Efficient solution-phase synthesis of multiple O-phosphoseryl-containing peptides related to casein and statherin

The multiple Ser(P)-containing peptides, H-Ser(P)-Ser(P)-Ser(P)-Glu-Glu-NHMe-TFA, H-Asp-Ser(P)-Ser(P)-Glu-Glu-NHMe-TFA and H-Glu-Ser(P)-Ser(^P)-Glu-Glu-NHMe-TFA were prepared by the use of Boc-Ser(PO₃Ph₂)-OH in the Boc mode of solution phase peptide synthesis followed by platinum-mediated hydrogenolytic de-protection of the Ser(PO₃Ph₂)-containing peptides. The protected peptides were assembled using the mixed anhydride coupling methods with 40% TFA/CH₂C1₂ used for removal of the Boc group from intermediate Boc-protected peptides.     

Efficient synthesis of metal binding peptides incorporating aminodiacetic acid based ligands

New N^x-Fmoc/Bu^t protected amino acids bearing half-EDTA side chains (CH₂)_nN(Ada-O-Bu¹)₂n= 1 (5), n= 2 ( 24 ), n= 3 ( 10 ), n= 4 ( 15 ) were prepared in satisfactory yields. These derivatives can be conveniently used in a solid-phase peptide synthesis as they are devoid of serious shortcomings of Boc/Bzl based syntheses of metallopeptides, such as preliminary peptide capping as well as undesired lactamization of 5 during the peptide synthesis.     

Fmoc/solid-phase synthesis of Tyr(P)-containing peptides through t-butyl phosphate protection

The synthesis is of Tyr(P)-containing peptides by the use of Fmoc-Tyr(PO₃Me₂)-OH in Fmoc/solid phase synthesis is complicated since, firstly, piperidine causes cleavage of the methyl group from the -Tyr(PO₃ Me₂)-residue during peptide synthesis and, secondly, harsh conditions are needed for its final cleavage. A very simple method for the synthesis of Tyr(P)-containing peptides using t-butyl phosphate protection is described. The protected phosphotyrosine derivative, Fmoc-Tyr(PO₃^tBu₂)-OH was prepared in high yield from Fmoc-Tyr-OH by a one-step procedure which employed di-t-butyl N,N-diethyl-phosphoramidite as the phosphorylation reagent. The use of this derivative in Fmoc/solid phase peptide synthesis is demonstrated by the preparation of the Tyr(P)-containing peptides, Ala-Glu-Tyr(P)-Ser-Ala and Ser-Ser-Ser-Tyr(P)-Tyr(P).     

The preparation of three new partially deuterated hexadecanethiols for applications in surface chemistry

The synthesis of three partially deuterated hexadecanethiols has been achieved. Thiols CH₃(CH₂)₇(CD₂)₈SH, CD₃(CD₂)₇(CH₂)₈SH and CD₃(CH₂)₁₅SH were targeted, as these compounds, after formation of self‐assembled monolayers on Au(1 1 1) or Au nanoparticles, can provide mechanistic information pertaining to reactive atom migrations within the assembly. The syntheses of each of these compounds called upon Grignard coupling chemistry, which was activated by Li₂CuCl₄. Applicable deuterium containing fragments were either commercially obtained or constructed from by way of an inexpensive and efficient ring opening, protection and dimerization of THF‐d₈. Sulfur incorporation was by thiolacetate substitution or addition reactions. The protocols presented possess general applicability in a number of syntheses requiring block‐deuterated fattyalkyl sections. Copyright © 2008 John Wiley & Sons, Ltd.     

