Energy generation from bioelectrochemical techniques: Concepts, reactor configurations and modeling approaches.

The process industries play a significant role in boosting the economy of any nation. However, poor management in several industries has been posing worrisome threats to an environment that was previously immaculate. As a result, the untreated waste and wastewater discarded by many industries contain abundant organic matter and other toxic chemicals. It is more likely that they disrupt the proper functioning of the water bodies by perturbing the sustenance of many species of flora and fauna occupying the different trophic levels. The simultaneous threats to human health and the environment, as well as the global energy problem, have encouraged a number of nations to work on the development of renewable energy sources. Hence, bioelectrochemical systems (BESs) have attracted the attention of several stakeholders throughout the world on many counts. The bioelectricity generated from BESs has been recognized as a clean fuel. Besides, this technology has advantages such as the direct conversion of substrate to electricity, and efficient operation at ambient and even low temperatures. An overview of the BESs, its important operating parameters, bioremediation of industrial waste and wastewaters, biodegradation kinetics, and artificial neural network (ANN) modeling to describe substrate removal/elimination and energy production of the BESs are discussed. When considering the potential for use in the industrial sector, certain technical issues of BES design and the principal microorganisms/biocatalysts involved in the degradation of waste are also highlighted in this review.

Electric field modulated peptide based hydrogel nanocatalysts.

The ability to modulate self-assembly is the key to manufacture application-oriented materials. In this study, we investigated the effect of three independent variables that can modulate the catalytic activity of self-assembling peptides. The first two variables, amino acid sequence and its stereochemistry, were examined for their specific roles in the epitaxial growth and hydrogelation properties of a series of catalytic tripeptides. We observed that aromatic π-π interactions that direct the self-assembly of designed peptides, and the catalytic properties of hydrogels, are governed by the position and chirality of the proline residue. Subsequently, the influence of the third variable, an external electric field, was also tested to confirm its catalytic efficiency for the asymmetric C-C bond-forming aldol reaction. In particular, the electric field treated pff and PFF gels showed 10 and 36% higher stereoselectivity, respectively, compared with the control. Structure-property analysis using CD and FTIR spectroscopy indicates the electric field-induced beta to non-beta conformational transition in the peptide secondary structure, which corroborates with its reduced cross-link density and fibril width, respectively. Amplitude sweep rheology of the gels suggests a decrease in the storage modulus, with increased field strength. The results showed that an electric field of optimal strength can modulate the physical characteristics of the hydrogel, which in turn is manifested in the observed difference in enantioselectivity.

Human intervertebral discs harbour a unique microbiome and dysbiosis determines health and disease.

BACKGROUND: To document the role of sub-clinical infections in disc disorders and investigate the existence of microbiome in intervertebral discs (IVD). METHODS: Genomic DNA from 24 lumbar IVDs [8-MRI normal discs (ND) from brain dead yet alive organ donors, 8-disc herniation (DH), 8-disc degeneration (DD)] was subjected to 16SrRNA sequencing for profiling the diversity of human disc microbiome in health and disease. The disc microbiome was further compared to established human gut and skin microbiomes. RESULTS: All healthy MRI normal discs from brain dead yet alive organ donors also had a rich bacterial presence. A total of 424 different species (355-ND, 346-DD, and 322-DH) were detected, with 42.75% OTUs being classified at kingdom level, 44% at the phylum level, 22.62% at genus level, and 5.5% at species level. Varying biodiversity and abundance between healthy and diseased discs were documented with protective bacteria being abundant in normal discs, and putative pathogens abundant in DD and DH. Propionibacterium acnes had a similar but lower abundance to other pathogens in all three groups ND (3.07%), DD (3.88%), DH (1.56%). Fifty-eight bacteria were common between gut and IVD microbiomes, 29 between skin and IVD microbiomes, and six common to gut/skin/IVD. CONCLUSION: Our study challenges the hitherto concept of sterility in healthy IVD and documented a microbiome even in MRI normal healthy discs. The varying abundance of bacteria between ND, DD, and DH documents 'dysbiosis' as a possible etiology of DD. Many known pathogens were identified in greater abundance than Propionibacterium acnes, and there was evidence for the presence of the gut/skin/spine microbiome axis.

Cellulose based nanocomposite hydrogel films consisting of sodium carboxymethylcellulose-grapefruit seed extract nanoparticles for potential wound healing applications.

