03069nas a2200289 4500008004100000022001400041245020400055210006900259260010300328300001600431490000800447520193000455100001602385700001902401700003702420700001702457700001802474700002502492700001702517700001702534700001702551700001902568700001502587700002102602700002402623856013202647 2012 eng d a0021-925800aNovel anti-inflammatory activity of epoxyazadiradione against macrophage migration inhibitory factor inhibition of tautomerase and proinflammatory activities of macrophage migration inhibitory factor0 aNovel antiinflammatory activity of epoxyazadiradione against mac a9650 ROCKVILLE PIKE, BETHESDA, MD 20814-3996 USAbAMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INCcJUL a24844-248610 v2873 a

Macrophage migration inhibitory factor (MIF) is responsible for proinflammatory reactions in various infectious and noninfectious diseases. We have investigated the mechanism of anti-inflammatory activity of epoxyazadiradione, a limonoid purified from neem (Azadirachta indica) fruits, against MIF. Epoxyazadiradione inhibited the tautomerase activity of MIF of both human (huMIF) and malaria parasites (Plasmodium falciparum (PfMIF) and Plasmodium yoelii (PyMIF)) non-competitively in a reversible fashion (K-i, 2.11-5.23 mu M). Epoxyazadiradione also significantly inhibited MIF (huMIF, PyMIF, and PfMIF)-mediated proinflammatory activities in RAW 264.7 cells. It prevented MIF-induced macrophage chemotactic migration, NF-kappa B translocation to the nucleus, up-regulation of inducible nitric-oxide synthase, and nitric oxide production in RAW 264.7 cells. Epoxyazadiradione not only exhibited anti-inflammatory activity in vitro but also in vivo. We tested the anti-inflammatory activity of epoxyazadiradione in vivo after co-administering LPS and MIF in mice to mimic the disease state of sepsis or bacterial infection. Epoxyazadiradione prevented the release of proinflammatory cytokines such as IL-1 alpha, IL-1 beta, IL-6, and TNF-alpha when LPS and PyMIF were co-administered to BALB/c mice. The molecular basis of interaction of epoxyazadiradione with MIFs was explored with the help of computational chemistry tools and a biological knowledge base. Docking simulation indicated that the binding was highly specific and allosteric in nature. The well known MIF inhibitor (S, R)-3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1) inhibited huMIF but not MIF of parasitic origin. In contrast, epoxyazadiradione inhibited both huMIF and plasmodial MIF, thus bearing an immense therapeutic potential against proinflammatory reactions induced by MIF of both malaria parasites and human.

1 aAlam, Athar1 aHaldar, Saikat1 aThulasiram, Hirekodathakallu, V.1 aKumar, Rahul1 aGoyal, Manish1 aIqbal, Mohd, Shameel1 aPal, Chinmay1 aDey, Sumanta1 aBindu, Samik1 aSarkar, Souvik1 aPal, Uttam1 aMaiti, Nakul, C.1 aBandyopadhyay, Uday uhttps://library.ncl.res.in/content/novel-anti-inflammatory-activity-epoxyazadiradione-against-macrophage-migration-inhibitory-001812nas a2200169 4500008004100000022001400041245011800055210006900173260011000242300001400352490000700366520106400373100001901437700002101456700003701477856012801514 2013 eng d a1463-926200aBiocatalysis: fungi mediated novel and selective 12 beta- or 17 beta-hydroxylation on the basic limonoid skeleton0 aBiocatalysis fungi mediated novel and selective 12 beta or 17 be aTHOMAS GRAHAM HOUSE, SCIENCE PARK, MILTON RD, CAMBRIDGE CB4 0WF, CAMBS, ENGLANDbROYAL SOC CHEMISTRYcMAR a1311-13170 v153 a

Basic limonoids carrying a 4,4,8-trimethyl-17-furanylsteroid skeleton are a class of triterpenoids and well-known for their insecticidal as well as a vast array of pharmacological activities. Rare and synthetically challenging 12 beta- and 17 beta-hydroxylation was achieved on the basic limonoid skeleton to produce a novel series of hydroxylated limonoids using fungi-mediated biocatalysis. The fungal system belonging to the genera of Mucor efficiently converted azadiradione, epoxyazadiradione, gedunin and their derivatives into corresponding 12 beta- and/or 17 beta-hydroxy derivatives. The position and stereochemistry of hydroxylation was determined by rigorous spectroscopic and crystallographic studies. This fungi-mediated stereo- and regio-selective hydroxylation process was highly efficient and mild enough to sustain chemically sensitive functional groups around the basic limonoid skeleton. Modifications of specific functional groups and variation in biocatalyst were shown to bring selectivity among 12 beta- or 17 beta-hydroxylation.