Copper(I) (Pseudo)Halide Complexes with Neocuproine and Aminomethylphosphines Derived from Morpholine and Thiomorpholine – In Vitro Cytotoxic and Antimicrobial Activity and the Interactions with DNA and Serum Albumins

Herein, a series of CuI or CuNCS complexes with neocuproine (2,9‐dimethyl‐1,10‐phenanthroline: dmp) and two tris(aminomethyl)phosphines derived from morpholine (P(CH₂N(CH₂CH₂)₂O)₃) or thiomorpholine (P(CH₂N(CH₂CH₂)₂S)₃) were tested as cytotoxic agents in vitro towards mouse colon carcinoma (CT26) and human lung adenocarcinoma (A549). The studies showed that the complexes exhibit potential antitumor properties, displayed by IC₅₀ values below 10 μm towards the tested cell lines, in the case of 4‐h incubation time with the examined compounds. Moreover, a high antimicrobial activity of all the complexes was observed against Staphylococcus aureus and Candida albicans with minimal inhibitory concentrations equal to 1–2 μg/mL. To gain insight into the molecular mechanism of biological activity of the complexes, we investigated also their interactions with plasmid DNA (pUC18) and the human and bovine serum albumins. Gel electrophoresis experiments demonstrated that all the compounds were comparably efficient in DNA degradation process; however, luminescence quenching showed surprising dependence on the interactions strength of the used compounds with the albumins. Apart from exceptionally effective [CuI(dmp)P(CH₂N(CH₂CH₂)₂O)₃], the complexes with P(CH₂N(CH₂CH₂)₂O)₃ quenched more strongly luminescence of bovine serum albumin, while the complexes with P(CH₂N(CH₂CH₂)₂S)₃ were more active in the quenching of human serum albumin luminescence.     

Solid-phase synthesis of protected peptides using new cobalt(III) ammine linkers†

Cobalt(III) ammine complexes of the type cis-[CoL₄(4-AMB)O-AA-Boc](CF₃SO₃)₂, where L₄= bisethylenediamine (en)₂ or tetraammine (NH₃)₄, and 4-AMB = 4-(aminomethyl)benzoic acid, have been synthesized and used as linkers to polystyrene resins for solid-phase synthesis of protected peptides. Boc/t-Bu-protected [Leu⁵]enkephalin was assembled on the two different Co(III) resins, and then cleaved from the resins by reduction of the Co(III) center in 93–96%; yield. HPLC-purified protected [Leu⁵]enkephalin was obtained in 67–69% overall yield and characterized by amino acid analysis and ¹H NMR. Stepwise synthesis on the Co(en)₂-resin was also used in the assembly of Boc-Asp(OcHex)-Arg(Mts)-Gly-Asp(OcHex)-Ala-Pro-Lys(2Cl-Z)-Gly-OH, a sequence from collagen α1 Type 1. The protected peptide was cleaved from the Co(III) resin in 74% yield, and the HPLC-purified nonapeptide was characterized by amino acid analysis, ¹H NMR and liquid secondary-ion mass spectrometry (LSIMS). New routes are described for the synthesis of isomerically pure Co(III) anchor complexes. The Co(III) resins were found to be compatible with both the tert-butyloxycarbonyl (Boc) and the 9-fluorenylmethoxycarbonyl (Fmoc) N^α-protecting group strategies used in solid-phase peptide synthesis.     

Efficient solution phase synthesis and use of multiple O-phosphothreonyl-containing peptides for calcium phosphate binding studies

The protected phosphothreonine derivative Boc-Thr(PO₃Ph₂)-OH was prepared in high yield from Boc-Thr-OH by a simple three-step procedure which involved 4-nitrobenzylcarboxyl protection, either phosphorotriester (diphenyl phosphorochloridate) or “phosphite-triester” (diphenyl N.N-diethylphosphoramidite) phosphorylation of the thrconine hydroxyl group of Boc-Thr-ONb followed by hydrogenolytic carboxyl deprotection. The three Thr(P)-containing peptides, H-Thr(P)-Glu-Glu-NHMe.TFA, H-Thr(P)-Thr(P)-Glu-Glu-NHMe.TFA and H-Thr(P)-Thr(P)-Thr(P)-Glu-Glu-NHMe.TFA, were prepared in high yield by the use of Boc-Thr(PO₃Ph₂)-OH in the Boc mode of peptide synthesis (mixed anhydride method) followed by platinum-mediated hydrogenolytic deprotection of the Thr(PO₃3Ph₂)-containing peptides. The use of the phosphopeptides in calcium phosphate binding studies showed that the triple Thr(P)-cluster was a basic structural requirement, since only the pentapeptide was able to bind calcium phosphate efficiently.     