The impact of grapefruit seed extract (GFSE) as an antibacterial agent on citric acid (CA) crosslinked sodium carboxymethylcellulose (NaCMC)/hydroxypropylmethylcellulose (HPMC) hydrogel films has been studied by incorporating different quantities of GFSE. The prepared films were examined for their physical, thermal, mechanical and antibacterial properties. It was observed that crystallinity and initial decomposition temperature of hydrogel films decreased with GFSE concentration. Furthermore, the swelling degree and tensile strength of hydrogel films were found to be 257.29 ± 5.08%-162.06 ± 1.78% and 11.61 ± 0.27-2.21 ± 0.94 MPa for increasing GFSE concentration varying from 0.25% - 1.5% (v/v). The presence of nanoparticles in the films was observed by FESEM and FETEM analysis. It was confirmed that the formation of nanoparticles (micelles) is due to the addition of NaCMC and GFSE, probably glycerides, which is one of the main components in GFSE. The hydrogel films have demonstrated excellent antimicrobial activity and elongation at break (%). Moreover, zeta potential of nanoparticles was recorded to be -55.26 mV ascertaining their stability in water that contributed to a higher antimicrobial activity against gram negative bacteria. All these outcomes prove the nanocomposite films to be a potential substitute for hydrogels loaded with synthetic drugs in wound healing and other biological applications.

Proteomic Signature of Nucleus Pulposus in Fetal Intervertebral Disc.

STUDY DESIGN: Profiling proteins expressed in the nucleus pulposus of fetal intervertebral disc (IVD). PURPOSE: To evaluate the molecular complexity of fetal IVDs not exposed to mechanical, traumatic, inflammatory, or infective insults to generate improved knowledge on disc homeostasis. OVERVIEW OF LITERATURE: Low back pain is the most common musculoskeletal disorder, causing a significant reduction in the quality of life, and degenerative disc disorders mainly contribute to the increasing socioeconomic burden. Despite extensive research, the causative pathomechanisms behind degenerative disc disorders are poorly understood. Precise molecular studies on the intricate biological processes involved in maintaining normal disc homeostasis are needed. METHODS: IVDs of nine fetal specimens obtained from medical abortions were used to dissect out the annulus fibrosus and nucleus pulposus under sterile operating conditions. Dissected tissues were transferred to sterile Cryovials and snap frozen in liquid nitrogen before transporting to the research laboratory for protein extraction and further liquid chromatography tandem mass spectrometry (LC-MS/ MS) analysis. Collected data were further analyzed using Gene Functional Classification Tool in DAVID and STRING databases. RESULTS: A total of 1,316 proteins were identified through LC-MS/MS analysis of nine fetal IVD tissues. Approximately 247 proteins present in at least four fetal discs were subjected to further bioinformatic analysis. The following 10 clusters of proteins were identified: collagens, ribosomal proteins, small leucine-rich proteins, matrilin and thrombospondin, annexins, protein disulfide isomerase family proteins and peroxiredoxins, tubulins, histones, hemoglobin, and prolyl 4-hydroxylase family proteins. CONCLUSIONS: This study provides fundamental information on the proteome networks involved in the growth and development of healthy fetal discs in humans. Systematic cataloging of proteins involved in various structural and regulatory processes has been performed. Proteins expressed most abundantly (collagen type XIV alpha 1 chain, biglycan, matrilin 1, and thrombospondin 1) in their respective clusters also elucidate the possibility of utilizing these proteins for potential regenerative therapies.

Inflammaging determines health and disease in lumbar discs-evidence from differing proteomic signatures of healthy, aging, and degenerating discs.

