Lactone Form Research Articles

Activation of fluorescence of lactone forms of rhodamine dyes by photodehydrogenation of aryl(hetaryl)pyrazolines

Aryl(hetaryl)pyrazolines are effective photogenerators of acid providing an irreversible photochemical activation of fluorescence of rhodamine group dyes. The effect of solvent on two consecutive reactions, photodehydrogenation of pyrazoline and lactone ring opening of the dye was studied. An increase in solvent polarity leads to an increase in the rate of both reactions by different degrees. The systems under study are believed to be promising for the formation of recording media for multilayer optical file type disks of ultrahigh data storage capacity with fluorescent readout.

Lovastatin lactone elicits human lung cancer cell apoptosis via a COX-2/PPARγ-dependent pathway.

Statins (3-hydroxy-3-methylglutaryl coenzyme A [HMG-CoA] reductase inhibitors) are well-established agents to treat hyperlipidemic states. Experimental and epidemiological evidence further implies an anticancer effect of these substances. This study investigates the mechanism underlying human lung cancer cell death by lovastatin and the role of the prostaglandin (PG)-synthesizing enzyme cyclooxygenase-2 (COX-2) in this process. In A549 and H358 lung carcinoma cells the lipophilic prodrug lovastatin lactone led to a concentration-dependent decrease of viability and induction of DNA fragmentation, whereas its HMG-CoA-inhibitory, ring-open acid form was inactive in this respect. Apoptotic cell death by lovastatin was accompanied by high intracellular levels of the lactone form, by upregulation of COX-2 mRNA and protein, as well as by increased formation of peroxisome proliferator-activated receptor γ (PPARγ)-activating PGD2 and 15-deoxy-Δ12,14-PGJ2. Cells were significantly less sensitive to lovastatin-induced apoptotic cell death, when the expression or activity of COX-2 was suppressed by siRNA or by the COX-2 inhibitor NS-398. Apoptosis by lovastatin was likewise reversed by the PPARγ antagonist GW9662. Fluorescence microscopy analyses revealed a lovastatin-induced cytosol-to-nucleus translocation of PPARγ that was inhibited by NS-398. Collectively, this study demonstrates COX-2 induction and subsequent COX-2-dependent activation of PPARγ as a hitherto unknown mechanism by which lovastatin lactone induces human lung cancer cell death.

New-to-nature sophorose analog: a potent inducer for gene expression in Trichoderma reesei

Controlled hydrolysis of lactonic sophorolipids from Starmerella bombicola yields a previously undescribed sophorose analog that potently induces cellulase in Trichoderma reesei Rut-C30. Acid treatment of natural sophorolipids results in a mixture of monoacetylated, deacetylated, and diacetylated sophorolipids in acidic and lactonic forms. Isolation of the active components of the mixture, followed by structure determination by MS and NMR, reveals a new chemical entity, in which the lactone ring has been opened at the C-1′ rather than at the C-4″ position of the sophorose moiety. This sophorose ester is resistant to degradation by the host and is at least 28 times more powerful an inducer than sophorose in shake-flask culture. Even at low concentrations (0.05mM), the chemically modified sophorolipid effectively induces cellulase. With further improvements, this highly enabling technology can potentially reduce the cost of enzymes produced in T. reesei and can facilitate the rapid deployment of enzyme plants to support the nascent cellulosic biofuels and biochemicals industries.

A New HPLC Method for Analysis of Natural Monacolin K in Red Yeast Rice Pharmaceutical Preparations

Monacolin K (lovastatin or mevinolin) is a naturally occurring hypocholesterolemic statin used to prevent cardiovascular diseases for those persons suffering from hypercholesterolemia, a condition characterized by very high levels of cholesterol in the blood. It is originally derived from the fungus Monascus ruber and Monascus purpureus family Monascaceae. Monacolin K is a lactone polyketide derived compound converted in the body to the active form, B-hydroxy acid of monacolin K. It acts by inhibiting the hepatic HMG-CoA Reductase enzyme. In this paper, we report a selective precise and accurate new HPLC method for analysis of both lactone form and acid form of monacolin k in red rice pharmaceutical preparations.