Incorporation of vinylogous scaffolds in the C‐terminal tripeptide of substance P

Abstract: Glycine‐9 and leucine‐10 of substance P (SP) are critical for (NK)‐1 receptor recognition and agonist activity. Proψ(Z)‐CH=CH(CH₃)‐CONH)Leu (or Met) and Proψ((E)‐CH=CH(CH₃)‐CONH)Leu (or Met) have been introduced in the sequence of SP, in order to restrict the conformational flexibility of the C‐terminal tripeptide, Gly‐Leu‐Met‐NH₂, of SP. Proψ((Z)‐CH=C(CH₂CH(CH₃)₂)‐CONH)Met‐NH₂, with an isobutyl substituent to mimic the Leu side‐chain, was also incorporated in place of the C‐terminal tripeptide. The substituted‐SP analogs were tested for their affinity to human NK‐1 receptor specific binding sites (NK‐1M and NK‐1m) and their potency to stimulate adenylate cyclase and phospholipase C in Chinese Hamster ovary (CHO) cells transfected with the human NK‐1 receptor. The most potent SP analogs [Pro⁹ψ((Z)CH=C(CH₃)CONH)Leu¹⁰]SP and [Pro⁹ψ ((E)CH=C(CH₃)CONH)Leu¹⁰]SP, are about 100‐fold less potent than SP on both binding sites and second messenger pathways. These vinylogous (Z)‐ or (E)‐CH=C(CH₃)‐ or (Z)‐CH=C(CH₂CH(CH₃)₂) moieties hamper the correct positioning of the C‐terminal tripeptide of SP within both the NK‐1M‐ and NK‐1m‐specific binding sites. The origin of these lower potencies is related either to an incorrect peptidic backbone conformation and/or an unfavorable receptor interaction of the methyl or isobutyl group.     

An improved synthesis of [11C]MENET via Suzuki coupling with [11C]methyl iodide

[¹¹C]MENET, a promising norepinephrine transporter imaging agent, was prepared by Suzuki cross coupling of 1 mg N‐t‐Boc pinacolborate precursor with [¹¹C]CH₃I in DMF using palladium complex generated in situ from Pd₂(dba)₃ and (o‐CH₃C₆H₄)₃P together with K₂CO₃ as the co‐catalyst, followed by deprotection with trifluoroacetic acid. This improved radiolabeling method provided [¹¹C]MENET in high radiochemical yield at end of synthesis (EOS, 51 ± 3%, decay‐corrected from end of ¹¹CH₃I synthesis, n = 6), moderate specific activity (1.5–1.9 Ci/µmol at EOS), and high radiochemical (>98%) and chemical purity (>98%) in a synthesis time of 60 ± 5 min from the end of bombardment. Copyright © 2013 John Wiley & Sons, Ltd.     

