Adaptive irrigation management by Balinese farmers reduces greenhouse gas emissions and increases rice yields.

The potential for changes in water management regimes to reduce greenhouse gases (GHG) in rice paddies has recently become a major topic of research in Asia, with implications for top-down versus bottom-up management strategies. Flooded rice paddies are a major source of anthropogenic GHG emissions and are responsible for approximately 11% of global anthropogenic methane (CH4) emissions. However, rice is also the most important food crop for people in low- and lower-middle-income countries. While CH4 emissions can be reduced by lessening the time the plants are submerged, this can trigger increased emissions of nitrous oxide (N2O), a more potent GHG. Mitigation options for CH4 and N2O are different, and minimizing one gas may increase the emission of the other. Accurate measurement of these gas emissions in rice paddies is difficult, and the results are controversial. We analysed these trade-offs using continuous high-precision measurements in a closed chamber in 2018-2020. Based on the results, we tested a bottom-up adaptive irrigation regime that improves nitrogen uptake by rice plants while reducing combined GHG emissions and nitrogen runoff from paddies to reefs in agricultural drainages. In 2023, we undertook a follow-up study in which farmers obtained higher rice yields with adaptive intermittent irrigation compared to uniformly flooded fields. These results use the polycentric, self-governing capacity of Balinese subaks for continuous adaptation. This article is part of the theme issue 'Climate change adaptation needs a science of culture'.

Modelling Singapore COVID-19 pandemic with a SEIR multiplex network model.

In this paper, we have implemented a large-scale agent-based model to study the outbreak of coronavirus infectious diseases (COVID-19) in Singapore, taking into account complex human interaction pattern. In particular, the concept of multiplex network is utilized to differentiate between social interactions that happen in households and workplaces. In addition, weak interactions among crowds, transient interactions within social gatherings, and dense human contact between foreign workers in dormitories are also taken into consideration. Such a categorization in terms of a multiplex of social network connections together with the Susceptible-Exposed-Infectious-Removed (SEIR) epidemic model have enabled a more precise study of the feasibility and efficacy of control measures such as social distancing, work from home, and lockdown, at different moments and stages of the pandemics. Using this model, we study an epidemic outbreak that occurs within densely populated residential areas in Singapore. Our simulations show that residents in densely populated areas could be infected easily, even though they constitute a very small fraction of the whole population. Once infection begins in these areas, disease spreading is uncontrollable if appropriate control measures are not implemented.

Social Cooperation and Disharmony in Communities Mediated through Common Pool Resource Exploitation.

It was theorized that when a society exploits a shared resource, the system can undergo extreme phase transition from full cooperation in abiding by a social agreement, to full defection from it. This was shown to happen in an integrated society with complex social relationships. However, real-world agents tend to segregate into communities whose interactions contain features of the associated community structure. We found that such social segregation softens the abrupt extreme transition through the emergence of multiple intermediate phases composed of communities of cooperators and defectors. Phase transitions thus now occur through these intermediate phases which avert the instantaneous collapse of social cooperation within a society. While this is beneficial to society, it nonetheless costs society in two ways. First, the return to full cooperation from full defection at the phase transition is no longer immediate. Community linkages have rendered greater societal inertia such that the switch back is now typically stepwise rather than a single change. Second, there is a drastic increase in social disharmony within the society due to the greater tension in the relationship between segregated communities of defectors and cooperators. Intriguingly, these results on multiple phases with its associated phenomenon of social disharmony are found to characterize the level of cooperation within a society of Balinese farmers who exploit water for rice production.

Radiotherapy with volumetric modulated arc therapy for hepatocellular carcinoma patients ineligible for surgery or ablative treatments.