Self-assembly of the active lactone form of a camptothecin–phospholipid complex for sustained nuclear drug delivery

(A) Illustration of the preparation of the CPT–SPC complex and its self-assembled nanoparticles (CPT–SPC NPs). (B) Illustration of drug delivery of the self-assembled CPT–SPC NPs.

Preparation and Characterization of SN-38-Encapsulated Phytantriol Cubosomes Containing α-Monoglyceride Additives.

SN-38 is a potent active metabolite of irinotecan that has been considered as an anticancer candidate. However, the clinical development of this compound has been hampered by its poor aqueous solubility and chemical instability. In this study, we developed SN-38-encapsulated cubosomes to resolve these problems. Six α-monoglyceride additives, comprising monocaprylin, monocaprin, monolaurin, monomyristin, monopalmitin, and monostearin, were used to prepare phytantriol (PHYT) cubosomes by probe sonication. The mean particle size, polydispersity index, and zeta potential values of these systems were around 190-230 nm, 0.19-0.25 and -17 to -22 mV, respectively. Small-angle X-ray scattering analyses confirmed that the SN-38-encapsulated cubosomes existed in the Pn̄3m space group both with and without the additives. The monoglyceride additives led to around a two-fold increase in the solubility of SN-38 compared with the PHYT cubosome. The drug entrapment efficiency of PHYT cubosomes with additives was greater than 97%. The results of a stability study at 25°C showed no dramatic changes in the particle size or polydispersity index characteristics, with at least 85% of the SN-38 existing in its active lactone form after 10 d, demonstrating the high stability of the cubosome nanoparticles. Furthermore, approximately 55% of SN-38 was slowly released from the cubosomes with additives over 96 h in vitro under physiological conditions. Taken together, these results show that the SN-38-encapsulated PHYT cubosome particles are promising drug carriers that should be considered for further in vivo experiments, including drug delivery to tumor cells using the enhanced permeability and retention effect.

Synthesis of a Far‐Red Photoactivatable Silicon‐Containing Rhodamine for Super‐Resolution Microscopy

AbstractThe rhodamine system is a flexible framework for building small‐molecule fluorescent probes. Changing N‐substitution patterns and replacing the xanthene oxygen with a dimethylsilicon moiety can shift the absorption and fluorescence emission maxima of rhodamine dyes to longer wavelengths. Acylation of the rhodamine nitrogen atoms forces the molecule to adopt a nonfluorescent lactone form, providing a convenient method to make fluorogenic compounds. Herein, we take advantage of all of these structural manipulations and describe a novel photoactivatable fluorophore based on a Si‐containing analogue of Q‐rhodamine. This probe is the first example of a “caged” Si‐rhodamine, exhibits higher photon counts compared to established localization microscopy dyes, and is sufficiently red‐shifted to allow multicolor imaging. The dye is a useful label for super‐resolution imaging and constitutes a new scaffold for far‐red fluorogenic molecules.

Synthesis of a Far-Red Photoactivatable Silicon-Containing Rhodamine for Super-Resolution Microscopy.

The rhodamine system is a flexible framework for building small‐molecule fluorescent probes. Changing N‐substitution patterns and replacing the xanthene oxygen with a dimethylsilicon moiety can shift the absorption and fluorescence emission maxima of rhodamine dyes to longer wavelengths. Acylation of the rhodamine nitrogen atoms forces the molecule to adopt a nonfluorescent lactone form, providing a convenient method to make fluorogenic compounds. Herein, we take advantage of all of these structural manipulations and describe a novel photoactivatable fluorophore based on a Si‐containing analogue of Q‐rhodamine. This probe is the first example of a “caged” Si‐rhodamine, exhibits higher photon counts compared to established localization microscopy dyes, and is sufficiently red‐shifted to allow multicolor imaging. The dye is a useful label for super‐resolution imaging and constitutes a new scaffold for far‐red fluorogenic molecules.

Review article: inhibition of methanogenic archaea by statins as a targeted management strategy for constipation and related disorders.

