again to 0° C., and the dicyclohexylurea is removed by centrifugation. The supernatant fluid is dripped into 30 ml of ice-cold methanol, whereupon the crude 22-peptide precipitates. It is vacuum dried, dissolved in 2,2,2-trifluoroethanol, and chromatographically purified on a 2,2,2-trifluoroethanol/LH-20 column (0.6×240 cm) by re-chromatographing the first peak until uniformity is achieved. 
     By splitting off the protective groups in the conventional manner, the desired fragment VIII of thymosin α 1  is obtained. 
     The 22-peptide is obtained as a glassy solid, which is insoluble in methanol, soluble in dimethylformamide, and easily soluble in 2,2,2-trifluoroethanol. The yield amounts to 67%. For further data see Tables VIII and XI. 
     
                                           TABLE I__________________________________________________________________________Thymosin α.sub.1 -Fragment V                      hRf α.sup.20    Yield        Appearance                 MP (°C.)                      I II                          578 nm                              365 nm                                  (g/100 ml)                                        Solvent__________________________________________________________________________Ddz-(28)-OBzl    61% white amorphous                 136-138                      49                        92                           -4,4°                               -12,3°                                  1,145 MethanolDdz-(27-28)-OBzl    100%        yellow crystalline                 125-132                      35                        92                           -4,8°                              -15,6°                                  1,465 MethanolDdz-(26-28)-OBzl    80% white foamy       -23,8°                              -80,3°                                  1,333 MethanolDdz-(25-28)-OBzl    78% white amorphous                 154-156                      24                        90                          -14,5°                              -51,1°                                  0,440 Methanol/                                        Dimethyl-                                        formamide                                        (8:2 v/v)__________________________________________________________________________ I: 7:1 II: 85:10:5 
    
     
                                           TABLE II__________________________________________________________________________Thymosin α.sub.1 -Fragment IV                hRf α.sup.20    Yield        Appearance                I II                    578 nm                        365 nm                            c (g/100 ml)                                  Solvent__________________________________________________________________________Ddz-(24)-OMe    100%        colorless oil                57                  92Ddz-(23-24)-OMe    100%        oil     47                  92Ddz-(22-24)-OMe    60% colorless oil                28                  92Ddz-(21-24)-OMe    94% colorless foamy                22                  88Ddz-(20-24)-OMe    99% colorless glass                05                  92                    -44,5°                        -155,9°                            0,53  Methanol__________________________________________________________________________ I: 7:1 II: 85:10:5 
    
     
                                           TABLE III__________________________________________________________________________Thymosin α.sub.1 -Fragment III                       hRf     α.sup.20    Yield %         Appearance                  MP (°C.)                       I II                           III                             IV                               578 nm                                   365 nm                                       c (g/100 ml)                                             Solvent__________________________________________________________________________Ddz-(19)-OMe    95   yellowish oil                  --   45  92  -6,2°                                   -16,4°                                       1,000 MethanolDdz-(18-19)-OMe    100  yellowish oil                  --   31  90  -12,8°                                       1,000 MethanolDdz-(17-19)-OMe    86   deliquescent foam                  --   15  86                             15                               -18,5°                                   -61,3°                                       1,539 MethanolDdz-(16-19)-OMe    77   white foamy                  --   21  84  -29,7°                                   -100,0°                                       0,740 MethanolDdz-(15-19)-OMe    84   white amorphous                  184-185°                       16  85  -21,8°                                   -70,0°                                       0,370 Dimethyl-                  (G.)                       formamidDdz-(14-19)-OMe    100  glass-like,                  --   01  92  -23,4°                                   -81,9°                                       0,475 Methanol         colorlessDdz-(13-19)-OMe    56   glass-like,                  --   00                         55                           88  -24,5°                                   -87,2°                                       0,375 Methanol         colorless__________________________________________________________________________ I: 7:1 II: 93:7 III: 85:10:5 IV: 4:1:1 
    