1 aHaldar, Saikat1 aKolet, Swati, P.1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/biocatalysis-fungi-mediated-novel-and-selective-12-beta-or-17-beta-hydroxylation-basic-001870nas a2200229 4500008004100000022001400041245010100055210006900156260007900225300001400304490000700318520099500325653002501320653002201345653001401367100002101381700002701402700001901429700002401448700003701472856013101509 2013 eng d a0039-128X00aBiocatalyst mediated production of 6 beta,11 alpha-dihydroxy derivatives of 4-ene-3-one steroids0 aBiocatalyst mediated production of 6 beta11 alphadihydroxy deriv a360 PARK AVE SOUTH, NEW YORK, NY 10010-1710 USAbELSEVIER SCIENCE INCcNOV a1152-11580 v783 a

Biotransformation of steroids with 4-ene-3-one functionality such as progesterone (I), testosterone (II), 17 alpha-methyltestosterone (III), 4-androstene-3,17-dione (IV) and 19-nortestosterone (V) were studied by using a fungal system belonging to the genera of Mucor (M881). The fungal system efficiently and quantitatively converted these steroids in regio- and stereo-selective manner into corresponding 6 beta,11 alpha-dihydroxy compounds. Time course experiments suggested that the transformation was initiated by hydroxylation at 6 beta- or 11 alpha-(10 beta-hydroxy in case of V) to form monohydroxy derivatives which upon prolonged incubation were converted into corresponding 613,11oc-dihydroxy derivatives. The fermentation studies carried out using 5 L table-top fermentor with substrates (I and II) clearly indicates that 6 beta,11 alpha-dihydroxy derivatives of steroids with 4-ene-3-one functionality can be produced in large scale by using M881. (C) 2013 Elsevier Ltd.

10a4-Ene-3-one steroids10aBiotransformation10aMucor sp.1 aKolet, Swati, P.1 aNiloferjahan, Siddiqui1 aHaldar, Saikat1 aGonnade, Rajesh, G.1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/biocatalyst-mediated-production-6-beta11-alpha-dihydroxy-derivatives-4-ene-3-one-steroids-001193nas a2200181 4500008004100000022001400041245010300055210006900158260011000227300001400337490000600351520042200357100001900779700002400798700002100822700003700843856013100880 2013 eng d a1759-966000aExpedient preparative isolation, quantification and characterization of limonoids from neem fruits0 aExpedient preparative isolation quantification and characterizat aTHOMAS GRAHAM HOUSE, SCIENCE PARK, MILTON RD, CAMBRIDGE CB4 0WF, CAMBS, ENGLANDbROYAL SOC CHEMISTRYcAUG a5386-53910 v53 a

Major basic limonoids from Neem fruits such as azadirone, epoxyazadiradione and azadiradione were isolated in preparative scale using an automated and rapid medium pressure liquid chromatography (MPLC)-based protocol. They were individually quantified using HPLC. An LC-ESI-MS/MS-based rapid identification technique was developed for the high-throughput screening of these limonoids in phytochemical extracts.

1 aHaldar, Saikat1 aPhapale, Prasad, B.1 aKolet, Swati, P.1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/expedient-preparative-isolation-quantification-and-characterization-limonoids-neem-fruits-001600nas a2200217 4500008004100000022001400041245009400055210006900149260007100218300001600289490000700305520078500312100001901097700001901116700002101135700002301156700001801179700002101197700003701218856012701255 2013 eng d a0022-326300aOne-pot fluorescent labeling protocol for complex hydroxylated bioactive natural products0 aOnepot fluorescent labeling protocol for complex hydroxylated bi a1155 16TH ST, NW, WASHINGTON, DC 20036 USAbAMER CHEMICAL SOCcOCT a10192-102020 v783 a

Tagging of small bioactive molecules with a fluorophore is a highly sensitive method to trace their cellular activities through real-time visual information. Here we disclose a 7-nitrobenzo-2-oxa-1,3-diazole (NBD)-based, high-yielding, one-pot labeling protocol for hydroxylated molecules using Yamaguchi coupling as the key reaction. This methodology was successfully applied on several sensitive and complex hydroxylated bioactive compounds including 7-deacetylazadiradione, simvastatin, camptothecin, andrographolide, cinchonine, beta-dihydroartemisinin, and azadirachtin A. Further, utility of this protocol was illustrated on the cytotoxic activity of azadiradione derivatives against several cancer cell lines through cell imaging of two qualified fluorescent probes.