SummaryBackgroundObservational studies show a strong association between delayed intestinal transit and the production of methane. Experimental data suggest a direct inhibitory activity of methane on the colonic and ileal smooth muscle and a possible role for methane as a gasotransmitter. Archaea are the only confirmed biological sources of methane in nature and Methanobrevibacter smithii is the predominant methanogen in the human intestine.AimTo review the biosynthesis and composition of archaeal cell membranes, archaeal methanogenesis and the mechanism of action of statins in this context.MethodsNarrative review of the literature.ResultsStatins can inhibit archaeal cell membrane biosynthesis without affecting bacterial numbers as demonstrated in livestock and humans. This opens the possibility of a therapeutic intervention that targets a specific aetiological factor of constipation while protecting the intestinal microbiome. While it is generally believed that statins inhibit methane production via their effect on cell membrane biosynthesis, mediated by inhibition of the HMG‐CoA reductase, there is accumulating evidence for an alternative or additional mechanism of action where statins inhibit methanogenesis directly. It appears that this other mechanism may predominate when the lactone form of statins, particularly lovastatin lactone, is administered.ConclusionsClinical development appears promising. A phase 2 clinical trial is currently in progress that evaluates the effect of lovastatin lactone on methanogenesis and symptoms in patients with irritable bowel syndrome with constipation. The review concludes with an outlook for the future and subsequent work that needs to be done.

Revisiting the nanoformulation design approach for effective delivery of topotecan in its stable form: an appraisal of its in vitro Behavior and tumor amelioration potential

Topotecan (TPT) is indicated against a variety of solid tumors, but has restricted clinical use owing to associated pharmaceutical caveats. This study is focused at formulating a successful TPT PLGA nanosystem which ameliorates the rapid conversion of active lactone form of drug to its inactive carboxylate form and consequently improvises its efficacy. TPT PLGA nanoparticles were formulated by a double emulsion-solvent evaporation technique with sequential optimization to obtain desired particle size, PDI, zeta potential, and entrapment efficiency. Stability of TPT was ensured by maintaining an acidic pH in the drug-containing phase and the system was evaluated for in vitro–in vivo performance including cytotoxic potency. The optimized nanosystem had a particle size of 187.33 ± 7.50 nm, a PDI of 0.179 ± 0.05, and an entrapment efficiency of 56 ± 1.2%. Low pH in the interior of nanoparticles stabilized the drug to remain in its active lactone form and revealed a biphasic release pattern till 15 d. Additionally, an in vitro cytotoxicity testing as well as in vivo antitumor efficacy demonstrated a significant potential of higher proliferation inhibition as compared with neat drug (TPT). Thus, the investigation summarized an innovative simple tool for developing stable TPT NPs for effective delivery for treating solid tumors.

Lovastatin Lactone Inhibits Methane Production in Human Stool Homogenates

Introduction: Research has linked an altered gut microbiome to symptoms of irritable bowel syndrome (IBS). In constipation-predominant IBS (IBS-C), methane production appears important as methane levels on breath are proportional to the degree of constipation, and antibiotic elimination of methane significantly improves constipation. The predominant methane-producing organism in humans is Methanobrevibacter smithii. Recent data suggests certain HMG-CoA reductase inhibitors (statins) may reduce methane production by inhibiting methanogenesis. This study examined the effects of various statins on methane production in fresh human stool homogenates. Methods: Five female subjects recruited based on high breath methane production (average=69.6ppm) provided a total of 8 fresh stool samples each. Samples were homogenized in 1XPBS (2g stool/3ml PBS) in an anaerobic chamber at 37°C and divided into 5 stoppered flasks. To assess statin effects, head gas withdrawn through a stopcock was collected at baseline, then every 30min for a total of 270min, and analyzed on a Quintron Model SC gas chromatograph for methane levels. Nine statins were initially assessed for methane inhibition at a concentration of 5mg/g stool: lovastatin lactone and hydroxyacid, pravastatin lactone and hydroxyacid, simvastatin lactone, mevastatin lactone, rosuvastatin lactone, and atorvastatin lactone and hydroxyacid. Once the ideal statin was determined, the effects of concentrations of 0.04, 0.12, 0.48, 1, 5, and 10mg/g stool were tested by sampling head gas every 90min after statin addition, up to 720min.Further assessments were then performed comparing three forms of lovastatin (lactone, diol and hydroxyacid). Results: Lovastatin lactone was identified as the only effective methane inhibitor (Figure 1), significantly inhibiting methane levels by -65% of the control stool. Lovastatin lactone at 5mg/g produced the maximum inhibiting effect resulting in an average methane level of 3% of the control over time (Figure 2). In a final validation comparison of 3 forms of lovastatin (5mg/g), both the lactone and diol forms proved to be effective. In all assessments, hydroxyacid forms were least able to inhibit methane production in fresh stool samples.Figure 1Figure 2Conclusion: Lovastatin lactone and lovastatin diol at concentrations of 5mg/g stool significantly reduce methane production in human stool homogenates. Future studies are needed to assess the effect of this statin on methane production and IBS-C symptoms in humans.