     
                                           TABLE IV__________________________________________________________________________Thymosin α.sub.1 -Fragment II                        hRf   α.sup.20    Yield        Appearance                 MP (°C.)                        I II                            III                              578 nm                                  365 nm                                      c (g/100 ml)                                            Solvent__________________________________________________________________________Ddz-(12)-OBzl    73% colorless oil                 --     58  59                              -17,5°                                  -53,5°                                      2,017 MethanolDdz-(11-12)-OBzl    85% fine needles                 116,5-117,5°                        57                          83  -20,6°                                  -69,1°                                      0,33  MethanolDdz-(10-12)-OBzl    89% colorless foam                 --     38                          92Ddz-(9-12)-OBzl    83% white amorphous                 172-177°                        34                          93   -4,3°                                  -19,8°                                      0,515 CH.sub.2 Cl.sub.2Ddz-(8-12)-OBzl    78% white needles                 191-192°                        28                          95   -2,2°                                  -16,5°                                      0,87  CH.sub.2 Cl.sub.2Ddz-(7-12)-OBzl    97% glassy amorphous                        17                          92                            58                               -5,1°                                   -6,9°                                      0,505 CH.sub.2 Cl.sub.2__________________________________________________________________________ I: 7:1 II: 85:10:5 III: 93:7 
    
     
                                           TABLE V__________________________________________________________________________Thymosin α.sub.1 -Fragment II        hRf α.sup.20    Yield        I II            578 nm                365 nm                    c (g/100 ml)                          Solvent__________________________________________________________________________Ddz-(11-12)-OMe    68,0%        53          92            -22,6°                -73,1°                    2     MethanolDdz-(10-12)-OMe    77,9%        37          81            -15,9°                -52,3°                    2     MethanolDdz-(9-12)-OMe    62,3%        22          73             -5,9°                -24,1°                    1     MethanolDdz-(8-12)-OMe    84,2%        11          72             -1,0°                -11,8°                    1     MethanolDdz-(7-12)-OMe    71,4%        12          78             -1,6°                -11,0°                    1     Methanol__________________________________________________________________________ I: 7:1 II: 93:7 
    
     
                                           TABLE VI__________________________________________________________________________Thymosin α.sub.1 -Fragment I                hRf       α.sup.20   Yield       Appearance                I II                    III                      IV                        V 578 nm                              365 nm                                   c (g/100 ml)                                         Solvent__________________________________________________________________________Ddz-(6)-OBzl   81% colorless oil                48  40    -19,8°                               -70,7°                                   1,388 MethanolAc-(1)-OH   96% granular crystals                07    71                        38                          +39,2°                              +133,3°                                   0,354 Ethyl acetateDdz-(5-6)-OBzl   100%       yellowish oil                58                  92       -6,5°                               -23,7°                                   8,44  MethanolDdz-(4-6)-OBzl   76% foam     27                  92      -47,4°                              -171,2°                                   0,515 MethanolDdz-(3-6)-OBzl   70% colorless amorph                24                  78      -54,6°                              -193,4°                                   0,71  MethanolDdz-(2-6)-OBzl   85% needles  05                  92      -24,6°                               -97,7°                                   0,435 CH.sub.2 Cl.sub.2Ac-(1-6)-OBzl   78% white foamy                00                  63                    35    -34,0°                              -145,5°                                   0,2   Dimethylsulfox-                                         ide/MethanolDdz-(1-6)-OBzl   84% colorless amorpho                04                  90                    47    -31,0°                              -142,0°                                   0,5   CH.sub.2 Cl.sub.2Ddz-Lys(Z)--   100%       colorless amorpho                16                  79      --  --   --    --(2-6)-OBzl__________________________________________________________________________ I: 7:1 II: 85:10:5 III: 93:7 IV: 4:1:1 V: sec. butanol/3% ammonia solution (100:44) 
    