1 aHaldar, Saikat1 aKumar, Santosh1 aKolet, Swati, P.1 aPatil, Harshal, S.1 aKumar, Dhiraj1 aKundu, Gopal, C.1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/one-pot-fluorescent-labeling-protocol-complex-hydroxylated-bioactive-natural-products-001282nas a2200205 4500008004100000022001400041245008100055210006900136260011000205300001600315490000600331520047900337100001900816700002100835700002700856700002100883700002400904700003700928856011100965 2014 eng d a2046-206900aBiocatalyst mediated functionalization of salannin, an insecticidal limonoid0 aBiocatalyst mediated functionalization of salannin an insecticid aTHOMAS GRAHAM HOUSE, SCIENCE PARK, MILTON RD, CAMBRIDGE CB4 0WF, CAMBS, ENGLANDbROYAL SOC CHEMISTRYcJUN a27661-276640 v43 a

Transformation of salannin, an insecticidal C-seco limonoid was investigated using a fungal system, Cunninghamella echinulata. Salannin was efficiently converted into two metabolites, where the C-17 furan moiety was transformed into gamma-hydroxybutenolide (salanninolide) and N-(2-hydroxyethyl)-alpha,beta-unsaturated-gamma-lactam (salanninactam) analogues. Present studies have indicated salanninolide to be a metabolite in the C-seco limonoid biosynthetic pathway.

1 aHaldar, Saikat1 aKolet, Swati, P.1 aDandekar, Devdutta, S.1 aKale, Balaji, S.1 aGonnade, Rajesh, G.1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/biocatalyst-mediated-functionalization-salannin-insecticidal-limonoid-002288nas a2200265 4500008004100000022001400041245012100055210006900176260007300245300000900318490000900327520130900336653003701645653000901682653001301691653002601704653001301730653001801743100001901761700002201780700002901802700002701831700003701858856012701895 2014 eng d a0021-967300aExpedient preparative isolation and tandem mass spectrometric characterization of C-seco triterpenoids from neem oil0 aExpedient preparative isolation and tandem mass spectrometric ch aPO BOX 211, 1000 AE AMSTERDAM, NETHERLANDSbELSEVIER SCIENCE BVcOCT a1-140 v13663 a

C-seco triterpenoids are widely bioactive class of natural products with high structural complexity and diversity. The preparative isolation of these molecules with high purity is greatly desirable, although restricted due to the complexity of natural extracts. In this article we have demonstrated a Medium Pressure Liquid Chromatography (MPLC) based protocol for the isolation of eight major C-seco triterpenoids of salannin skeleton from Neem (Azadirachta indica) oil. Successive application of normal phase pre-packed silica-gel columns for the fractionation followed by reverse phase in automated MPLC system expedited the process and furnished highly pure metabolites. Furthermore, eight isolated triterpenoids along with five semi-synthesized derivatives were characterized using ultra performance liquid chromatography-electrospray ionization-quadrupole/orbitrap-MS/MS spectrometry as a rapid and sensitive identification technique. The structure-fragment relationships were established on the basis of plausible mechanistic pathway for the generation of daughter ions. The MS/MS spectral information of the triterpenoids was further utilized for the identification of studied molecules in the complex extract of stem and bark tissues from Neem. (C) 2014 Elsevier B.V. All rights reserved.

10aESI(+)-quadrupole/orbitrap-MS/MS10aMPLC10aNeem oil10aPreparative isolation10aSalannin10aTriterpenoids1 aHaldar, Saikat1 aMulani, Fayaj, A.1 aAarthy, Thiagarayaselvam1 aDandekar, Devdutta, S.1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/expedient-preparative-isolation-and-tandem-mass-spectrometric-characterization-c-seco-001705nas a2200241 4500008004100000022001400041245008800055210006900143260010600212300001400318490000700332520081800339653001101157653001901168653002301187653001101210653002201221100001901243700002101262700002301283700003701306856012001343 2014 eng d a0040-403900aLipase mediated separation of triterpene structural isomers, alpha- and beta-amyrin0 aLipase mediated separation of triterpene structural isomers alph aTHE BOULEVARD, LANGFORD LANE, KIDLINGTON, OXFORD OX5 1GB, ENGLANDbPERGAMON-ELSEVIER SCIENCE LTDcMAY a3122-31250 v553 a

Pentacyclic triterpenoids alpha- and beta-amyrin possess a wide range of biological and pharmacological activities. High structural similarity between these two structural isomers makes their chromatographic separation an ineffective and tedious choice. In this study, Candida rugosa lipase catalyzed separation protocol for the isolation of individual isomers has been developed. In the presence of vinyl acetate as the acyl donor, Candida rugosa lipase carried out acetylation of beta-amyrin more efficiently as compared to alpha-amyrin leading to a kinetic separation. The conditions of transesterification reaction were optimized systematically, which was utilized to separate alpha- and beta-amyrin from a mixture obtained from the latex of Plumeria obtusa. (C) 2014 Elsevier Ltd. All rights reserved.