BACKGROUND CONTEXT: The true understanding of aging and disc degeneration (DD) is still elusive. MRI has not helped our attempts to understand the health and disease status of the discs as it reflects mainly the end morphologic changes and not the changes at a molecular level. Understanding degeneration at a molecular level through proteomics might allow differentiation from normal aging and also aid in the development of biomarkers for early diagnosis and preventive therapies. PURPOSE: To utilize proteomics to understand the molecular basis of healthy, aging, and degenerating discs and conclusively differentiate normal aging and degeneration. STUDY DESIGN: Proteomic analysis of human intervertebral disc samples. METHODS: L4-L5 disc samples from three groups were acquired and subjected to proteomic analysis. Samples from individuals aged in the second, third, and fourth decades were used to represent young healthy discs (Group A). Those from MRI normal donors aged in the fifth, sixth, and seventh decades represented normal aging (Group B). Five degenerated discs obtained from patients at surgery represented degeneration (Group C). The entire proteome map and alteration in protein expressions were further analyzed using bioinformatics analysis. This was a self-funded project. RESULTS: There were 84 common proteins. Specific proteins numbered 225 in A, 315 in B, and 283 in C. By gene ontology biological process identification, Group A predominated with extracellular matrix organization, cytoskeletal structural and normal metabolic proteins. Group B differed in having additional basal expression of immune response, complement inhibitors, and senescence proteins. Group C was different, with upregulation of proteins associated with oxidative stress response, positive regulators of apoptosis, innate immune response, complement activation and defense response to gram positive bacteria indicating ongoing inflammaging. CONCLUSIONS: Our study documented diverse proteome signatures between the young, aging and degenerating discs. Inflammaging was the main differentiator between normal biological aging and DD. CLINICAL SIGNIFICANCE: Multiple inflammatory molecules unique to DD were identified, allowing the possibility of developing specific biomarkers for early diagnosis and thereby provide evidence-based metrics for preventive measures rather than surgical intervention and also to monitor progress of the disease.

Implementation of artificial neural network model for continuous hydrogen production using confectionery wastewater.

In the present study, an artificial neural network (ANN) was implemented to estimate the hydrogen production from confectionery wastewater. From the experimental investigation, it could be concluded that maximum COD removal efficiency of 99% and hydrogen production rate of 6570 mL/d was achieved at 7.00 kg COD/m3d and 24 h HRT. To validate this, a back propagation ANN configuration of 4-12-4-2 was opted. The modelling was performed using the input parameters like time, influent chemical oxygen demand (COD), effluent pH and volatile fatty acids (VFA). The correlation coefficient between the experimental and predicted hydrogen production rate was 0.996. The result of the tested data for hydrogen production rate was successful. The calculated average percentage error (APE) for hydrogen production rate was 0.0004. As the APE values were closer to zero, the trained ANN model fitted well with the experimental data.

Fabrication, characterization and drug loading efficiency of citric acid crosslinked NaCMC-HPMC hydrogel films for wound healing drug delivery applications.

In this study, hydrogel films were prepared from sodium carboxymethylcellulose (NaCMC) and hydroxypropylmethylcellulose (HPMC) using citric acid (CA) as crosslinker. The chemical crosslinking (ester bond) between NaCMC and HPMC was confirmed by FTIR analysis. It was found that swelling degree, crystallinity and water contact angle of hydrogel films based on NaCMC-HPMC (2wt%) decrease with increase in CA from 5% to 20% (by weight). The increase in Tg and decrease in initial decomposition temperature were observed for increasing CA in NaCMC-HPMC films and were confirmed by DSC and TGA analysis, respectively. Tensile strength of hydrogel films decreases while elongation at break (%) increases with increase in CA. Highly interconnected nanoporous network and decrease in average pore diameter from micron to nanosize in cryofixed hydrogel films were found by FESEM and mercury intrusion porosimetry, respectively. The drug loading efficiency of hydrogel films was significantly higher for methylene blue compared to tetracycline. It was observed that hydrogel films release the drugs in sustained manner for 72h. The hydrogel films showed significant antibacterial activity after three days of release at 37°C in PBS (pH7.4). These findings strongly recommend that the prepared hydrogel films can be used as potential wound healing materials.

Microwave-assisted synthesis of cellulose/zinc-sulfate­calcium-phosphate (ZSCAP) nanocomposites for biomedical applications.

In this study, zinc-sulfate-calcium-phosphate (ZSCAP) ceramics was prepared by calcination in the presence of microwave irradiation using precursors ZnSO4:ZnO:CaO:P2O5 in a ratio of 15:30:30:25 (by weight). The calcined ZSCAP ceramics was mixed with microcrystalline cellulose and it was further heated by microwave radiation for the preparation of cellulose/ZSCAP nanocomposites. It was found that microwave heating time played an important role in the crystalline phase of synthesized nanocomposites of cellulose/ZSCAP. Well-crystalline phases of calcium phosphate, zinc oxide and zinc sulfate were observed in the nanocomposites at 20 min of microwave heating time. Fourier transformed infrared spectroscopy (FTIR) and Raman spectroscopy confirmed that the obtained products were of cellulose/ZSCAP nanocomposites. Field emission scanning electron microscopy (FESEM) and transmission electron microscopy (TEM) images indicated that reinforced ZSCAP nanostructures were embedded into cellulose matrix. Energy dispersive X-ray spectroscopy (EDS) further supported the presence of Zn, S, Ca and P in cellulose/ZSCAP nanocomposites. The thermal behavior of the products was studied using thermogravimetric analysis (TGA) and differential thermal analysis (DTA). The prepared nanocomposites showed antibacterial activity against Staphylococcus aureus and Escherichia coli. In vitro cytotoxicity assay revealed that the prepared nanocomposites had no influence on proliferation of MG-63 cells. This rapid microwave-assisted method is simple, fast and suitable for the production of cellulose/ZSCAP nanocomposites, which finds its biomedical applications in tissue engineering and bone repair.