PURPOSE: The aim of this article is to report the dosimetric and clinical findings in the treatment of primary hepatocellular carcinoma (HCC) with volumetric modulated arc therapy (VMAT, RapidArc). METHODS AND MATERIALS: A total of 138 patients were investigated. Dose prescription ranged from 45-66 Gy. Most patients (88.4 %) presented AJCC stage III or IV and 83 % were N0-M0. All were classified as Barcelona Clinic Liver Cancer (BCLC) stage A-C. All patients were treated using 10 MV photons with single or multiple, coplanar or non-coplanar arcs, and cone-down technique in case of early response of tumors. RESULTS: The patients' median age was 66 years (range 27-87 years), 83 % were treated with 60 Gy (12 % at 45 Gy, 6 % at 66 Gy), 62 % with cone-down, 98 % with multiple arcs. The mean initial planning target volume (PTV) was 777 ± 632 cm(3); the mean final PTV (after the cone-down) was 583 ± 548 cm(3). High target coverage was achieved. The final PTV was V98% > 98 %. Kidneys received on average 5 and 8 Gy (left and right), while the maximum dose to the spinal cord was 22 Gy; mean doses to esophagus and stomach were 23 Gy and 15 Gy, respectively. The average volume of healthy liver receiving more than 30 Gy was 294 ± 145 cm(3). Overall survival at 12 months was 45 %; median survival was 10.3 months (95 % confidence interval 7.2-13.3 months). Actuarial local control at 6 months was 95 % and 93.7 % at 12 months. The median follow-up was 9 months and a maximum of 28 months. CONCLUSION: This study showed from the dosimetric point of view the feasibility and technical appropriateness of RapidArc for the treatment of HCC. Clinical results were positive and might suggest, with appropriate care, to consider RapidArc as an additional therapeutic opportunity for these patients.

Antinociceptive effects of two deltorphins analogs in the tail-immersion test in rats.

The antinociceptive effects of analogs of deltorphins: cyclo(Nδ,Nδ-carbonyl-D-Orn2, Orn4)deltorphin (DEL-6) and deltorphin II N-(ureidoethyl)amide (DK-4) after intracerebroventricular (i.c.v.) administration were investigated in the tail-immersion test in rats. Morphine, the most commonly used µ-opioid receptors (MOR) agonist, was employed as a reference compound. The contribution of the MOR, δ-(DOR) and κ-opioid receptors (KOR) in antinociceptive effects of the deltorphins analogs was studies using selective antagonists of these receptors. The results indicated that DK-4 (5, 10 and 20 nmol) and DEL-6 (5, 10 and 20 nmol) were the most effective in alleviating thermal pain at the dose of 20 nmol. The antinociceptive potency of DEL-6 at the dose of 20 nmol was approximately equal but DK-4 at the dose of 20 nmol was less effective than morphine at the dose of 13 nmol. DOR antagonist - naltrindole (NTI, 5 nmol) very strongly and, to the lower extent MOR antagonist - ß-funaltrexamine (ß-FNA, 5 nmol), inhibited antinociceptive effect of DK-4 (20 nmol). In turn, ß-FNA was more potent than NTI in inhibition of the antinociceptive effects of DEL-6. Co-administration of DEL-6 and morphine at doses of 5 nmol, which do not produce measurable antinociception, generated additive antinociceptive effect. Chronic intraperitoneal (i.p.) injection of morphine (9 days) displayed a marked analgesic tolerance to the challenge dose of morphine and a slight cross-tolerance to challenge doses of DEL-6 and DK-4, given i.c.v. These findings indicate that the new deltorphin analogs recruit DOR and MOR to attenuate the nociceptive response to acute thermal stimuli.

Network extreme eigenvalue: from mutimodal to scale-free networks.

The extreme eigenvalues of adjacency matrices are important indicators on the influence of topological structures to the collective dynamical behavior of complex networks. Recent findings on the ensemble averageability of the extreme eigenvalue have further authenticated its applicability to the study of network dynamics. However, the ensemble average of extreme eigenvalue has only been solved analytically up to the second order correction. Here, we determine the ensemble average of the extreme eigenvalue and characterize its deviation across the ensemble through the discrete form of random scale-free network. Remarkably, the analytical approximation derived from the discrete form shows significant improvement over previous results, which implies a more accurate prediction of the epidemic threshold. In addition, we show that bimodal networks, which are more robust against both random and targeted removal of nodes, are more vulnerable to the spreading of diseases.

Dependence of entanglement dynamics on the global classical dynamical regime.

We analyze the connections between the dynamical generation of continuous variable entanglement and the underlying classical trajectories in pairs of coupled oscillators. In the quantization of a periodic cycle, we find periodic entanglement which has twice the frequency of the corresponding classical motion. Such frequency doubling continues to hold true in the entanglement dynamics for a second model that exhibits a two-frequency orbit in the classical domain. In addition, the periodicity and the quasiperiodicity of the entanglement are found to be independent of the local classical dynamical behavior. Finally, in our third model, the entanglement production rate is found to be (i) higher in the chaotic regime and (ii) insensitive toward the choice of regular or chaotic initial condition in the mixed regime. In summary, we have illustrated through our sample models that the generation of dynamical pattern of entanglement can depend completely on the global classical dynamical regime without being influenced by the local classical behavior.