10aAmyrin10aCandida rugosa10aKinetic separation10aLipase10aStructural isomer1 aHaldar, Saikat1 aKale, Balaji, S.1 aJadhav, Dipesh, D.1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/lipase-mediated-separation-triterpene-structural-isomers-alpha-and-beta-amyrin-001674nas a2200241 4500008004100000022001400041245013300055210006900188260007900257300000900336490000700345520074600352653002501098653002201123653001901145653001701164653001701181100002101198700001901219700002701238700003701265856013001302 2014 eng d a0039-128X00aMucor hiemalis mediated 14 alpha-hydroxylation on steroids: In vivo and in vitro investigations of 14 alpha-hydroxylase activity0 aMucor hiemalis mediated 14 alphahydroxylation on steroids In viv a360 PARK AVE SOUTH, NEW YORK, NY 10010-1710 USAbELSEVIER SCIENCE INCcJUL a6-120 v853 a

Transformation of testosterone and progesterone into synthetically challenging 14 alpha-hydroxy derivatives was achieved by using fungal strain Mucor hiemalis. Prolonged incubation led to the formation of corresponding 6 beta/7 alpha,14 alpha-dihydroxy metabolites. The position and stereochemistry of newly introduced hydroxyl group was determined by detailed spectroscopic analyses. The time course experiment indicated that fungal strain initiated transformation by hydroxylation at 14 alpha-position followed by at 6 beta- or 7 alpha-positions. Studies using cell-free extracts suggest that the 14 alpha-hydroxylase activity is NADPH dependent and belongs to the cytochrome P450 family. (C) 2014 Elsevier Inc. All rights reserved.

10a14 alpha-Hydroxylase10aBiotransformation10aMucor hiemalis10aProgesterone10aTestosterone1 aKolet, Swati, P.1 aHaldar, Saikat1 aNiloferjahan, Siddiqui1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/mucor-hiemalis-mediated-14-alpha-hydroxylation-steroids-vivo-and-vitro-investigations-14-002652nas a2200205 4500008004100000022001400041245013700055210006900192260009100261300001300352490000700365520181000372100002102182700001902203700002202222700002402244700003302268700002202301856012302323 2015 eng d a1932-620300aGedunin and azadiradione: human pancreatic alpha-amylase inhibiting limonoids from neem (azadirachta indica) as anti-diabetic agents0 aGedunin and azadiradione human pancreatic alphaamylase inhibitin a1160 BATTERY STREET, STE 100, SAN FRANCISCO, CA 94111 USAbPUBLIC LIBRARY SCIENCEcOCT ae01401130 v103 a

Human pancreatic alpha-amylase (HPA) inhibitors offer an effective strategy to lower postprandial hyperglycemia via control of starch breakdown. Limonoids from Azadirachta indica known for their therapeutic potential were screened for pancreatic alpha-amylase inhibition, a known anti-diabetic target. Studies were carried out to reveal their mode of action so as to justify their hypoglycemic potential. Of the nine limonoids isolated/semi-synthesized from A. indica and screened for alpha-amylase inhibition, azadiradione and exhibited potential inhibition with an IC50 value of 74.17 and 68.38 mu M, respectively against HPA under in vitro conditions. Further screening on AR42J alpha-amylase secretory cell line for cytotoxicity and bioactivity revealed that azadiradione and gedunin exhibited cytotoxicity with IC50 of 11.1 and 13.4 mu M. Maximal secreted alpha-amylase inhibition of 41.8% and 53.4% was seen at 3.5 and 3.3 mu M, respectively. Michaelis-Menten kinetics suggested a mixed mode of inhibition with maltopentaose (K-i 42.2, 18.6 mu M) and starch (K-i' 75.8, 37.4 mu M) as substrate with a stiochiometry of 1: 1 for both azadiradione and gedunin, respectively. The molecular docking simulation indicated plausible p-alkyl and alkyl-alkyl interactions between the aromatic amino acids and inhibitors. Fluorescence and CD confirmed the involvement of tryptophan and tyrosine in ligand binding to HPA. Thermodynamic parameters suggested that binding is enthalpically and entropically driven with Delta G degrees of -21.25 kJ mol(-1) and -21.16 kJ mol(-1) for azadiradione and gedunin, respectively. Thus, the limonoids azadiradione and gedunin could bind and inactivate HPA (anti-diabetic target) and may prove to be lead drug candidates to reduce/control post-prandial hyperglycemia.

1 aPonnusamy, Sudha1 aHaldar, Saikat1 aMulani, Fayaj, A.1 aZinjarde, Smita, S.1 aThulasiram, Hirekodathakallu1 aRaviKumar, Ameeta uhttps://library.ncl.res.in/content/gedunin-and-azadiradione-human-pancreatic-alpha-amylase-inhibiting-limonoids-neem-103334nas a2200277 4500008004100000022001400041245014500055210006900200260008100269300002400350490000700374520225400381653002302635653002702658653001802685653001802703100002202721700002702743700001902770700002202789700002702811700002202838700002902860700003702889856013002926 2015 eng d a1471-222900aTriterpenoid profiling and functional characterization of the initial genes involved in isoprenoid biosynthesis in neem (Azadirachta indica)0 aTriterpenoid profiling and functional characterization of the in a236 GRAYS INN RD, FLOOR 6, LONDON WC1X 8HL, ENGLANDbBIOMED CENTRAL LTDcSEP aArticle Number: 2140 v153 a