Peptide N-alkylamides by solid phase synthesis*

Three new resins have been developed that allow for the solid phase synthesis of C-terminal peptide N-alkylamides using Boc amino acids, usual side chain protecting groups and hydrogen fluoride cleavage and deprotection. These resins were prepared by reacting the appropriate alkylamine (NH₂ CH₃, NH₂ CH₂ CH₃, NH₂ CH₂ CF₃) to Merrifield's 1% divinylbenzene cross-linked chloromethylated polystyrene resin. The application of these resins to the synthesis of C-terminal GnRH N-alkylamides illustrates the versatility of this approach. GnRH analogs were tested for their ability to release LH from cultured rat anterior pituitary cells. [D Glu⁶, Pro⁹-NHCH₂ CH₃]-GnRH was synthesized for the first time using the solid phase approach and found to be three times more potent than [D Glu⁶]-GnRH. Other analogs including [D Trp⁶, Pro⁹-NHCH₂ CH₃]-GnRH, [D Ala⁶, Pro⁹-NHCH₂ CF₃]-GnRH and related peptides were found to be equipotent and to have the same properties (HPLC retention times, amino acid analysis and specific rotation) as the corresponding peptides synthesized using less amenable strategies; yields were equivalent or better than those reported earlier.     

Orthogonal solid-phase synthesis of human gastrin-I under mild conditions*

An approach to the solid-phase segment condensation synthesis of the 17-peptide amide human gastrin-I has been developed. N^α-amino and side-chain protection were provided by 9-fluorenylmethyloxycarbonyl (Fmoc) and tert.-butyl groups, and a series of anchors cleavable under mild conditions were used. The N-terminal pentapeptide pGlu-Gly-Pro-Trp-Leu-OH was prepared using a p-alkoxybenzyl ester linkage made by a preformed handle strategy. Cleavage, in 65% yield, was with the new Reagent M: CF₃ COOH—CH₂ Cl₂—β-mercaptoethanol-anisole (70:30:2:1), which was optimized to preserve the labile tryptophan residue. A new preformed handle procedure expedited solid-phase synthesis of the protected “middle” hexapeptide, Fmoc-(Glu(OtBu))₅-Ala-OH, anchored as an o-nitrobenzyl ester. Chains were not lost during this assembly, and final photolytic cleavage (350nm) in toluene—CF₃ CH₂ OH (4:1) occurred in 59% yield. Both protected intermediates were purified by simple gel filtration, whereupon they were shown to be pure by analytical HPLC, and gave satisfactory NMR and FABMS spectra. Last, the C-terminal hexapeptide, Tyr(tBu)-Gly-Trp-Met-Asp(OtBu)-Phe, was assembled on a tris(alkoxy)benzylamide “PAL” support. For the polymer-supported segment condensation, the middle and N-terminal pieces were added respectively in > 98% and 89% yields (judged by amino acid analysis and solid-phase sequencing), by overnight couplings in N,N-dimethylformamide (DMF) mediated by benzotriazolyl N-oxytrisdimethylaminophosphonium hexafluorophosphate (BOP) in the presence of 1-hydroxybenzotriazole (HOBt) and N-methylmorpholine (NMM). Racemization was 4% and 11% respectively at Ala and Leu. Cleavage with Reagent M followed by reversed-phase chromatography gave pure gastrin-I in an overall 30% isolated yield. These results compare favorably with those from a stepwise assembly.     

Synthesis and pharmacological activity of the N-terminal dermorphin tetrapeptide analogs with CH2-NH peptide bond isosteres

The synthesis of pseudotetrapeptides H-Tyr-D-Ala-Phe-NH-(CH₂)₂-NH₂ (1a), H-Tyr-D-Ala-Phe-ψ(CH₂-NH)-Gly-NH₂ (2a), H-Tyr-D-Ala-ψ(CH₂-NH)-Phe-Gly-NH₂ (3a), and H-Tyr-ψ(CH₂-NH)-D-Ala-Phe-Gly-NH₂ (4a), representing the N-terminal tetrapeptide sequence of dermorphin, in which amide bonds are replaced by CH₂-NH bond, is described. N-acetyl-Tyr and desamino-Tyr pseudopeptide analogs (1-4b), (1-3c) are also described. The analogs were assayed in binding studies based on displacement of μ and δ-receptor selective radiolabels from rat brain membrane and in a bioassay using guinea pig ileum (GPI). Pseudopeptides in which the C-terminal (1a) or D-Ala-Phe (3a) amide bond are substituted, exhibit higher μ-affinities and μ-receptor selectivity than the corresponding Phe-Gly or Tyr-D-Ala analogs (2a, 4a). Acetyl-and desamino-Tyr pseudopeptide analogs (1-4b) and (1-3c) did not exhibit μ and δ-opioid receptor affinity at nM concentration. The relevance of the single peptide replacement and of its association to acetylation or amino group elimination of Tyr, is discussed on the basis of a receptor model for μ and δ opioids.     