Improving Antitumor Activity with N-Trimethyl Chitosan Entrapping Camptothecin in Colon Cancer and Lung Cancer.

Camptothecin (CPT) exerts very strong antitumor activities by suppressing the activity of DNA topoisomerase I, but its application is greatly limited owing to its low solubility and the instability of the active lactone form. To overcome this bottleneck, we prepared the novel camptothecin nanocolloids based on N-trimethyl chitosan (CPT-TMC) to efficiently administer CPT systemically. In this study, we investigated the antitumor activity of CPT-TMC against both colon cancer and lung cancer. In vitro cell experiments both CPT and CPT-TMC significantly inhibited the growth of CT26 cells and LL/2 cells, but no statistical difference was observed between CPT-TMC and CPT. In vivo studies, CPT-TMC more effectively inhibited tumor growth and prolonged survival time than CPT both in the CT26 colon carcinoma subcutaneous model and in the LL/2 Lewis lung carcinoma subcutaneous model. In addition, results of PCNA and CD31 immunohistochemical staining of tumor tissues also confirmed the improved antitumor effect of CPT-TMC. These findings suggest that N-trimethyl chitosan could increase the antitumor effect of CPT. Consequently, CPT delivery by N-trimethyl chitosan is a potential approach for effective treatment of cancer.

Analysis of Chemical Equilibrium of Silicon-Substituted Fluorescein and Its Application to Develop a Scaffold for Red Fluorescent Probes

Fluorescein is a representative green fluorophore that has been widely used as a scaffold of practically useful green fluorescent probes. Here, we report synthesis and characterization of a silicon-substituted fluorescein, i.e., 2-COOH TokyoMagenta (2-COOH TM), which is a fluorescein analogue in which the O atom at the 10' position of the xanthene moiety of fluorescein is replaced with a Si atom. This fluorescein analogue forms a spirolactone ring via intramolecular nucleophilic attack of the carboxylic group in a pH-dependent manner. Consequently, 2-COOH TM exhibits characteristic large pH-dependent absorption and fluorescence spectral changes: (1) 2-COOH TM is colorless at acidic pH, whereas fluorescein retains observable absorption and fluorescence even at acidic pH, and the absorption maximum is also shifted; (2) the absorption spectral change occurs above pH 7.0 for 2-COOH TM and below pH 7.0 for fluorescein; (3) 2-COOH TM shows a much sharper pH response than fluorescein because of its pKa inversion, i.e., pKa1 > pKa2. These features are also different from those of a compound without the carboxylic group, 2-Me TokyoMagenta (2-Me TM). Analysis of the chemical equilibrium between pH 3.0 and 11.0 disclosed that 2-COOH TM favors the colorless and nonfluorescent lactone form, compared with fluorescein. Substitution of Cl atoms at the 4' and 5' positions of the xanthene moiety of 2-COOH TM to obtain 2-COOH DCTM shifted the equilibrium so that the new derivative exists predominantly in the strongly fluorescent open form at physiological pH (pH 7.4). To demonstrate the practical utility of 2-COOH DCTM as a novel scaffold for red fluorescent probes, we employed it to develop a probe for β-galactosidase.