Synthesis and biological evaluation of constrained analogues of the opioid peptide H-Tyr-D-Ala-Phe-Gly-NH2 using the 4-amino-2-benzazepin-3-one scaffold.

The synthesis of conformationally restricted dipeptidic moieties 4-amino-1,2,4,5-tetrahydro-2-benzazepin-3-one (Aba)-Gly ([(4S)-amino-3-oxo-1,2,4,5-tetrahydro-1H-2-benzazepin-2-yl]-acetic acid) and 8-hydroxy-4-amino-1,2,4,5-tetrahydro-2-benzazepin-3-one (Hba)-D-Ala ([(4S)-amino-8-hydroxy-3-oxo-1,2,4,5-tetrahydro-benzo[c]azepin-2-yl]-propionic acid) was based on a synthetic strategy that uses an oxazolidinone as an N-acyliminium precursor. Introducing these Aba scaffolds into the N-terminal tetrapeptide of dermorphin (H-Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH2)-induced remarkable shifts in affinity and selectivity towards the opioid mu- and delta-receptors. This paper provides the synthesis and biological in vitro and in vivo evaluation of constricted analogues of the N-terminal tetrapeptide H-Tyr-D-Ala-Phe-Gly-NH2, which is the minimal subunit of dermorphin needed for dermorphin-like opiate activity.

Novel endomorphin-2 analogs with mu-opioid receptor antagonist activity.

A series of position 4-substituted endomorphin-2 (Tyr-Pro-Phe-Phe-NH2) analogs containing 3-(1-naphthyl)-alanine (1-Nal) or 3-(2-naphthyl)-alanine (2-Nal) in L- or D-configuration, was synthesized. The opioid activity profiles of these peptides were determined in the mu-opioid receptor representative binding assay and in the Guinea-Pig Ileum assay/Mouse Vas Deferens assay (GPI/MVD) bioassays in vitro, as well as in the mouse hot-plate test of analgesia in vivo. In the binding assay the affinity of all new analogs for the mu-opioid receptor was reduced compared with endomorphin-2. The two most potent analogs were [D-1-Nal(4)]- and [D-2-Nal4]endomorphin-2, with IC50 values 14 +/- 1.25 and 19 +/- 2.1 nM, respectively, compared with 1.9 +/- 0.21 nM for endomorphin-2. In the GPI assay these analogs were found to be weak antagonists and they were inactive in the MVD assay. The in vitro GPI assay results were in agreement with those obtained in the in vivo hot-plate test. Antinociception induced by endomorphin-2 was reversed by concomitant intracerebroventricula (i.c.v.) administration of [D-1-Nal4]- and [D-2-Nal4]-endomorphin-2, indicating that these analogs were mu-opioid antagonists. Their antagonist activity was compared with that of naloxone. At a dose 5 microg per animal naloxone almost completely inhibited antinociceptive action of endomorphin-2, while [D-1-Nal4]endomorphin-2 in about 46%.

Type and location of fluorescent probes incorporated into the potent mu-opioid peptide [Dmt]DALDA affect potency, receptor selectivity and intrinsic efficacy.

The dermorphin-derived tetrapeptide H-Dmt-d-Arg-Phe-Lys-NH(2) (Dmt = 2',6'-dimethyltyrosine) ([Dmt(1)]DALDA) is a highly potent and selective mu-opioid agonist capable of crossing the blood-brain barrier and producing a potent, centrally mediated analgesic effect when given systemically. For the purpose of biodistribution studies by fluorescence techniques, [Dmt(1)]DALDA analogues containing various fluorescent labels [dansyl, anthraniloyl (atn), fluorescein, or 6-dimethylamino-2'-naphthoyl] in several different locations of the peptide were synthesized and characterized in vitro in the guinea-pig ileum and mouse vas deferens assays, and in mu-, delta- and kappa-opioid receptor-binding assays. The analogues showed various degrees of mu receptor-binding selectivity, but all of them were less mu-selective than the [Dmt(1)]DALDA parent peptide. Most analogues retained potent, full mu-agonist activity, except for one with fluorescein attached at the C-terminus (3a) (partial mu-agonist) and one containing beta-(6'-dimethylamino-2'-naphthoyl)alanine (aladan) in place of Phe(3) (4) (mu- and kappa-antagonist). The obtained data indicate that the receptor-binding affinity, receptor selectivity and intrinsic efficacy of the prepared analogues vary very significantly, depending on the type of fluorescent label used and on its location in the peptide. The results suggest that the biological activity profile of fluorescence-labeled peptide analogues should always be carefully determined prior to their use in biodistribution studies or other studies. One of the analogues containing the atn group (2a) proved highly useful in a study of cellular uptake and intracellular distribution by confocal laser scanning microscopy.