Oxidative metabolism of the GABAA receptor antagonist t-butylbicycloortho[3[H]benzoate

1. The trioxabicyclooctane ring of t-butylbicycloortho[³H]benzoate (TBOB), (CH₃)₃CC(CH₂O)₃CC₆H₅, is cleaved to yield the 3-oxo-benzoate, (CH₃)₃CC(CHO)(CH₂OH)CH₂OC(O)C₆H ₅, on O-methylene hydroxylation by microsomes from mouse liver or houseflies in the presence of NADPH.

2. The methyl and phenyl substituents are tentatively identified as additional sites of oxidative metabolism.

3. The 3-oxo-benzoate from oxidative cage opening and the bis-(hydroxymethyl)-benzoate, (CH₃)₃CC(CH₂OH)CH₂OC(O)C₆H ₅, from enzymic reduction of the 3-oxobenzoate undergo esteratic hydrolysis to benzoic acid.

4. Metabolites of TBOB excreted by rats and houseflies include the bis-(hydroxymethyl)-benzoate and benzoic and hippuric acids.

5. Metabolic hydroxylation of TBOB at O-methylene, alkyl and aryl substituents may serve as a model for detoxication reactions of related potent GABA_A receptor antagonists and insecticides.     

DESIGN AND SYNTHESIS OF POTENT IN VIVO ANTAGONISTS OF OXYTOCIN

We have previously shown that the substitution of 8-ornithine and 2-O-methyltyrosine alone and in combination in [1-deaminopenicillamine] oxytocin (dPOT) brought about enhancements in antagonistic potencies to responses to oxytocin in vivo. To explore the effects of these subtitutions in analogs of dPOT containing larger alkyl substitutents on the β carbon at position 1 and on the tyrosine residue at position two, the following six analogs were synthesized: [1-(β-mercapto-β, β-diethylpropionic acid), 8-ornithine] vasotocin (1, dEt₂OVT); [1-β-mercapto-β, β-cyclopentamethylenepropionic acid), 8-ornithine] vasotocin [2, d(CH₂)₅OVT); [1-β-mercapto-β, β-diethylpropionic acid), 2-O-methyltyrosine, 8-ornithine]vasotocin [3, dEt₂ Tyr(Me)OVT]; [1-(β-mercapto-β, β-diethylpropionic acid), 2-O-ethyltyrosine, 8-ornithine] vasotocin [4, dEt₂ Tyr(Et)OVT]; [1-β-mercapto-β', β-cyclopentamethylenepropionic acid), 2-O-methyltyrosine, 8-ornithine] vasotocin [5, d(CH₂)₅ Tyr(Me)OVT]; [1-β-mercapto-β, β-cyclopentamethylenepropionic acid), 2-O-methyltyrosine, 8-ornithine] vasotocin [6, d(CH₂)₅ Tyr(Et)OVT]. The required protected intermediates were synthesized by a combination of solid-phase synthesis and by individual 8 + 1 couplings in solution. All six analogs antagonize the actions of oxytocin on the rat uterus in the absence of Mg²⁺, in the presence of 0.5 mM Mg²⁺ and in situ. They also antagonize milk ejection responses to oxytocin, and the vasopressor responses to arginine vasopressin, and all have very low antidiuretic activities. With pA₂ values of 7.35 ± 0.08 and 7.37 ± 0.17, respectively, compounds 3 and 5 are the two most potent in vivo antagonists of oxytocin reported to date.     