Background: Neem tree (Azadirachta indica) is one of the richest sources of skeletally diverse triterpenoids and they are well-known for their broad-spectrum pharmacological and insecticidal properties. However, the abundance of Neem triterpenoids varies among the tissues. Here, we delineate quantitative profiling of fifteen major triterpenoids across various tissues including developmental stages of kernel and pericarp, flower, leaf, stem and bark using UPLC-ESI (+)-HRMS based profiling. Transcriptome analysis was used to identify the initial genes involved in isoprenoid biosynthesis. Based on transcriptome analysis, two short-chain prenyltransferases and squalene synthase (AiSQS) were cloned and functionally characterized. Results: Quantitative profiling revealed differential abundance of both total and individual triterpenoid content across various tissues. RNA from tissues with high triterpenoid content (fruit, flower and leaf) were pooled to generate 79.08 million paired-end reads using Illumina GA.. platform. 41,140 transcripts were generated by d e novo assembly. Transcriptome annotation led to the identification of the putative genes involved in isoprenoid biosynthesis. Two short-chain prenyltransferases, geranyl diphosphate synthase (AiGDS) and farnesyl diphosphate synthase (AiFDS) and squalene synthase (AiSQS) were cloned and functionally characterized using transcriptome data. RT-PCR studies indicated five-fold and ten-fold higher relative expression level of AiSQS in fruits as compared to leaves and flowers, respectively. Conclusions: Triterpenoid profiling indicated that there is tissue specific variation in their abundance. The mature seed kernel and initial stages of pericarp were found to contain the highest amount of limonoids. Furthermore, a wide diversity of triterpenoids, especially C-seco triterpenoids were observed in kernel as compared to the other tissues. Pericarp, flower and leaf contained mainly ring-intact triterpenoids. The initial genes such as AiGDS, AiFDS and AiSQS involved in the isoprenoids biosynthesis have been functionally characterized. The expression levels of AiFDS and AiSQS were found to be in correlation with the total triterpenoid content in individual tissues.

10aAzadirachta indica10aQuantitative profiling10aTranscriptome10aTriterpenoids1 aPandreka, Avinash1 aDandekar, Devdutta, S.1 aHaldar, Saikat1 aUttara, Vairagkar1 aVijayshree, Shinde, G.1 aMulani, Fayaj, A.1 aAarthy, Thiagarayaselvam1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/triterpenoid-profiling-and-functional-characterization-initial-genes-involved-isoprenoid-101423nas a2200181 4500008004100000022001400041245010600055210006900161260007100230300001400301490000700315520069000322100001901012700002201031700002901053700003701082856012201119 2015 eng d a0022-326300aWhole-cell mediated 11 beta-hydroxylation on the basic limonoid skeleton by cunninghamella echinulata0 aWholecell mediated 11 betahydroxylation on the basic limonoid sk a1155 16TH ST, NW, WASHINGTON, DC 20036 USAbAMER CHEMICAL SOCcJUN a6490-64950 v803 a

Regio- and stereoselective 11 beta-hydroxylation was achieved on the basic limonoid skeleton through microbial transformation. Whole cells of Cunninghamella echinulata efficiently converted basic limonoids such as epoxyazadiradione, azadiradione, and gedunin to their 11 beta-hydroxy analogues as the sole metabolite. Fermentation conditions affecting the efficiency (96%) of biotransformation including substrate concentration, incubation period, pH, and temperature were optimized. The position and stereochemistry of hydroxyl functionality on the isolated metabolites were established through extensive spectroscopic and spectrometric studies (1D, 2D NMR, ESI-MS, and MS/MS).

1 aHaldar, Saikat1 aMulani, Fayaj, A.1 aAarthy, Thiagarayaselvam1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/whole-cell-mediated-11-beta-hydroxylation-basic-limonoid-skeleton-cunninghamella-002044nas a2200265 4500008004100000022001400041245011200055210006900167260010600236300001400342490000700356520101800363653003301381653002101414653003101435653002101466100002301487700002201510700001901532700001801551700002201569700003701591700001801628856013201646 2016 eng d a0960-894X00aAzadirachta indica triterpenoids promote osteoblast differentiation and mineralization in vitro and in vivo0 aAzadirachta indica triterpenoids promote osteoblast differentiat aTHE BOULEVARD, LANGFORD LANE, KIDLINGTON, OXFORD OX5 1GB, ENGLANDbPERGAMON-ELSEVIER SCIENCE LTDcAUG a3719-37240 v263 a

Terpenoids were isolated using chromatographic purification through solvent purification technique and identified as Azadirone (1), Epoxyazadiradione (2) Azadiradione (3) Gedunin (4) Nimbin (5) Salannin (6) Azadirachtin A (7) and Azadirachtin B (8) from Azadirachta indica. Out of eight compounds, only three compounds had osteogenic activity and enhanced osteoblast proliferation, differentiation and mineralization in osteoblast cells. Active compounds stimulated osteogenic genes ALP, RunX-2 and OCN expressions in vitro, but Azadirachtin A had a maximum ability to stimulate osteoblast differentiation and mineralization compared to other two active compounds. For in vivo study, Azadirachtin A injected subcutaneously in pups, which enhanced osteogenic gene expressions and promoted bone formation rate significantly. Here, we conclude that active compounds of Azadirachta indica have osteogenic activity and Azadirachtin A has a beneficial effects on bone. (C) 2016 Elsevier Ltd. All rights reserved.