Cyclic enkephalin analogs containing various para-substituted phenylalanine derivatives in place of Tyr1 are potent opioid agonists.

The cyclic enkephalin analog H-Tyr-c[D-Cys-Gly-Phe(pNO(2))-D-Cys]NH(2) is a highly potent opioid agonist with IC(50)s of 35 pm and 19 pm in the guinea-pig ileum (GPI) and mouse vas deferens (MVD) assays, respectively. The Phe(1)-analog of this peptide showed 370-fold and 6790-fold lower agonist potency in the GPI and MVD assays, respectively, indicating the importance of the Tyr(1) hydroxyl-group in the interaction with mu and delta opioid receptors. In the present study, the effect of various substituents (-NH(2), -NO(2), -CN, -CH(3), -COOH, -COCH(3), -CONH(2)) introduced in the para-position of the Phe(1)-residue of H-Phe-c[D-Cys-Gly-Phe(pNO(2))-D-Cys]NH(2) on the in vitro opioid activity profile was examined. Most analogs showed enhanced mu and delta agonist potencies in the two bioassays, except for the Phe(pCOOH)(1)-analog, which was weakly active, probably as a consequence of the negative charge. The most potent compounds were the Phe(pCOH(3))(1)- and the Phe(pCONH(2))(1)-analogs. The latter compound showed subnanomolar mu and delta agonist potencies and represents the most potent enkephalin analog lacking the Tyr(1) hydroxyl-group reported to date. Taken together, these results indicate that various substituents introduced in the para-position of Phe(1) enhance opioid activity via hydrogen bonding or hydrophobic interactions with the receptor. Comparison with existing structure-activity relationship on phenolic hydroxyl replacements in morphinans indicates that these nonpeptide opiates and some of the cyclic enkephalin analogs described here may have different modes of binding to the receptor.

A chimeric opioid peptide with mixed mu agonist/delta antagonist properties.

There is evidence to indicate that opioid compounds with mixed mu agonist/delta antagonist properties are analgesics with low propensity to produce tolerance and physical dependence. A chimeric peptide containing the potent and selective mu agonist H-Dmt-D-Arg-Phe-Lys-NH2 ([Dmt1]DALDA) (Dmt=2',6'-dimethyltyrosine) and the potent and selective delta antagonist H-Tyr-TicPsi[CH2-NH]Cha-Phe-OH (TICP[Psi]) (Cha=cyclohexylalanine), connected 'tail-to-tail' via a short linker, was synthesized using a combination of solid-phase and solution techniques. The resulting peptide, H-Dmt-->D-Arg-->Phe-->Lys-NH-CH2-CH2-NH-Phe<--Cha[NH-CH2]PsiTic<--Tyr-H, showed the expected mu agonist/delta antagonist profile in the guinea-pig ileum and mouse vas deferens assays. Its mu and delta receptor binding affinities were in the low nanomolar range, as determined in rat brain membrane binding assays.

Induction chemotherapy with mitomycin, epirubicin, cisplatin, fluorouracil, and leucovorin followed by radiotherapy in the treatment of locoregionally advanced nasopharyngeal carcinoma.