Cadmium Mobilization in Vivo by Intraperitoneal or Oral Administration of Monoalkyl Esters of meso–2, 3–Dmercaptosuccinic Acid in the Mouse

Abstract: The relative activities of a series of nine monoalkyl esters of meso-2, 3–dimercaptosuccinic acid have been examined as agents for the mobilization of cadmium from mice one week after intraperitoneal administration of cadmium chloride. Eight of these are newly synthetized; all are of the type ROOCCH(SH)CH(SH)COOH, were R=Me, MMDMS; R = C₂H₅, MEDMS; R = (CH₂)₂CH₃, Mn-PDMS; R = CHMe₂) Mi-PDMS; R = (CH₂)₃CH₃, Mn-BDMS; R = CH₂CHMe₂, Mi-BDMS; R = (CH₂)₄CH₃, Mn-ADMS; R=(CH₂)₂CHMe₂, Mi-ADMS; and R = (CH₂)₅CH₃, Mn-HDMS. All are soluble in dilute sodium bicarbonate solutions and can be administered as aqueous solutions. Cadmium mobilization data were collected on each compound using mice previously loaded with cadmium; the monoesters were administered at a level of 0.40 mmol/kg intraperitoneally daily for five days. Data on whole body cadmium mobilization indicated that the monoester with the isoamyl group was the most effective under the conditions used. The relative whole body cadmium mobilization increased with the number of carbon atoms in the alkyl group of the monoester up to C₅ and then decreased for the C₆ compound. Cadmium removal from the kidneys and liver was also measured. It was found that the monoisoamyl ester was the most effective in removing cadmium from both the liver and the kidneys. The monoisoamyl ester also proved to be very effective in mobilizing cadmium from both the liver and the kidneys when given orally. This is the first compound which is reported capable of mobilizing cadmium in vivo from aged deposits after oral administration.     

Efficient Fmoc/solid-phase synthesis of Abu(P)-containing peptides using Fmoc-Abu(PO3Me2)-OH

The synthesis of the two 4-phosphono-2-aminobutanoyl-containing peptides, Leu-Arg-Arg-Val-Abu(P)-Leu-Gly-OH.CF₃CO₂H and Ile-Val-Pro-Asn-Abu(P)-Val-Glu-Glu-OH.CF₃CO₂H was accomplished by the use of Fmoc-Abu(PO₃Me₂)-OH in Fmoc solid-phase peptide synthesis. The protected phosphoamino acid, Fmoc-Abu(PO₃Me₂)-OH, was prepared from Boc-Asp-O'Bu in seven steps, the formation of the C—P linkage being effected by the treatment of Boc-Asa-O'Bu with dimethyl trimethylsilyl phosphite. Peptide synthesis was performed using Wang Resin as the polymer support with both peptides assembled by the use of PyBOP® for the coupling of Fmoc amino acids and 20%, piperidine for cleavage of the Fmoc group from the Fmoc-peptide after each coupling cycle. Cleavage of the peptide from the resin and peptide deprotection was accomplished by the treatment of the peptide-resin with 5%, thioanisole/TFA followed by cleavage of the methyl phosphonate group by 1 M bromotrimethylsilane/l M thioanisole in TFA.     

Platelet Aggregation Inhibiting and Anticoagulant Effects of Oligoamines,Part 33 From Tetradecane- to Octacosanediamines

Twenty alkanediamines were designed according to structure-activity relationships drawn from previous parts of this series and synthesized. Their general structure is CH₃-(CH₂)n-CHNH₂-(CH₂)_m-CHNH₂-(CH₂)_n-CH₃, (n = 2–10; m = 3–6). Twelve of them inhibited the aggregation of human blood platelets in concentrations between 3–10 μmol/L halfmaximally (Born test, inducer collagen). With increasing m a decreasing n is necessary to achieve the optimum activity. In the most active compounds ( 7b, 7e, 7p ) it is found that m + n = 9. When the nitrogen functions are hydroxyalkylated secondary amines with similar antiplatelet effects are obtained. The conversion of the amino groups into sydnonimines is accompanied by the loss of activity. The bisethoxycarbonylderivatives of 7f and 7m ( 8f, 8m ) exhibited antithrombotic effects in rats after oral administration.     