     
                                           TABLE VII__________________________________________________________________________Thymosin α.sub.1 -FragmentsI-IV and IX in the form of the peptide acidsFrag-                           hRf    α.sup.20                                           c (g/100ment         Yield            Appearance                    MP (°C.)                           I  II                                III                                   578 nm                                       365 nm                                             ml)  Solvent__________________________________________________________________________IV  Ddz-(20-24)-OH        76% glass-like                    --        0 57III Ddz-(13-19)-OH        86% white powder                    180-182 (Z)                           80.sup.(1)                              0 56 -24,5°                                       -87,2°                                            0,375 MethanolII  Ddz-(7-12)-OH        57% white powder                    --        0 74 -18,0°                                       -46,7°                                            0,15  MethanolI   Ac-(1-6)-OH        67% white amorph.                    219-221° (Z)                              0 22 -47,1°                                       -190,0°                                            0,014 2,2,2-trifluoro-IX  Ac-(1-12)-OH        85% colorless glass                    --        0 25.sup.(2)                                   -33,3°                                       -98,3°                                            0,06  ethanol__________________________________________________________________________ I: 7:1 II: 93:7 III: 85:10:5 .sup.(1) 4:1:1 .sup.(2) CHCl.sub.3 :2,2,2trifluoroethanol/acetic acid 
    
     
                                           TABLE VIII__________________________________________________________________________Thymosin α.sub.1 -Fragments VI-IXFrag-                         hRf    α.sup.20ment          Yield             Appearance                    MP (°C.)                         I II III                                578 nm                                    365 nm                                         c (g/100 ml)                                               Solvent__________________________________________________________________________VI   Ddz-(20-28)-OBzl         70% glass-like                    252° (Z)                         25                           72   +13,1°                                    +29,4°                                         0,435 CHCl.sub.3VII  Ddz-(13-28)-OBzl         60% white powder                    260° (Z)                         00                           60 88                                 -8,1°                                    -49,8°                                         0,235 CHCl.sub.3 Methanol                                               (8:2)VIII Ddz-(7-28)-OBzl         67% glass-like                    --     50    -9,7°                                    -25,2°.sup.(1)                                         2,593 2,2,2-trifuoroethanol                                               1IX   Ac-(1-12)-OBzl         61% white powder                    263° (Z)                         34                           77   -28,7°                                    -116,2°                                         0,16  2,2,2-trifluoroethanol__________________________________________________________________________ I: 93:7 II: 85:10:5 III: 4:1:1 .sup.(1) 436 nm 
    
     
                                           TABLE IX__________________________________________________________________________Amino acid analysis of thymosin α.sub.1 -Fragments IV and VFrag-        Hydrolysisment         Method              Asp Glu Ala Val Lys__________________________________________________________________________V   Ddz-(25-28)-OBzl        a     1,00 (1)                  1,90 (2)                      1,20 (1)IV  Ddz-(20-24)-OMe        a         1,82 (2)                          2,00 (2)                              1,05 (1)__________________________________________________________________________ Hydrolysis method: a = HCl/propionic acid 160° C. 45 to 60 minutes 
    
     
                                           TABLE X__________________________________________________________________________Amino acid analysis of Thymosin α.sub.1 -Fragments I to IIIFrag-        Hydrolysisment         Method              Asp Thr Ser Glu Ala Val Ile Leu Lys__________________________________________________________________________III Ddz-(13-19)-OMe        a     0,96 (1)                  0,6 (1) 1,05 (1)        1,00 (1)                                              2,75 (3)II  Ddz-(7-12)-OBzl        b         1,82 (2)                      1,96 (2)                          1,08 (1)    1,00 (1)I   Ac-(1-6)-OBzl        b     1,97 (2)                      0,99 (1)                              1,95 (2)                                  1,00 (1)__________________________________________________________________________ Hydrolysis method: a = HCl/propionic acid 160° C., 45 to 60 minutes b = 6 n HCl 110° C., 18 to 24 hours 
    