Abstract 2294: Sophorolipid-mediated inhibition of colorectal tumor cell growth in vitro and in vivo

Abstract Background: Sophorolipids (SL), are amphiphilic biosurfactant molecules which consist of a sophorose molecule with 2 variable chain length (C10 - C22) They contain double bonds at the 3” 4” positions and fatty acids at the 1” positions. SL exist in either a lactonic (SLL; open ring) or acidic (SLA; closed ring) forms. SL is produced in crude mixtures by the yeast Candidia bombicola in economically viable amounts, with variable levels of anti-proliferative activity on tumour cell lines in vitro. As biosurfactants are well tolerated in the GI tract and currently used in a variety of food products, we tested the hypotheses that purified forms of either SLL or SLA have differential effects on colo-rectal tumour versus “normal” cells as well as in a well-established model of pre-cancerous lesions; viz the Apcmin+/− mouse. Methodology: The colo-rectal cancer cell lines HT29, HT115, HCT116, Caco-2 in addition to CCD-841 colonic epithelium and MRC5 lung fibroblasts were exposed to 10 - 100 μg/ml of either SLL or SLA (96% or 94%) pure respectively by HPLC/MS analysis) for 24 hours following serum starvation and an MTT assay performed. The mechanism of cell death in cell lines was assessed by acridine orange/ethidium bromide staining followed by microscopic examination. Five-week old APCmin+ mice or wild-type littermate mice were treated orally with 50mg/kg of either SLL or SLA, or vehicle-only control every other day for 14 weeks. Weights, water and food consumption were measured on a daily basis. On completion of the experiment, mice were euthanized, the digestive tract was excised, washed and fixed with 10% BFS. Polyp size, number and location were recorded and samples were blocked for paraffin embedding, sectioning and H&E/immunofluorescence staining and analysis. Results and Discussion: In vitro, SLL caused a decrease in colorectal cancer cell viability at >70μg/ml. However, it also caused an unfavourable effect on the colonic epithelium and lung fibroblast cell lines by reducing cell viability at a lower concentration (<10μg/ml). In-vivo, SLL treatment exacerbated the growth of polyps along the digestive tract of APCmin+ mice. SLL treatment resulted in an increase in polyp number and size compared to vehicle treated APCmin+ mice. SLL treated mice presented with reduced haematocrits (28.5% vs 34.5% p<0.03) and increased spleen weights (0.56g vs 0.70g p<0.01) compared to vehicle treated control APCmin+ mice. SLA induced a dose dependent decrease in cell viability of colo-rectal cancer cell lines (20μg/ml) without affecting the viability of the colonic epithelial and lung cell lines. SLA selectively induced cell death in the colo-rectal cancer cells by means of necrosis. However the effects of SLA in vivo are yet to be established. Contrary to current literature, purified SLA appears to have an advantage over the SLL form in vitro. To date, this is the first study investigating anti-cancer effects of sophorolipids in vivo. Citation Format: Breedge Callaghan, Sophie Roelants, Niki Baccile, Helen Lydon, Inge Van Bogaert, Ibrahim M. Banat, Roger Marchant, Christopher A. Mitchell. Sophorolipid-mediated inhibition of colorectal tumor cell growth in vitro and in vivo. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2294. doi:10.1158/1538-7445.AM2015-2294

Second-order calibration method applied to process three-way excitation–emission-kinetic fluorescence data: A novel tool for real-time quantitative analysis of the lactone hydrolysis of irinotecan in human plasma