PURPOSE: Survival in advanced nasopharyngeal carcinoma (NPC) is compromised by distant metastasis. Because mitomycin is active against hypoxic and G0 cells, which may help to eradicate micrometastasis, we investigated the effect of mitomycin-containing cisplatin-based induction chemotherapy. PATIENTS AND METHODS: Recruited for this study were American Joint Committee on Cancer (AJCC) 1992 staging system stage IV NPC patients with the following adverse features: obvious intracranial invasion, supraclavicular or bilateral neck lymph node metastasis, large neck node (> 6 cm), or elevated serum lactate dehydrogenase (LDH) level. Patients were given three cycles of chemotherapy before radiotherapy. The chemotherapy comprised a 3-week cycle of mitomycin, epirubicin, and cisplatin on day 1 and fluorouracil and leucovorin on day 8 (MEPFL). RESULTS: From January 1994 to December 1997, 111 patients were recruited. The median follow-up period was 43 months. The actuarial 5-year overall survival rate was 70% (95% confidence interval [CI], 60% to 80%; n = 111). For patients having completed radiotherapy (n = 100), the 5-year locoregional control rate was 70% (95% CI, 55% to 84%) and the distant metastasis-free rate was 81% (95% CI, 73% to 89%). The 5-year distant metastasis-free rate of N3a and N3b disease of AJCC 1997 staging system were 79% (95% CI, 62% to 95%) and 74% (95% CI, 60% to 89%), respectively. By Cox multivariate analysis, high pretreatment serum LDH level (P = .04) and neck nodal enlargement before radiotherapy (P = .001) were adverse prognostic factors of survival. CONCLUSION: The good 5-year survival of N3 disease supports the effectiveness of induction MEPFL in the primary treatment of advanced NPC. Further investigation to incorporate concurrent chemoradiotherapy is warranted.

Pseudoproline-containing analogues of morphiceptin and endomorphin-2: evidence for a cis Tyr-Pro amide bond in the bioactive conformation.

Analogues of the opioid peptides [D-Phe(3)]morphiceptin (H-Tyr-Pro-D-Phe-Pro-NH(2)) and endomorphin-2 (H-Tyr-Pro-Phe-Phe-NH(2)) containing the pseudoproline (Psi Pro) (4R)-thiazolidine-4-carboxylic acid (Cys[Psi(R1,R2)pro]) or (4S)-oxazolidine-4-carboxylic acid (Ser[Psi(R1,R2)pro]) in place of Pro(2) were synthesized. The pseudoproline ring in these compounds was either unsubstituted (R(1), R(2) = H) or dimethylated (R(1), R(2) = CH(3)) at the 2-C position. 2-C-dimethylated pseudoprolines are known to be quantitative or nearly quantitative inducers of the cis conformation around the Xaa(i-1)-Xaa(i)[Psi(CH(3),CH)(3)pro)] imide bond. All dihydropseudoproline-containing analogues (R(1), R(2) = H) showed good mu opioid agonist potency in the guinea pig ileum (GPI) assay, high mu receptor binding affinity in the rat brain membrane binding assay, and, like their parent peptides, excellent mu receptor binding selectivity. (1)H NMR spectroscopic analysis of the Cys[Psi(H,H)pro](2)- and Ser[Psi(H,H)pro](2)-containing analogues in DMSO-d(6) revealed that they existed in a conformational equilibrium around the Tyr-Xaa[Psi(H,H)pro] peptide bond with cis/trans ratios of 40:60 and 45:55, respectively. The dimethylated thiazolidine- and oxazolidine-containing [D-Phe(3)]morphiceptin- and endomorphin-2 analogues (R(1), R(2) = CH(3)) all retained full mu agonist potency in the GPI assay and displayed mu receptor binding affinities in the nanomolar range and high mu receptor selectivity. As expected, no conformers of the latter analogues with a trans conformation around the Tyr-Xaa[Psi(CH(3),CH(3)pro)] imide bond were detected by (1)H NMR spectral analysis, indicating that in these compounds the cis conformation is highly predominant (>98%). These results represent the most direct evidence obtained so far to indicate that morphiceptin and endomorphin-2 have the cis conformation around the Tyr-Pro peptide bond in their bioactive conformations.

[2',6'-Dimethyltyrosine]dynorphin A(1-11)-NH2 analogues lacking an N-terminal amino group: potent and selective kappa opioid antagonists.