     
                                           TABLE XI__________________________________________________________________________Amino acid analysis of Thymosin α.sub.1 -Fragments I to IXFrag-        Hydrolysisment         Method              Asp Thr Ser Glu Ala Val Ile Leu Lys__________________________________________________________________________V  Ddz-(25-28)-OBzl        a     1,00 (1)    1,90 (2)                              1,20 (1)IV Ddz-(20-24)-OH        a                 1,82 (2)                                  2,00 (2)    1,05 (1)VI Ddz-(20-28)-OBzl        b     1,70 (1)    4,3 (4)                              1,00 (1)                                  1,36 (2)    1,00 (1)III   Ddz-(13-19)-OH        b     1,12 (1)                  0,85 (1)                          1,02 (1)        1,00 (1)                                              2,75 (3)VII   Ddz-(13-28)-OBzl        b     1,94 (2)                  0,7 (1) 4,44 (5)                              0,71 (1)                                  1,44 (2)                                          1,00 (1)                                              3,76 (4)II Ddz-(7-12)-OMe        a         1,52 (2)                      1,38 (2)                          0,77 (1)    1,00 (1)II Ddz-(7-12)-OH        b         1,75 (2)                      1,78 (2)                          1,16 (1)    1,00 (1)VIII   Ddz-(7-28)-OBzl        a     2,09 (2)                  2,56 (3)                      1,07 (2)                          5,22 (6)                              0,78 (1)                                  1,44 (2)                                      0,85 (1)                                          1,15 (1)                                              3,91 (4)I  Ac-(1-6)-OH        b     1,84 (2)                      0,91 (1)                              1,97 (2)                                  1,0.0 (1)IX Ac-(1-12)-OBzl        b     2,08 (2)                  1,87 (2)                      2,5 (3)                          0,56 (1)                              2,42 (2)                                  1,00 (1)                                      0,96 (1)IX Ac-(1-12)--OH        b     2,12 (2)                  1,77 (2)                      2,45 (3)                          1,00 (1)                              2,12 (2)                                  1,00 (1)                                      1,10 (1)Ia Ddz-Lys-(2-6-OBzl        a     2,10 (2)        1,92 (2)                                  1,00 (1)    1,03 (1)IVa   Ddz-(20-23)-Gla-        a                 2,11 (2)                                  2,00 (2)    1,02 (1)   OMeVa Ddz-Gla-(25-28)-        a     0,87 (1)    2,09 (2)                              1,00 (1)   OBzlVIa   Ddz-(20-23-Gla.sub.2 -        a     1,11 (1)    4,23 (4)                              1,00 (1)                                  1,79 (2)    0,98 (1)   25-28)-OBzl__________________________________________________________________________ Hydrolysis method: a = HCl/propionic acid 160° C., 45 to 60 minutes b = 6 n HCl 110° C., 18 to 24 hours 
    
     EXAMPLE 9 
     In order to prove the immunostimulant and immunoregulating properties of the thymosin α 1  fragments of the invention, immunological-pharmacological tests were performed. 
     The immunological test methods used were the E-rosette test according to Wara, Ammann et al. (N.Y. Acad. Sci. 249 (1975) 308 and N. Engl. J. Med. 292 (1975) 70) and the method of mixed lymphocyte culture, both methods involving additional inhibition of cellular RNS polymerase by alphaamanitin. The E-rosette test is based on the fact that the percentage of E-rosette forming cells in peripheral human blood is a measure of the content of fully mature T-cells, which form with sheep&#39;s red blood cells characteristic aggregates which can be counted under the microscope. Experiments by Wara, Ammann et al (loc. cit.) have shown that in patients with thymus hypoplasia an increase from 20% to 45% can be achieved in the E-rosette formation, provided that an optimally active thymosin preparation is added. The normal level of E-rosette formation in the healthy human subject is 56%. 
     For the performance of the E-rosette test, peripheral human lymphocytes are isolated by a gradient centrifugation. Their RNS polymerase is blocked by alpha-amanitin. Then red blood cells from sheep and thymosin or a thymosinα 1  fragment is added, which removes the alpha-amanitin blockage of the E-rosette formation, so that the normal level of the cell aggregate formation, which is 56%, can be achieved. This value is taken as 100% in a relative numerical scale, and after the alpha-amanitin blockage it is taken to be 0%; see Table XII. 
     The test in mixed lymphocyte cultures is based on the fact that T-cells are made to proliferate by stimulation with allogenic antigens. For the performance of the test, the lymphocytes of donor A (&#34;responder&#34;) are incubated with those of donor B (&#34;stimulator&#34;) for 4 to 5 days. Then  3  H-thymidin is added and the rate of proliferation is measured on the basis of the increasing radioactivity of the cell culture due to the installation of the radioactive material. Since both populations A and B might be made to proliferate, resulting in an unclear effect, population B is blocked with mitomycin C in all metabolical activities. This test is routinely used before organ transplants to determine the immunological compatibility of donor and receiver tissues. 
     The results obtained in these pharmacological studies are given in the following Tables XII and XIII. 
     