Fabrication and characterization of chitosan, polyvinylpyrrolidone, and cellulose nanowhiskers nanocomposite films for wound healing drug delivery application.

This study describes the preparation of composite film using chitosan (CS) and polyvinylpyrrolidone (PVP) with incorporated cellulose nanowhiskers (CNWs) for drug delivery application. CNWs were prepared by acid hydrolysis of cellulose with sulfuric acid. Field emission scanning electron microscopy studies revealed nanofibrous morphology of CNWs with 20-30 nm diameter and 200-250 nm in length. X-ray powder diffraction analysis confirmed highly crystalline nature of CNWs with 92.81% crystallinity. Incorporation of CNWs enhanced the thermal and mechanical properties of films. Fourier transform infrared spectroscopy data showed physical interactions between polymer-polymer and polymer-drug. Films prepared with CNWs showed improved swelling behavior which resulted in sustained drug release from polymeric matrix. In vitro curcumin release data were fitted with two-step release model; Step 1 as desorption from the outer surface of the film, and Step 2 as diffusion from within the film and subsequent desorption. The release kinetics confirmed biphasic release profile with different release rates along with diffusion controlled curcumin release. Prepared films showed high biocompatibility with excellent antibacterial activities. Overall, the performed studies confirmed CS-PVP-CNWs based release system can as a potential candidate for wound dressing applications with sustained drug release. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2391-2404, 2017.

ISSLS PRIZE IN CLINICAL SCIENCE 2017: Is infection the possible initiator of disc disease? An insight from proteomic analysis.

STUDY DESIGN: Proteomic and 16S rDNA analysis of disc tissues obtained in vivo. OBJECTIVE: To address the controversy of infection as an aetiology for disc disorders through protein profiling. There is raging controversy over the presence of bacteria in human lumbar discs in vivo, and if they represent contamination or infection. Proteomics can provide valuable insight by identifying proteins signifying bacterial presence and, also host defence response proteins (HDRPs), which will confirm infection. METHODS: 22 discs (15-disc herniations (DH), 5-degenerate (DD), 2-normal in MRI (NM) were harvested intraoperatively and immediately snap frozen. Samples were pooled into three groups and proteins extracted were analysed with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Post identification, data analysis was performed using Uniprotdb, Pantherdb, Proteome discoverer and STRING network. Authentication for bacterial presence was performed by PCR amplification of 16S rDNA. RESULTS: LC-MS/MS analysis using Orbitrap showed 1103 proteins in DH group, compared to 394 in NM and 564 in DD. 73 bacterial specific proteins were identified (56 specific for Propionibacterium acnes; 17 for Staphylococcus epidermidis). In addition, 67 infection-specific HDRPs, unique or upregulated, such as Defensin, Lysozyme, Dermcidin, Cathepsin-G, Prolactin-Induced Protein, and Phospholipase-A2, were identified confirming presence of infection. Species-specific primers for P. acnes exhibited amplicons at 946 bp (16S rDNA) and 515 bp (Lipase) confirming presence of P. acnes in both NM discs, 11 of 15 DH discs, and all five DD discs. Bioinformatic search for protein-protein interactions (STRING) documented 169 proteins with close interactions (protein clustering co-efficient 0.7) between host response and degenerative proteins implying that infection may initiate degradation through Ubiquitin C. CONCLUSION: Our study demonstrates bacterial specific proteins and host defence proteins to infection which strengthen the hypothesis of infection as a possible initiator of disc disease. These results can lead to a paradigm shift in our understanding and management of disc disorders.

Molecular interactions in alcohol-ethyl methacrylate mixtures.