10aALP and mineralization assay10aBalb/c mice pups10aCalvarial osteoblast cells10aGene expressions1 aKushwaha, Priyanka1 aKhedgikar, Vikram1 aHaldar, Saikat1 aGautam, Jyoti1 aMulani, Fayaj, A.1 aThulasiram, Hirekodathakallu, V.1 aTrivedi, Ritu uhttps://library.ncl.res.in/content/azadirachta-indica-triterpenoids-promote-osteoblast-differentiation-and-mineralization-vitro02898nas a2200313 4500008004100000022001400041245015200055210006900207260000800276300000700284490000700291520187700298653001702175653001402192653001802206653001402224653001502238653000902253100001802262700001902280700002002299700001902319700002202338700002002360700001702380700003702397700002102434856012902455 2018 eng d a1471-240700aEpoxyazadiradione suppresses breast tumor growth through mitochondrial depolarization and caspase-dependent apoptosis by targeting PI3K/Akt pathway0 aEpoxyazadiradione suppresses breast tumor growth through mitocho cJAN a520 v183 a

Background: Breast cancer is one of the most commonly diagnosed invasive cancers among women around the world. Among several subtypes, triple negative breast cancer (TNBC) is highly aggressive and chemoresistant. Treatment of TNBC patients has been challenging due to heterogeneity and devoid of well-defined molecular targets. Thus, identification of novel effective and selective agents against TNBC is essential. Methods: We used epoxyazadiradione to assess the cell viability, mitochondrial potential, ROS level, cell migration, apoptosis and protein expression in cell culture models of TNBC MDA-MB-231 and ER+MCF-7 breast cancer cells. The molecular mechanism was examined in two different type of breast cancer cells in response to epoxyazadiradione. We have also analyzed the effect of epoxyazadiradione on breast tumor growth using in vivo mice model. Results: In this study, we for the first time investigated that out of 10 major limonoids isolated from Azadirachta indica, epoxyazadiradione exhibits most potent anti-cancer activity in both TNBC and ER+breast cancer cells. Epoxyazadiradione induces apoptosis and inhibits PI3K/Akt-mediated mitochondrial potential, cell viability, migration and angiogenesis. It also inhibits the expression of pro-angiogenic and pro-metastatic genes such as Cox2, OPN, VEGF and MMP-9 in these cells. Furthermore, epoxyazadiradione attenuates PI3K/Akt- mediated AP-1 activation. Our in vivo data revealed that epoxyazadiradione suppresses breast tumor growth and angiogenesis in orthotopic NOD/SCID mice model. Conclusion: Our findings demonstrate that epoxyazadiradione inhibits PI3K/Akt-dependent mitochondrial depolarisation, induces apoptosis and attenuates cell migration, angiogenesis and breast tumor growth suggesting that this compound may act as a potent therapeutic agent for the management of breast cancer.