                                           TABLE XII__________________________________________________________________________Examples of the stimulation of human peripheral lymphocytes withthymosin α.sub.1 fragments in the E-rosette test after the cellsareblocked with α-amanitin (E-rosette number being then 0%;The normal level without any addition = 100%)Fragment   Ia     II       III         IV           V  VI                VII                   IX                     Thymosin                           (Synthetic)__________________________________________________________________________E-rosette   93     29       29         45            42              16                41 40                     100number (%)Optimum 16     80       80         83           113              10                 6  4                     1.6concentr. (μM)__________________________________________________________________________ 
    
     
                       TABLE XIII______________________________________Examples of the stimulation of human peripheral lymphocytes bythymosin α.sub.1 fragments in the test with mixed lymphocytecultures,with and without blockage by α-amanitin.(1 μg thymosin α.sub.1 fragment per ml of cell culture; 1 μgofα-amanitin per ml of cell culture (+) = stimulation (-) =inhibition (%) of the culture by addition of thymosin α.sub.1fragmentsThe reaction of the mixed lymphocyte culture withoutsuch addition is taken as 0%)Fragment   Ia     II     III  IV    V     VI    IX______________________________________Cell pairsAB      -8     --     -64  -53   +223  +153  -74CD      -73    -53    --13 +37   +45   +57   -63BA      -15    -18    -12  +565  +791  +1136 --DC      -31    +30    +12  +2    +37   ±0 -77Cell pairs*AB      -10    -11    - 44 -87   -47   +279  -30CD      -53    -98    -20  +35   --    +83   +63BA      --     -80    +6   -97   -94   -96   -78DC      -7     +20    +80  +100  --    +160  +60______________________________________ *Cultures blocked with amanitin 
    
     From the above Tables XII and XIII is is immediately apparent that the thymosin α 1  fragments of the invention, as shown by the Examples in Tables XII and XIII, exercise a very pronounced immunological action which enables them to be used in human therapy and in veterinary medicine. 
     If the immunological activities of the fragments, as shown in the above example, are compared with those of the parent substance, totally synthetic thymosin α 1  (shown in Table XIV for the E-rosette test and in Table XV for the mixed lymphocyte culture), it can be concluded that all of the fragments studied have a pronounced action on the differentiating mechanism and on the maturation of thymus-related lymphocytes to immune-competent T-cells, as well as in their proliferation process. 
     Comparisons in detail of the immunological activities of the fragments with thymosin α 1  clearly show that mechanisms of T-cell activation which express themselves in the E-rosette test are more strongly stimulated by N-terminal thymosin α 1  fragments (VII, IX) than by the C-terminal fragments IV, V and VI. Here a special position is assumed by the especially active, unnatural fragment Ia, which, with its additional basic function (lysine) apparently exercises an effect on T-lymphocytes, which in the case of the natural fragments is brought about only in combination with median sections of the sequence. These fragments, which are especially active in the E-rosette test, also display a surprisingly pronounced inhibiting action in the mixed lymphocyte culture (see Table XIII for fragments Ia and IX) on the opposite cell-bound antigenicity of lymphocytes from different donors. This finding can acquire considerable importance in transplantation practice due to the possibility of the preparation of medicaments having an immunosuppressive action on the basis of small, easy-to-synthesize peptides. 
     On the basis of the comparisons of thymosin α 1  with the fragments IV, V and VI, however, it can be concluded from both of the bioassays that the immunologically stimulative center of thymosin α 1  is located in the C-terminal area. Special attention must be called to the easy-to-synthesize peptides Glu-Ala-Glu-Asn (fragment V) and Lys-Glu-Val-Val-Glu (fragment IV), which show activity in the E-rosette test only at a relatively high dosage, but develop a very considerable stimulation of T-cell proliferation in the mixed lymphocyte culture. 
     This is because, when there is a lack of immune-competent T-cells (T-lymphocytes), the immunological recognition (cell-bound antibody formation) and defense of abnormal body cells is reduced. Cellular abnormalities may be due to virus infections, and various forms of development of cancer. 
     The results presented indicate that those fragments of thymosin α 1  which display activities equivalent to or more powerful than the parent substance ought to be especially suitable for the preparation of medicaments against immunological deficiency diseases, senilities and virus infections, but above all against various forms of cancer (leukemia, pulmonary metastases). 
     