Molecular interaction between alcohols (1-propanol, 1-butanol, s-butanol, t-butanol, 1-pentanol, 1-heptanol, 1-octanol and 1-decanol) with ethyl methacrylate has been studied in n-heptane, CCl(4) and benzene at 298K using FTIR spectroscopic and dielectric methods. The result obtained from both the methods indicates only the existence of most likely 1:1 complex formation between the alcohol and ethyl methacrylate in these systems. The alkyl chain length of alcohol and the solvent used play a significant role in the strength of hydrogen bond (O-H:O=C) determined on the basis of spectral and dielectric parameters.

Hydrogen bonding interaction between acrylic esters and monohydric alcohols in non-polar solvents: An FTIR study.

The association between acrylic esters (methyl methacrylate, ethyl methacrylate and butyl methacrylate) and some monohydric (primary, secondary and tertiary) alcohols in non-polar solvents, viz. n-heptane, CCl4 and benzene has been investigated by means of FTIR spectroscopy. The most likely association complex between alcohol and acrylic ester is 1:1 stoichiometric complex through the hydroxyl group of alcohol and the carbonyl group of acrylic ester. The formation constant of the 1:1 complexes has been calculated using Nash method. It appears that the primary alcohols have larger formation constant than the secondary and tertiary alcohols. The results show that the proton donating power of the alcohols decreases in the order primary>secondary>tertiary and the association constant increases with the increase in carbon chain of the alkyl group of acrylic esters and alcohols. Also the results show a significant dependence of the association constant upon the solvents used. The solvent effect on the formation of hydrogen bond equilibria is discussed in terms of specific interaction between the solute and solvent.

Molecular interaction studies of acrylic esters with 1-alcohols.

Hydrogen bonding between 1-alcohols and acrylic esters in n-heptane has been studied by FTIR spectroscopic method. The formation constant of the 1:1 complexes has been calculated using Nash method. The values of formation constant and free energy change vary with alcohol and ester chain length, which suggests that the strengths of the intermolecular O-H...O=C bonds are shown to be dependent on the alkyl group of acrylic esters and the 1-alcohols and the results shows that the proton donating ability of 1-alcohols is in the order: 1-propanol<1-butanol<1-pentanol and the accepting ability of acrylic esters is in the order: methyl methacrylate

Mycobacterial lipoarabinomannans modulate cytokine production in human T helper cells by interfering with raft/microdomain signalling.

Lipoarabinomannans (LAMs) are major lipoglycans of the mycobacterial envelope and constitute immunodominant epitopes of mycobacteria. In this paper, we show that mannose-capped (ManLAM) and non-mannose-capped (PILAM) mycobacterial lipoglycans insert into T helper cell rafts without apparent binding to known receptors. T helper cells modified by the insertion of PILAM responded to CD3 cross-linking by decreasing type 1 (IL-2 and IFN-gamma) and increasing type 2 (IL-4 and IL-5) cytokine production. Modification by the mannose-capped ManLAMs had similar, but more limited effects on T helper cell cytokine production. When incorporated into isolated rafts, PILAMs modulated membrane-associated kinases in a dose-dependent manner, inducing increased phosphorylation of Src kinases and Cbp/PAG in Th1 rafts, while decreasing phosphorylation of the same proteins in Th2 rafts. Mycobacterial lipoglycans thus modify the signalling machineries of rafts/microdomains in T helper cells, a modification of the membrane organization that eventually leads to an overall enhancement of type 2 and inhibition of type 1 cytokine production.

Role of keratinocytes in antigen presentation and polarization of human T lymphocytes.

Allergic contact dermatitis is a T-cell-mediated inflammatory skin disease in which interaction between skin keratinocytes and migrating T lymphocytes may play a critical part. In this study, the role of keratinocytes as allergen-/antigen-presenting cells (APCs) leading to activation of T lymphocytes is investigated using a human epidermal cell line A431. It is known that cultured cells do not express human leucocyte antigen (HLA) and hence can be used as APCs independent of HLA profile of both APCs and T cells from human volunteers. This cell line responded to common allergens and irritants by inducing or upregulating the cell-surface expression of HLA-DR, and intercellular adhesion molecule-1 and B7 mRNA transcripts in keratinocytes. In addition, allergen-primed A431 cells also induced allergen-specific proliferation of human T lymphocytes in cocultures. Anti-HLA-DR, interleukin-1alpha (IL-1alpha) antibodies and lysosomotropic agent chloroquine inhibited the proliferation. Allergens also upregulated cytokines IL-1alpha, granulocyte-macrophage colony-stimulating factor, Gro-alpha and IL-12 in keratinocytes. Further, keratinocytes activated by allergens induced polarization of activated T lymphocytes to the Th1 phenotype.

