Identification of potential inhibitors of the folate biosynthesis enzymes HPPK of Salmonella enterica and Escherichia coli and pteridine reductase of Trypanosoma brucei through molecular docking and enzyme assays
- Authors: Gerwel, Tiaan Marc
- Date: 2021-10-29
- Subjects: Antimicrobial resistance , African trypanosomiasis , Trypanosoma brucei , Salmonella enterica , Escherichia coli , Molecular docking
- Language: English
- Type: Master's theses , text
- Identifier: http://hdl.handle.net/10962/192419 , vital:45224
- Description: Antimicrobial resistance has become a serious threat to the survival of the human species especially those living in rural areas where access to medicine and the knowledge for proper use is scarce. It has been estimated that the number of extreme untreatable resistant infections in Africa will increase to as much as 10 million by the year 2050. Thus the need for novel drugs to act as therapeutic agents is becoming more compelling each year. The subspecies of Trypanosoma brucei (T. brucei) is responsible for the Human African Trypanosomiasis (HAT) also known as sleeping sickness and results in large numbers of deaths and loss of income for many homes. Resistance to current therapeutic agents has been observed and is on the rise increasing the difficulty to treat the infection. Salmonella enterica (S. enterica) serotypes are responsible for acute diarrhoeal disease in humans ranging from invasive typhoid to non-invasive non-typhoid disease, resulting in a large number of deaths, with an estimated 180 million people falling ill annually. The pathogen is spread via the faecal-oral route or through food contaminated with bacterium and prepared in an unsanitary environment. The chance of recovery in rural populations is low. Escherichia coli (E. coli), forming part of commensal gut flora, spread via the faecal-oral route or unhygienic practices, causes a diarrhoeal disease which can progress to a haemorrhagic phase. More than 241 million annual infections are caused by enterotoxigenic E. coli. A common strategy to develop antimicrobial agents is to target the biosynthesis of essential biological molecules, thereby rendering the microbes less viable. One such group of molecules are folates, which are generally synthesised de novo by bacteria. Higher organisms have a scavenger mechanism to obtain their folates from the extracellular environment and in some cases, organisms have both mechanisms. In this study, two enzymes falling into the folate scavenger and de novo synthesis groups were examined to identify potential agents to act as inhibitors. Pteridine reductase 1 (PTR1) is a scavenger enzyme used by a variety of trypanosomes and 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) is an enzyme that overlaps the kingdoms of protozoa and bacteria forming part of the de novo biosynthetic pathway. Homology modelling was performed on the HPPK enzyme of S. enterica using Yersinia pestis (Y. pestis) as a template providing a model to use for docking studies. The E. coli HPPK enzyme structure was retrieved from the Protein Data Bank (PDB) and bound molecules were removed to render the enzyme in an apo-state. Docking studies using the generated S. enterica homology model and apo E. coli HPPK was performed using the ZINC 15 database and resulted in 9 hit compounds which showed high binding affinities and binding energy to the enzyme. The HPPK and PTR1 enzyme coding sequences were cloned into pET-28a(+) plasmids and supplied by GenScript, to enable the expression of histidine-tagged proteins in T7 Express lysY competent E. coli cells. Analytical scale expression studies showed the recombinant proteins to be in a soluble form and purification was achieved using nickel-NTA affinity chromatography. The purified PTR1 recombinant protein was used to establish and optimise an NADPH absorbance microplate assay to screen compounds previously identified in docking studies by Kimuda, Laming, Hoppe, & Bishop, (2019). The assay yielded a Z´-factor of above 0.8 indicating an excellent assay to use for screening. An unsuccessful attempt was made to use resazurin reduction as an alternative method to demonstrate PTR1 enzyme activity. HPPK purified recombinant proteins were used to establish and optimise a luminescence microplate assay for the screening of compounds identified in in silico docking studies against the HPPK enzymes of S. enterica and E. coli. The Z´-factor of the luminescence assay was above 0.5, indicative of a functional assay with good separation between enzyme activity signal and negative control of reaction without enzyme. The target-based enzyme screening resulted in the confirmation of compound 3 (3-chloro-N-[(4-oxo-3,4-dihydrophthalazin-1-yl)methyl]benzamide) as an inhibitor of S. enterica HPPK with an IC₅₀ of 10.4 μM. At 50 μM none of the compounds decreased E. coli HPPK enzyme activity by 50%. Further bacterial studies would provide more compelling data to motivate the optimisation of compound 3 as an S. enterica inhibitor. This study demonstrated the effectiveness of using computational methods in the drug discovery process, correlating in silico results with those obtained from target-based assays producing a hit compound that can be used for future drug optimisation. , Thesis (MSc (Pharm)) -- Faculty of Pharmacy, Pharmacy, 2021