10aAngiogenesis10aApoptosis10aBreast cancer10aLimonoids10aMetastasis10aPI3K1 aKumar, Dhiraj1 aHaldar, Saikat1 aGorain, Mahadeo1 aKumar, Santosh1 aMulani, Fayaj, A.1 aYadav, Amit, S.1 aMiele, Lucio1 aThulasiram, Hirekodathakallu, V.1 aKundu, Gopal, C. uhttps://library.ncl.res.in/content/epoxyazadiradione-suppresses-breast-tumor-growth-through-mitochondrial-depolarization-and03475nas a2200205 4500008004100000020001400041245017500055210006900230260000800299490000700307520265300314100002902967700002202996700002203018700001803040700002703058700001903085700003703104856012803141 2018 eng d a1471-222900a Tracing the biosynthetic origin of limonoids and their functional groups through stable isotope labeling and inhibition in neem tree (Azadirachta indica) cell suspension0 aTracing the biosynthetic origin of limonoids and their functiona cOCT0 v183 aBackground Neem tree serves as a cornucopia for triterpenoids called limonoids that are of profound interest to humans due to their diverse biological activities. However, the biosynthetic pathway that plant employs for the production of limonoids remains unexplored for this wonder tree. Results Herein, we report the tracing of limonoid biosynthetic pathway through feeding experiments using C-13 isotopologues of glucose in neem cell suspension. Growth and development specific limonoid spectrum of neem seedling and time dependent limonoid biosynthetic characteristics of cell lines were established. Further to understand the role of mevalonic acid (MVA) and methylerythritol phosphate (MEP) pathways in limonoid biosynthesis, Ultra Performance Liquid Chromatography (UPLC)- tandem mass spectrometry based structure-fragment relationship developed for limonoids and their isotopologues have been utilized. Analyses of labeled limonoid extract lead to the identification of signature isoprenoid units involved in azadirachtin and other limonoid biosynthesis, which are found to be formed through mevalonate pathway. This was further confirmed by treatment of cell suspension with mevinolin, a specific inhibitor for MVA pathway, which resulted in drastic decrease in limonoid levels whereas their biosynthesis was unaffected with fosmidomycin mediated plastidial methylerythritol 4-phosphate (MEP) pathway inhibition. This was also conspicuous, as the expression level of genes encoding for the rate-limiting enzyme of MVA pathway, 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGR) was comparatively higher to that of deoxyxylulose-phosphate synthase (DXS) of MEP pathway in different tissues and also in the in vitro grown cells. Thus, this study will give a comprehensive understanding of limonoid biosynthetic pathway with differential contribution of MVA and MEP pathways. Conclusions Limonoid biosynthesis of neem tree and cell lines have been unraveled through comparative quantification of limonoids with that of neem tree and through C-13 limonoid isotopologues analysis. The undifferentiated cell lines of neem suspension produced a spectrum of C-seco limonoids, similar to parental tissue, kernel. Azadirachtin, a C-seco limonoid is produced in young tender leaves of plant whereas in the hard mature leaves of tree, ring intact limonoid nimocinol accumulates in high level. Furthermore, mevalonate pathway exclusively contributes for isoprene units of limonoids as evidenced through stable isotope labeling and no complementation of MEP pathway was observed with mevalonate pathway dysfunction, using chemical inhibitors.1 aAarthy, Thiagarayaselvam1 aMulani, Fayaj, A.1 aPandreka, Avinash1 aKumar, Ashish1 aNandikol, Sharvani, S.1 aHaldar, Saikat1 aThulasiram, Hirekodathakallu, V. uhttps://library.ncl.res.in/content/tracing-biosynthetic-origin-limonoids-and-their-functional-groups-through-stable-isotope03076nas a2200265 4500008004100000022001400041245011900055210006900174260000800243300001400251490000600265520217000271100002302441700002202464700002402486700002002510700002002530700001902550700002202569700002102591700003702612700001802649700002002667856012302687 2019 eng d a2470-134300aAzadirachtin-A from azadirachta indica impacts multiple biological targets in cotton bollworm helicoverpa armigera0 aAzadirachtinA from azadirachta indica impacts multiple biologica cMAY a9531-95410 v43 a

Azadirachtin-A (AzaA) from the Indian neem tree (Azadirachta indica) has insecticidal properties; however, its molecular mechanism remains elusive. The ``targeted and nontargeted proteomic profiling'', metabolomics, matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) imaging, gene expression, and in silico analysis provided clues about its action on Helicoverpa armigera. Fourth instar H. armigera larvae fed on AzaA-based diet (AzaD) suffered from significant mortality, growth retardation, reduced larval mass, complications in molting, and prolonged development. Furthermore, death of AzaD-fed larvae was observed with various phenotypes like bursting, blackening, and half-molting. Liquid chromatography-mass spectrometry (LC-MS) data showed limited catabolic processing of ingested AzaA and dramatic alternations of primary metabolism in H. armigera. MALDI-TOF imaging indicated the presence of AzaA in midgut of H. armigera. In the gut, out of 79 proteins identified, 34 were upregulated, which were related to digestion, immunity, energy production, and apoptosis mechanism. On the other hand, 45 proteins were downregulated, including those from carbohydrate metabolism, lipid metabolism, and energy transfer. In the hemolymph, 21 upregulated proteins were reported to be involved in immunity, RNA processing, and mRNA-directed protein synthesis, while 7 downregulated proteins were implicated in energy transfer, hydrolysis, lipid metabolism, defense mechanisms, and amino acid storage-related functions. Subsequently, six target proteins were identified using labeled AzaA that interacted with whole insect proteins. In silico analysis suggests that AzaA could be efficiently accommodated in the hydrophobic pocket of juvenile hormone esterase and showed strong interaction with active site residues, indicating plausible targets of AzaA in H. armigera. Quantitative polymerase chain reaction analysis suggested differential gene expression patterns and partly corroborated the proteomic results. Overall, data suggest that AzaA generally targets more than one protein in H. armigera and hence could be a potent biopesticide.