                       TABLE XIV______________________________________Data on a comparison of synthetic thymosin α.sub.1 with anatural polypeptide prepared from thymus glands,in the Erosette test with inhibition by α-amanitin.Normal cell population:             64%.sup.(a)                       E rosettesInhibited with α-amanitin:             25%.sup.(a)                       E-rosettesThymus polypeptide.sup.(b)             Synthetic thymosin α.sub.1.sup.(c)μg per culture (%)             μg per culture (%).sup.(d)______________________________________6.25    10            5.00      333.12    67            2.50      571.25    100           1.00      640.62    62            0.50      1000.31     6            0.25      39______________________________________ .sup.(a) Absolute value .sup.(b) Isolated from thymus glands .sup.(c) Analytically pure .sup.(d) Relative number of the Erosettes produced by stimulation with thymus peptides after their previous inhibition with amanitin, from peripheral human lymphocytes. 
    
     
                       TABLE XV______________________________________The-stimulation by synthetic thymosin α.sub.1 of an allogenicmixed culture from peripheral human lymphocytes(stimulator cells blocked with mitomycin)                  Stimulation.sup.(c)Cell pairs     (%).sup.(a)                  .sup.(b)______________________________________AB             +20     +86AD              +4     +37DB             +27     +1550BA             +27     +47BD             +45     +51DA             +44      ±0______________________________________ .sup.(a) Culture not inhibited .sup.(b) Culture inhibited with 1 microgram of amanitin per ml of the cel suspension .sup.(c) Culture stimulated by the addition of 1 microgram of thymosin α.sub.1 per ml of cell suspension; the lymphocyte stimulation in th culture without the addition of thymosin α.sub.1 was taken as 0%. 
    
     EXAMPLE 10 
     Preparation of Ddz-Lys(Z)-Lys(Z)-Glu(OBu t )-Val-Val-Glu(OBu 6 )-OMe, protected on all sides (19 to 24) 
     Reaction Solution A: 
     1.0 g of Ddz(20-24)-OMe was dissolved in 14 ml of dichloromethane to which 0.7 ml of trifluoroacetic acid had been added, in order to split off the Ddz protective group. After 30 minutes at 20° C., neutralization was performed with 1.03 ml of N-methylmorpholine. The gel that formed was redissolved by the addition of 5 ml of dimethylformamide. 
     Reaction Solution B 
     693 mg of Ddz-Lys(Z)OH was dissolved in 15 ml of dichloromethane under anhydrous conditions, chilled to -15° C., and 152 μl of N-methylmorpholine and 162 μl of isobutyloxycarbonyl chloride with vigorous agitation. After 8 minutes of reaction at -15° C., the above Solution A was added and the mixture was stirred for another 3 hours without refrigeration. 
     For working up, the reaction mixture was diluted with 100 ml of chloroform, extracted at 0° C. with 0.5M KHSO 4  solution and 5% KHCO 3  solution and water, and concentrated in vacuo at 30° C. 
     After crystallization from a mixture of ethyl acetate and ether, yield 1.125 g (91% of the theory), MP 216°-218° C. (decomposition). 
     Amino acid analysis: Glu 2.20(2), Val 1.94(2) Lys 2.00(2) thin layer chromatography (Merck silica gel F 254 , 0.250 mm). 
     
