ORIGINAL PAPER Toward the synthesis and biological evaluation of hirsutide Rajiv Dahiya Æ Monika Maheshwari Æ Anil Kumar Received: 24 July 2008 / Accepted: 30 July 2008 / Published online: 14 October 2008 Ó Springer-Verlag 2008 Abstract The present investigation deals with the synthesis of a N-methylated cyclotetrapeptide, hirsutide (2), by coupling of the dipeptide units Boc-L-phenylalanyl- L-N-methylphenylalanine-OH and L-valyl-L-N-methylphe- nylalanine-OMe followed by cyclization of the linear tetrapeptide fragment. The chemical structure was estab- lished on the basis of analytical as well as spectroscopic data. The newly synthesized cyclic peptide was subjected to pharmacological screening and found to be highly potent against the gram-negative bacteria Pseudomonas aerugin- osa and Klebsiella pneumoniae at 6 lg cm -3 . In addition, potent antihelmintic activity against the earthworms Megascoplex konkanensis and Pontoscotex corethruses at 1 and 2 mg cm -3 , and potent cytotoxic activity against Dalton’s lymphoma ascites and Ehrlich’s ascites carci- noma cell lines with IC 50 values of 14 and 22 lM were also observed. Studies revealed that the pentafluorophenyl ester method employing a catalytic amount of N-methylmor- pholine proved to be better for cyclization of the linear tetrapeptide unit. Keywords Natural products Á Hirsutide Á Peptides Á Cyclizations Á Total synthesis Á Biological activity Introduction Literature is enriched with several findings suggesting the vital role of natural products in pharmaceutical research as biomedically useful agents or as lead compounds for drug development. Among these, cyclopeptides and related congeners having unique structures and wide pharmaco- logical profiles have emerged as an important class of organic compounds, which may prove better candidates to overcome the problem of resistance towards conventional agents [1–3]. Fungi-derived natural cyclic peptides exhibit a variety of bioactivities, such as cytotoxic activity [4, 5], antimalarial activity [6], insecticidal activity [7], antibacterial activity [8], antimycotic activity [9], anti- dinoflagellate activity [10], antimycobacterial activity [11], and inhibitory activity toward the spore germination of fungi [12]. Hirsutide (2), a cyclotetrapeptide, has been isolated from the spider-derived entomopathogenic fungus Hirsutella sp. using semipreparative HPLC under isocratic conditions [13] and is unique in having two N-methylated phenylalanine units in its structure. Having only minute quantities of this cyclopeptide obtained from natural resources has restricted scientists from investigating its biological profile in detail. Further, the widespread increase of resistance against conventional antibiotics encourages the development of novel bioactive congeners with unex- ploited mechanisms of action. Hence, keeping in view the biological potential of extracts of Hirsutella sp. and to obtain a bioactive peptide in good yield, the present work was aimed at the first total synthesis of the natural peptide hirsutide (2) using the solution-phase technique in a simple and economical manner. The study also includes testing of the peptide for its antibacterial, antifungal, antihelmintic, and cytotoxic effects. R. Dahiya (&) Á M. Maheshwari Á A. Kumar Department of Pharmaceutical Chemistry, Rajiv Academy for Pharmacy, PO Chattikara, Mathura 281 001, Uttar Pradesh, India e-mail: rajivdahiya02@yahoo.com; rajivdahiya77@rediffmail.com 123 Monatsh Chem (2009) 140:121–127 DOI 10.1007/s00706-008-0052-z