A novel extracellular protein of Streptomyces peucetius binds to daunorubicin but does not inhibit the bioactivity of the antibiotic.

Extracellular proteins of Streptomyces peucetius that bind to a red pigment were identified during the course of isolation of mutants defective in daunorubicin production. Two pigment-protein complexes were partially purified and this complex inhibited the growth of Bacillus subtilis. Routine solvent extraction could not separate the pigment from purified pigment-protein complex. Treatment with 2% SDS at 100 degrees C followed by solvent extraction also failed to separate the protein from the pigment. However, the pigment could be separated from the purified pigment-protein complex by heating it in 0.1M HCl at 100 degrees C followed by solvent extraction. The pigment extracted from the complex was analyzed by HPLC and found to be daunorubicin.

Large-restriction-fragment polymorphism analysis of Mycobacterium chelonae and Mycobacterium terrae isolates.

Mycobacterium chelonae and Mycobacterium terrae were reported to be frequently present in the environment of the Mycobacterium bovis BCG trial area in south India. Six isolates of M. chelonae and four isolates of M. terrae obtained from different sources in this area were analyzed by pulsed-field gel electrophoresis (PFGE) to examine large-restriction-fragment (LRF) polymorphism using the chromosomal DNA digested with DraI and XbaI restriction enzymes. With the exception of one isolate of M. terrae, DNA from all other isolates could be digested with DraI and XbaI and resulted in separable fragments. Visual comparison of the LRFs showed a unique pattern for each of the isolates tested. A computer-assisted dendrogram of the percent similarity demonstrated a high degree of genetic diversity in this group of isolates. This study demonstrates that species of nontuberculous mycobacteria, particularly M. chelonae and M. terrae, can be successfully typed by their LRF pattern using PFGE, which does not require species-specific DNA probes.

Purification, cloning, and DNA sequence analysis of a chitinase from an overproducing mutant of Streptomyces peucetius defective in daunorubicin biosynthesis.

Extracellular chitinases of Streptomyces peucetius and a chitinase overproducing mutant, SPVI, were purified to homogeneity by ion exchange and gel filtration chromatography. The purified enzyme has a molecular mass of 42 kDa on SDS-PAGE, and the N-terminal amino acid sequence of the protein from the wild type showed homology to catalytic domains (Domain IV) of several other Streptomyces chitinases such as S. lividans 66, S. coelicolor A3(2), S. plicatus, and S. thermoviolaceus OPC-520. Purified SPVI chitinase cross-reacted to anti-chitinase antibodies of wild-type S. peucetius chitinase. A genomic library of SPVI constructed in E. coli using lambda DASH II was probed with chiC of S. lividans 66 to screen for the chitinase gene. A 2.7 kb fragment containing the chitinase gene was subcloned from a lambda DASH II clone, and sequenced. The deduced protein had a molecular mass of 68 kDa, and showed domain organization similar to that of S. lividans 66 chiC. The N-terminal amino acid sequence of the purified S. peucetius chitinase matched with the N-terminus of the catalytic domain, indicating the proteolytic processing of 68 kDa chitinase precursor protein to 42 kDa mature chitinase containing the catalytic domain only. A putative chiR sequence of a two-component regulatory system was found upstream of the chiC sequence.

Isolation and characterization of stable mutants of Streptomyces peucetius defective in daunorubicin biosynthesis.

Daunorubicin and its derivative doxorubicin are antitumour anthracycline antibiotics produced by Streptomyces peucetius. In this study we report isolation of stable mutants of S. peucetius blocked in different steps of the daunorubicin biosynthesis pathway. Mutants were screened on the basis of colony colour since producer strains are distinctively coloured on agar plates. Different mutants showed accumulation of aklaviketone, epsilon-rhodomycinone, maggiemycin or 13-dihydrocarminomycin in their culture filtrates. These results indicate that the mutations in these isolates affect steps catalysed by dnrE (mutants SPAK and SPMAG), dnrS (SPFS and SPRHO) and doxA (SPDHC) gene products.