Methods to monitor the metabolic transformations of pharmaceuticals in a real-time and rapid fashion are required to develop safe and effective drugs. Lactone hydrolysis is one of the most important metabolic transformations; it occurs readily in pharmaceuticals containing lactones by both enzymatic and non-enzymatic processes. In this study, a novel strategy that combines three-way excitation–emission-kinetic (EEM-kinetic) fluorescence data with second-order calibration method based on alternating normalization-weighted error (ANWE) algorithm was developed to monitor the lactone hydrolysis of irinotecan (CPT-11). Accurate real-time concentrations together with reasonable resolutions of the excitation and emission profiles could be estimated for both the lactone form (CPT-11-L) and carboxylate form (CPT-11-C) of CPT-11 during its hydrolysis, even when the fluorescence spectra of CPT-11-L and CPT-11-C seriously overlapped with each other and unknown fluorescent components coexisted in the plasma. Satisfactory results were obtained for static validation samples, spiked plasma samples, and kinetic samples. The average recoveries of CPT-11-L and CPT-11-C in three types of samples mentioned above ranged from 83.7% to 116.7%. The lactone hydrolysis of CPT-11 follows 1st-order degradation kinetics with rate constants of 0.0318±0.0031min−1 and 0.0323±0.0008min−1, and half-lives of 21.9±2.0min and 21.5±0.5min in PBS and plasma samples, respectively. The strategy described herein minimized or eliminated traditionally time-consuming sample pre-treatments and facilitated the quantification of the target drug in its native environment. This method was proved to be simple, real-time, fast, low-cost, high-efficiency and can be potentially applied to other types of dynamic systems.

Biological activity of sophorolipids and their possible use as antiviral agents

Sophorolipids are surface active glycolipids consisting of a hydrophilic sophorose unit and a hydrophobic portion composed of a fatty acid tail. Crude sophorolipid sample contains both acidic and lactonic forms of sophorolipid with different degrees of acetylation and varying lengths of the fatty acid chains depending on the substrates used in the production process. Carboxylic end in the acidic form of the fatty acid is free, whereas in the lactonic form, it is internally esterified. Sophorolipids show different physicochemical properties with wide range of applications for each structural compound. Lactonic form of sophorolipids shows surface tension reducing ability and biological activity, whereas the acidic form possesses better foam forming ability and higher solubility. Presence of acetyl groups gives hydrophilic nature to the sophorolipids which promotes its antiviral and cytokine-stimulating properties. The aim of this review is to explore and suggest the plausibility of sophorolipids as therapeutic and prophylactic agents for the treatment of viral diseases.

Directional Dynamic Covalent Motion of a Carbonyl Walker on a Polyamine Track

Controlled directional displacement of a molecular group has been achieved based on dynamic covalent motions implementing the reactional features of the imine bond. ortho-Carboxybenzaldehyde derivatives are able to form stable adducts with both primary and secondary amines as imines or as amino lactones, respectively, depending on the acidity of the medium. They may thus perform pH-driven intramolecular "walking" along a non-symmetric polyamine chain, in which an imine serves as the terminus under basic conditions on one end of the chain and a lactone formed on a secondary hydroxylamine nitrogen on the other end serves as the terminal site upon addition of acid. The displacement between the termini occurs stochastically through reversible change in valency at the carbon site of the carbonyl group between imine, aminal, iminium and amino lactone form. On the other hand, the directionality results from the stabilisation of the terminal products under given pH conditions. By its ability to undergo interconversion between C=N and O-C-N moieties, the ortho-carboxybenzaldehyde group extends the realm of dynamic covalent chemistry of imines to secondary amines and opens new perspectives in this field.

Accumulation of Glycoconjugates of 3-Methyl-4-hydroxyoctanoic Acid in Fruits, Leaves, and Shoots of Vitis vinifera cv. Monastrell following Foliar Applications of Oak Extract or Oak Lactone

Grapevines are capable of absorbing volatile compounds present in the vineyard during the growing season, and in some cases, volatiles have been found to accumulate in fruits or leaves in glycoconjugate forms, that is, with one or more sugar moieties attached. The presence of oak lactone in wine is usually attributable to oak maturation, but oak lactone has been detected in wines made with fruit from grapevines treated with oak extract or oak lactone. This study investigated the accumulation of glycoconjugates of 3-methyl-4-hydroxyoctanoic acid (i.e., the ring-opened form of oak lactone) in the fruits, leaves, and shoots of Monastrell grapevines following foliar application of either oak extract or oak lactone at approximately 7 days postveraison. Fruits, leaves, and shoots were collected at three different time points, including at maturity. The oak lactone content of fruit was determined by gas chromatography-mass spectrometry, with declining concentrations observed in fruit from grapevines treated with oak lactone with ripening. The concentrations of a β-d-glucopyranoside of 3-methyl-4-hydroxyoctanoic acid in fruits, leaves, and shoots was determined by liquid chromatography-tandem mass spectrometry, with the highest oak lactone glucoside levels observed in leaves of grapevines treated with oak lactone. A glucose-glucose disaccharide was also tentatively identified. These results demonstrate both ring-opening and glycosylation of oak lactone occurred after experimental treatments were imposed.