Mixed carboxylic-carbonic acid anhydrides of acylamino acids and peptides as a convenient source of 2,4-dialkyl-5(4H)-oxazolones

Racemic 2-R¹-4-R²-5(4H)-oxazolones (R¹= CH₃, C₆H₅, C₆H₅ CH₂ OCONHCH₂; R²= aliphatic and C₆H₅CH₂) have been prepared in good yields by reaction of the parent N-acetyl-, N-benzoyl-, and N-benzyloxycarbonylglycyl-amino acids with methyl chloroformate in cold dichloromethane in the presence of N-methylmorpholine, followed by washing the mixtures with aqueous solutions and removal of the solvent. The products derived from optically active substrates contained enantiometric excesses of 83–98%. The method cannot be applied to the preparation of oxazolones from N-formylamino acids.     

Protection of Guinea Pigs against Soman Poisoning by Pretreatment withp-Nitrophenyl Phosphoramidates

Severalp-nitrophenyl phosphoramidates with the general formula RO(NH₂)P(O)OC₆H₄-p-NO₂, in which R = CH₂CH₃,CH₂CH₂F, CH₂CHF₂, CH₂CF₃, CH₂CH₂CH₂CH₃, and CH₂CH₂Cl, as well as (NH₂)₂P(O)OC₆H₄-p-NO₂were administered intravenously to guinea pigs as pretreatment compounds for protection against the lethal effects of 1,2,2-trimethylpropyl methylphosphonofluoridate (soman) poisoning. Administration of phosphoramidates at a dose that produces 30% acetylcholinesterase (AChE) inhibition in the blood of atropinized guinea pigs at the moment of soman poisoning increases the subcutaneous LD50 of soman up to almost fivefold depending on which compound was used. A synergistic effect with atropine was observed. Three of these compounds offered a higher degree of protection against soman poisoning than pyridostigmine. Furthermore, the surviving animals pretreated with the two phosphoramidates that provided the highest protective ratio were in a better condition at 24 hr after soman intoxication than those pretreated with pyridostigmine. Due to the limited number of compounds and their different characteristics, it appeared difficult to demonstrate a relationship between the rate of spontaneous reactivation of AChE inhibited by the pretreatment compound and the protection against soman. Nevertheless, the results indicate that a several-fold decrease in the rate of spontaneous reactivation relative to that of pyridostigmine may increase the protective ratio against soman.     

Synthesis and biological activities of linear and cyclic enkephalin analogues containing a Ψ (E,CH=CH) or Ψ (CH2CH2) isosteric replacement

The peptide CO–NH function was replaced by a trans carbon-carbon double bond or by a CH₂–CH₂ isostere in enkephalin analogues of DADLE, DCDCE–NH₂ or DPDPE. In DADLE the 2-3 and the 3-4 peptide bond was modified, whereas in the cyclic analogues the Gly³-Phe⁴ bond was replaced by the isosteres GlyΨ (E,CH=CH)Phe [5-amino-2-(phenylmethyl)-3(E)-pentenoic acid] or GlyΨ(CH₂CH₂)Phe [5-amino-2-(phenylmethyl)pentanoic acid]. In general, the modification results in a drop in potency which is the largest for the flexible CH₂–CH₂ replacement. The Gly³Ψ (E,CH=CH)Phe⁴ DCDCE–NH₂ analogue retains considerable potency. These results confirm the importance of the peptide function at the 2-3 and 3-4 position in enkephalin analogues for biological potency.