1 aDawkar, Vishal, V.1 aBarage, Sagar, H.1 aBarbole, Ranjit, S.1 aFatangare, Amol1 aGrimalt, Susana1 aHaldar, Saikat1 aHeckel, David, G.1 aGupta, Vidya, S.1 aThulasiram, Hirekodathakallu, V.1 aSvatos, Ales.1 aGiri, Ashok, P. uhttps://library.ncl.res.in/content/azadirachtin-azadirachta-indica-impacts-multiple-biological-targets-cotton-bollworm02116nas a2200313 4500008004100000022001400041245012800055210006900183260000800252300001600260490000800276520108500284653000801369653002001377653001301397653001901410653002101429100002301450700001801473700002001491700001801511700001901529700002201548700002501570700002401595700003301619700001801652856013201670 2019 eng d a0021-954100aEstrogen receptor activation in response to Azadirachtin A stimulates osteoblast differentiation and bone formation in mice0 aEstrogen receptor activation in response to Azadirachtin A stimu cDEC a23719-237350 v2343 a

The positive effectsof the sex hormone in sustaining bone homeostasis areexercised by maintaining the equilibrium betweencell activity and apoptosis. In this regard, the importance of estrogen receptors in maintaining the bone is that it is anattractive drug target,if devoid of known side effects. In this study, we show that a natural pure compound Azadirachtin A (Aza A) isolated from Azadirachta indica binds selectively to a site in the estrogen receptor, identifying itself to bea selective tissue modifier. Using computational and medicinal chemistry, we show that Aza A binds potentially and selectively to estrogen receptor-alpha (ER alpha) as compared with ER beta. This preferential binding of Aza A to ER alpha with good pharmacokinetic distribution in the body forms metabolites, showing that it is well absorbed. In in vivo estrogen deficiency models for osteoporosis, Aza A at a much lower dose enhances new bone formation at both sites of the trabecular and cortical bone with increased bone strength and presentswith no hyperplastic effect in the uterus.

10aALP10aGene expression10amicro-CT10amineralization10aosteoblast cells1 aKushwaha, Priyanka1 aAhmad, Naseer1 aDhar, Yogeshwar1 aVerma, Ashwni1 aHaldar, Saikat1 aMulani, Fayaj, A.1 aTrivedi, Prabodh, K.1 aMishra, Prabhat, R.1 aThulasiram, Hirekodathakallu1 aTrivedi, Ritu uhttps://library.ncl.res.in/content/estrogen-receptor-activation-response-azadirachtin-stimulates-osteoblast-differentiation-and02740nas a2200181 4500008004100000022001400041245012200055210006900177260000900246300001600255490000600271520207900277100002202356700002702378700001902405700000502424856012902429 2021 eng d a2470-134300aAccurate identification of bioactive meliaceae limonoids by UHPLC-MS/ms based structure-fragment relationships (SFRs)0 aAccurate identification of bioactive meliaceae limonoids by UHPL cOCT  a26454-264760 v63 aY Limonoids are bioactive plant specialized metabolites found in the Meliaceae family. The basic limonoids, i.e., azadiradione, epoxyazadiradione, and gedunin have been exploited for various bioactivities and therefore are the potential drug leads for tomorrow. However, their low abundance, structural similarity, and lack of adequate mass fragmentation data have hampered their accurate identification and quantification from various sources. In the present study, basic limonoids such as azadirone, azadiradione, epoxyazadiradione, and gedunin isolated from Neem were utilized for the synthesis of their derivatives and isotopologs. A total of 30 one compounds were used in this study among which five were isolated, two were biotransformed, and 24 were synthesized. Among the synthesized compounds nine are novel compounds including six deuterated analogs/isotopologs which are (1,3-H-2)-1,2dihydro-3 beta-hydroxyazadiradione (9), (1,3,16-H-2)-1,2-dihydro-3 beta-16 beta-dihydroxyazadiradione (10), 3 beta-hydroxyazadiradione (11), 3 beta-16 beta-dihydroxyazadiradione (12), (3-H-2)-3 beta-hydroxyazadiradione (13), (3,16-H-2)-3 beta-16 beta-dihydroxyazadiradione (14), (1,3,7-H-2)-1,2-dihydro-3 beta-hydroxy-7-deacetylazadiradione (15), 1,2,20,21,22,23-hexahydroazadiradione (17), and (1,3-H-2)-1,2-dihydro-3 beta-hydroxygedunin (29). These limonoids along with their semisynthesized derivatives were subjected to ultra high performance liquid chromatography mass spectrometry (UHPLC-MS/MS) and the fragmentation pathway was established based on structure-fragment relationships (SFRs), utilizing high resolution MS/MS data. We have developed a most reliable and easily reproducible protocol describing in depth analysis of SFRs based on the structural modifications and synthesis of isotopologs. Also, the MS/MS fragment library of these basic limonoids generated in this study acts as a fingerprint for accurate identification and quantification of limonoids by MS/MS analysis in various plant tissue extracts, phytopharmaceutical formulations and biological samples.1 aMulani, Fayaj, A.1 aNandikol, Sharvani, S.1 aHaldar, Saikat1 a uhttps://library.ncl.res.in/content/accurate-identification-bioactive-meliaceae-limonoids-uhplc-msms-based-structure-fragment