Recent studies showed that dermorphin and enkephalin analogues containing two methyl groups at the 2',6'-positions of the Tyr(1) aromatic ring and lacking an N-terminal amino group were moderately potent delta and mu opioid antagonists. These results indicate that a positively charged N-terminal amino group may be essential for signal transduction but not for receptor binding and suggested that its deletion in agonist opioid peptides containing an N-terminal 2',6'-dimethyltyrosine (Dmt) residue may represent a general way to convert them into antagonists. In an attempt to develop dynorphin A (Dyn A)-derived kappa opioid antagonists, we prepared analogues of [Dmt(1)]Dyn A(1-11)-NH2 (1), in which the N-terminal amino group was either omitted or replaced with a methyl group. This was achieved by replacement of Tyr(1) with 3-(2,6-dimethyl-4-hydroxyphenyl)propanoic acid (Dhp) or (2S)-2-methyl-3-(2,6-dimethyl-4-hydroxyphenyl)propanoic acid [(2S)-Mdp]. Compounds were tested in the guinea pig ileum and mouse vas deferens bioassays and in rat and guinea pig brain membrane receptor binding assays. All analogues turned out to be potent kappa antagonists against Dyn A(1-13) and the non-peptide agonist U50,488 and showed only weak mu and delta antagonist activity. The most potent and most selective kappa antagonist of the series was [(2S)-Mdp(1)]Dyn A(1-11)-NH2 (5, dynantin), which showed subnanomolar kappa antagonist potency against Dyn A(1-13) and very high kappa selectivity both in terms of its K(e) values determined against kappa, mu, and delta agonists and in terms of its ratios of kappa, mu, and delta receptor binding affinity constants. Dynantin is the first potent and selective Dyn A-derived kappa antagonist known and may complement the non-peptide kappa antagonists norbinaltorphimine and GNTI as a pharmacological tool in opioid research.

Highly potent side-chain to side-chain cyclized enkephalin analogues containing a carbonyl bridge: synthesis, biology and conformation.

Six novel cyclic enkephalin analogues have been synthesized. Cyclization of the linear peptides containing basic amino acid residues in position 2 and 5 was achieved by treatment with bis(4-nitrophenyl)carbonate. It was found that some of the compounds exibit unusually high mu-opioid activity in the guinea pig ileum (GPI) assay. The 18-membered analogue cyclo(N(epsilon),N(beta)-carbonyl-D-Lys2,Dap5)-enkephalinamide turned out to be one of the most potent mu-agonists reported so far. NMR spectra of the peptides were recorded and structural parameters were determined. The conformational space was exhaustively examined for each of them using the electrostatically driven Monte Carlo method. Each peptide was finally described as an ensemble of conformations. A model of the bioactive conformation of this class of opioid peptides was proposed.

Stereospecific synthesis of (2S)-2-methyl-3-(2',6'-dimethyl-4'-hydroxyphenyl)-propionic acid (Mdp) and its incorporation into an opioid peptide.

To examine the effect of replacing the N-terminal amino group in opioid peptides with a methyl group on biological activity, a stereospecific synthesis of the tyrosine analogue (2S)-2-methyl-3-(2',6'-dimethyl-4'-hydroxyphenyl)-propionic acid (Mdp) was performed. The enkephalin analogue (2S)-Mdp-D-Ala-Gly-Phe-Leu-NH2 turned out to be a quite potent delta opioid antagonist and a somewhat less potent mu antagonist, indicating that a positively charged N-terminal amino group is not a conditio sine qua non for the binding of opioid peptides to delta and mu receptors but may be required for signal transduction.

DALDA analogues containing alpha-hydroxymethylamino acids.

To evaluate the role of aromatic amino-acids residues, four analogues of the mu-selective opioid peptide agonist DALDA (H-Tyr-D-Arg-Phe-Lys-NH2) containing the amphiphilic, a,a-disubstituted amino acid (R)- or (S)-alpha-hydroxymethyltyrosine (HmTyr) in position 1 and (R)- or (S)-alpha-hydroxymethylphenylalanine (HmPhe) in position 3 of the peptide sequence were synthesized. Only the [(R)-HmPhe3)]DALDA analogue displayed full agonistic activity in both the guinea pig ileum and the mouse vas deferens assays and turned out to be a delta receptor-selective opioid agonist.

A novel cis-peptide bond motif inducing beta-turn type VI. The synthesis of enkephalin analogues modified with 4-aminopyroglutamic acid.

A new pathway leading to a mixture of four isomers of 4-aminopyroglutamic acid is described. Michael type addition of Z-deltaAla-OMe to enolates prepared from acylaminomalonates, followed by hydrolysis and decarboxylation give protected 4-aminopyroglutamic acid with the cis:trans ratio approximately 3:2. This mixture was incorporated into Leu-enkephalin (position 2-3). After separation of peptides it appeared that all analogues were essentially inactive in guinea pig ileum and mouse vas deferens bioassays.