         ______________________________________R.sub.f × 100    Solvent______________________________________73       CHCl.sub.3 /CH.sub.3 OH/CH.sub.3 CO.sub.2 H                        85:10:5 (V/V)60       CHCl.sub.3 /C.sub.2 H.sub.5 OH                        9:1 (V/V)______________________________________ 
    
     EXAMPLE 11 
     Preparation of [18-24] protected on all sides, Ddz-Glu(OBu t )-Lys(Z)-Glu(OBu t )Val-Val-Glu(OBu t )OMe 
     735 mg of Ddz[19-24]OMe, obtained in accordance with Example 1, is freed of the Ddz- protective group by the above-described Procedure A and reacted with the reaction solution of Procedure B, of 787 mg of Ddz-Glu(OBu t )CHA (CHA=cyclohexylammonium salt) and 97 microliters of isobutyloxycarbonyl chloride for 5 hours, and the product is worked up as described therein. 
     After crystallization from ethyl acetate, yield 730 mg (87% of the theory) amino acid analysis: Glu 2.99(3), Val 2.00(2), Lys 1.91(2). 
     
         ______________________________________R.sub.f × 100    Solvent______________________________________69       CHCl.sub.3 /CH.sub.3 OH/CH.sub.3 CO.sub.2 H                        85:10:5 (V/V)59       CHCl.sub.3 /CH.sub.3 OH                        9:1 (V/V)______________________________________ 
    
     EXAMPLE 12 
     Preparation of [17-24] protected on all sides, Ddz-Lys(Z)-Glu(OBu t )-Lys(Z)-Glu(OBu t )-Val-Val-Glu(OBu t )OMe 
     350 mg of Ddz[18-24]OMe, obtained as in Example 2, is stripped of the Ddz protective group in accordance with Procedure A and reacted for 5 hours with the reaction solution of Procedure B of 186 mg of Ddz-Lys(Z)OH and 43.2 microliters of isobutyloxycarbonyl chloride, and the product is worked up as described therein. 
     After recrystallizing twice from boiling methanol, yield 245 mg (60% of the theory). Amino acid analysis: Glu 3.38(3) Val 2.00(2) Lys 2.80(3) 
     
         ______________________________________R.sub.f × 100    Solvent______________________________________67       CHCl.sub.3 /CH.sub.3 OH/CH.sub.3 CO.sub.2 H                        85:10:5 (V/V)______________________________________ 
    
     EXAMPLE 13 
     Preparation of [25-27] protected on all sides, Ddz-Glu(OBu t )-Ala-Glu(OBu t )OBu t   
     1.5 g of Ddz-Ala-Glu(OBu t )OBu t  is freed of the Ddz protective group by Procedure A and reacted for 2 hours with the reaction solution corresponding to Procedure B of 2.84 g of Ddz-Glu-(OBu t )CHA and 0.62 ml of isobutyloxycarbonyl chloride. After working up as described in that procedure, the product was chromatographically purified on Sephadex LH 20 in methanol and crystallized from a mixture of ether and benzine (40° C.). 
     Yield 1.7 g (85% of the theory), MP 88° to 90° C. 
     Amino acid analysis: Glu 1.70 (2) Ala 1.00 (1) 
     
         ______________________________________R.sub.f × 100      Solvent______________________________________45         CHCl.sub.3 /CH.sub.3 COCH.sub.3                    85:15 (V/V)______________________________________ 
    
     EXAMPLE 14 
     Preparation of [20-25] protected on all sides, Ddz-Lys(Z)-Glu(OBu t )-Val-Val-Glu(OBu t )-Glu(OBu t )OBu t   
     250 mg of Ddz-[20-24]OH, prepared in accordance with Example 1, and 73.5 mg of 1-hydroxybenzotriazole (HOBT) are dissolved in 20 ml of absolute tetrahydrofuran and reacted at 0° C. with 50 mg of dicyclohexylcarbodiimide, followed by 65 mg of HGlu(OBu t )OBu t , for 2 hours of reaction time at 0° C. and 4 hours at 20° C. After concentration in vacuo at 30° C., the product is dissolved with 50 ml of ethyl acetate, the dicyclohexylurea is filtered out, and the filtrate is extracted with 0.5M KHSO 4  solution, 5% KHCO 3  solution and water. The organic phase is filtered through bas[ic] Al 2  O 3  (column 3×10 cm) which is eluted with 250 ml of ethyl acetate. The eluate is concentrated in vacuo at 30° C., leaving the end product as a colorless glass in pure form. 
     Yield 160 mg (52% of the theory). 
     Amino acid analysis: Glu 2.90 (3), Val 2.00 (2), Lys 0.99 (1). 
     