Structure-function analysis of peptide signaling in the Clostridium perfringens Agr-like quorum sensing system.

The accessory growth regulator (Agr)-like quorum sensing (QS) system of Clostridium perfringens controls the production of many toxins, including beta toxin (CPB). We previously showed (J. E. Vidal, M. Ma, J. Saputo, J. Garcia, F. A. Uzal, and B. A. McClane, Mol Microbiol 83:179-194, 2012, http://dx.doi.org/10.1111/j.1365-2958.2011.07925.x) that an 8-amino-acid, AgrD-derived peptide named 8-R upregulates CPB production by this QS system. The current study synthesized a series of small signaling peptides corresponding to sequences within the C. perfringens AgrD polypeptide to investigate the C. perfringens autoinducing peptide (AIP) structure-function relationship. When both linear and cyclic ring forms of these peptides were added to agrB null mutants of type B strain CN1795 or type C strain CN3685, the 5-amino-acid peptides, whether in a linear or ring (thiolactone or lactone) form, induced better signaling (more CPB production) than peptide 8-R for both C. perfringens strains. The 5-mer thiolactone ring peptide induced faster signaling than the 5-mer linear peptide. Strain-related variations in sensing these peptides were detected, with CN3685 sensing the synthetic peptides more strongly than CN1795. Consistent with those synthetic peptide results, Transwell coculture experiments showed that CN3685 exquisitely senses native AIP signals from other isolates (types A, B, C, and D), while CN1795 barely senses even its own AIP. Finally, a C. perfringens AgrD sequence-based peptide with a 6-amino-acid thiolactone ring interfered with CPB production by several C. perfringens strains, suggesting potential therapeutic applications. These results indicate that AIP signaling sensitivity and responsiveness vary among C. perfringens strains and suggest C. perfringens prefers a 5-mer AIP to initiate Agr signaling. Clostridium perfringens possesses an Agr-like quorum sensing (QS) system that regulates virulence, sporulation, and toxin production. The current study used synthetic peptides to identify the structure-function relationship for the signaling peptide that activates this QS system. We found that a 5-mer peptide induces optimal signaling. Unlike other Agr systems, a linear version of this peptide (in addition to thiolactone and lactone versions) could induce signaling. Two C. perfringens strains were found to vary in sensitivity to these peptides. We also found that a 6-mer peptide can inhibit toxin production by some strains, suggesting therapeutic applications.

Castor oil as secondary carbon source for production of sophorolipids using Starmerella bombicola NRRL Y-17069.

Sophorolipids (SLs), a prominent member of the biosurfactants family are produced in acidic and/or lactonic form by yeast Starmerella bombicola NRRL Y-17069 when grown on hydrophilic or hydrophobic or both carbon sources. In current study, ricinoleic acid rich castor oil (10%) was used as hydrophobic and glycerol (10%) was used as hydrophilic carbon source. The yields of 24.5 ± 0.25 g/l sophorolipids were analyzed by anthrone and HPLC method which further increased upto 40.24 ± 0.76 g/l sophorolipids using fed batch process at 5L fermenter level. The structures of sophorolipids synthesized on castor oil were elucidated by liquid chromatography-mass spectrometer (LC-MS), (13)C and (1)H NMR. The results indicated that the ricinoleic acid (RA) gets hydroxylated at ω-1 position but incorporated into sophorolipids through already available hydroxyl group at 12(th) position. It resulted in the production of a novel sophorolipids with hydroxyl fatty acid as side chain and has applications as surfactant for novel drug delivery, anti microbial agent, cosmetic ingredient and emulsifier.