         ______________________________________R.sub.f × 100    Solvent______________________________________91       CHCl.sub.3 /CH.sub.3 OH/CH.sub.3 CO.sub.2 H                        85:10:5 (V/V)58       CHCl.sub.3 /C.sub.2 H.sub.5 OH                        9:1 (V/V)______________________________________ 
    
     EXAMPLE 15 
     Preparation of [25-26] protected on all sides, Ddz-Glu(OBu t )-AlaOBzl 
     In accordance with Procedure B, 2.1 g of Ddz-Glu(OBu t )CHA was activated with 468 microliters of isobutyloxycarbonyl chloride and reacted with 500 mg of HCl.AlaOBzl and 443 microliters of N-methylmorpholine. After concentration in vacuo at 30° C., the oily concentrate is dissolved with 5 ml of ethyl acetate and filtered with basic Al 2  O 3  on a column (3×10 cm). 
     After elution with 250 ml of ethyl acetate and concentration in vacuo at 30° C., the pure product is obtained as a colorless glass. 
     Yield: 1.31 g (96% of the theory) 
     
         ______________________________________R.sub.f × 100    Solvent______________________________________85       CHCl.sub.3 /CH.sub.3 OH/CH.sub.3 CO.sub.2 H                        85:10:5 (V/V)51       CHCl.sub.3 /CH.sub.3 COCH.sub.3                        85:15 (V/V)______________________________________ 
    
     EXAMPLE 16 
     Preparation of [20-26] protected on all sides, Ddz-Lys(Z)-Glu(OBu t )-Val-Val-Glu(OBu t )-Glu(OBu t )-AlaOBzl 
     By Procedure A, the protective group Ddz- was split off from 176 mg of Ddz[25-26]-OBzl prepared as in Example 6, and the product was neutralized with 330 microliters [word or words omitted] and concentrated in vacuo at 20° C. The concentrate was dissolved with 2 ml of dimethylformamide and added to the batch next described. 
     250 mg of Ddz[20-24]OH and 73.5 mg of HOBT were dissolved in 3 ml of dimethylformamide, activated at 0° C. with 100 mg of dicyclohexylcarbodiimide, and combined with the above solution of H[25-26]Bzl. After one hour of reaction at 0° C. and three days at 20° C., the product was worked up in accordance with the synthesis of fragment [20-25] and purified. 
     After recrystallization from methanol, yield 190 mg (58% of the theory) 
     Amino acid analysis: Glu 3.06 (3) Ala 1.00 (1) Val 1.77 (2) Lys 1.05 (1). 
     
         ______________________________________R.sub.f × 100        Solvent______________________________________78           CHCl.sub.3 /C.sub.2 H.sub.5 OH                     9:1 (V/V)______________________________________ 
    
     The R f  -values for a number of α 1  -thymosin fragments in accordance with the invention are given in the following Table 2. They were determined on thin-coated silica gel plates, 0.25 mm, Merck 60 F 254 , and are given as R f  ×100 values. 
     
         ______________________________________Solvent 1: n-butanol/pyridine/glacial acetic acid/water      5:5:1:4 (V/V)Solvent 2: acetic ester/pyridine/glacial acetic acid/      water 15:20:6:11 (V/V)Solvent 3: N--pentanol/pyridine/2-butanone/formic acid/      water 40:28:11:5:15 (V/V)______________________________________ 
    
     The fragments are identified by the numbering given in Table A. 
     It will be understood that the specification and examples are illustrative but not limitative of the present invention and that other embodiments within the spirit and scope of the invention will suggest themselves to those skilled in the art.