- Full Text:
- Authors: Gerwel, Tiaan Marc
- Date: 2021-10-29
- Subjects: Antimicrobial resistance , African trypanosomiasis , Trypanosoma brucei , Salmonella enterica , Escherichia coli , Molecular docking
- Language: English
- Type: Master's theses , text
- Identifier: http://hdl.handle.net/10962/192419 , vital:45224
- Description: Antimicrobial resistance has become a serious threat to the survival of the human species especially those living in rural areas where access to medicine and the knowledge for proper use is scarce. It has been estimated that the number of extreme untreatable resistant infections in Africa will increase to as much as 10 million by the year 2050. Thus the need for novel drugs to act as therapeutic agents is becoming more compelling each year. The subspecies of Trypanosoma brucei (T. brucei) is responsible for the Human African Trypanosomiasis (HAT) also known as sleeping sickness and results in large numbers of deaths and loss of income for many homes. Resistance to current therapeutic agents has been observed and is on the rise increasing the difficulty to treat the infection. Salmonella enterica (S. enterica) serotypes are responsible for acute diarrhoeal disease in humans ranging from invasive typhoid to non-invasive non-typhoid disease, resulting in a large number of deaths, with an estimated 180 million people falling ill annually. The pathogen is spread via the faecal-oral route or through food contaminated with bacterium and prepared in an unsanitary environment. The chance of recovery in rural populations is low. Escherichia coli (E. coli), forming part of commensal gut flora, spread via the faecal-oral route or unhygienic practices, causes a diarrhoeal disease which can progress to a haemorrhagic phase. More than 241 million annual infections are caused by enterotoxigenic E. coli. A common strategy to develop antimicrobial agents is to target the biosynthesis of essential biological molecules, thereby rendering the microbes less viable. One such group of molecules are folates, which are generally synthesised de novo by bacteria. Higher organisms have a scavenger mechanism to obtain their folates from the extracellular environment and in some cases, organisms have both mechanisms. In this study, two enzymes falling into the folate scavenger and de novo synthesis groups were examined to identify potential agents to act as inhibitors. Pteridine reductase 1 (PTR1) is a scavenger enzyme used by a variety of trypanosomes and 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) is an enzyme that overlaps the kingdoms of protozoa and bacteria forming part of the de novo biosynthetic pathway. Homology modelling was performed on the HPPK enzyme of S. enterica using Yersinia pestis (Y. pestis) as a template providing a model to use for docking studies. The E. coli HPPK enzyme structure was retrieved from the Protein Data Bank (PDB) and bound molecules were removed to render the enzyme in an apo-state. Docking studies using the generated S. enterica homology model and apo E. coli HPPK was performed using the ZINC 15 database and resulted in 9 hit compounds which showed high binding affinities and binding energy to the enzyme. The HPPK and PTR1 enzyme coding sequences were cloned into pET-28a(+) plasmids and supplied by GenScript, to enable the expression of histidine-tagged proteins in T7 Express lysY competent E. coli cells. Analytical scale expression studies showed the recombinant proteins to be in a soluble form and purification was achieved using nickel-NTA affinity chromatography. The purified PTR1 recombinant protein was used to establish and optimise an NADPH absorbance microplate assay to screen compounds previously identified in docking studies by Kimuda, Laming, Hoppe, & Bishop, (2019). The assay yielded a Z´-factor of above 0.8 indicating an excellent assay to use for screening. An unsuccessful attempt was made to use resazurin reduction as an alternative method to demonstrate PTR1 enzyme activity. HPPK purified recombinant proteins were used to establish and optimise a luminescence microplate assay for the screening of compounds identified in in silico docking studies against the HPPK enzymes of S. enterica and E. coli. The Z´-factor of the luminescence assay was above 0.5, indicative of a functional assay with good separation between enzyme activity signal and negative control of reaction without enzyme. The target-based enzyme screening resulted in the confirmation of compound 3 (3-chloro-N-[(4-oxo-3,4-dihydrophthalazin-1-yl)methyl]benzamide) as an inhibitor of S. enterica HPPK with an IC₅₀ of 10.4 μM. At 50 μM none of the compounds decreased E. coli HPPK enzyme activity by 50%. Further bacterial studies would provide more compelling data to motivate the optimisation of compound 3 as an S. enterica inhibitor. This study demonstrated the effectiveness of using computational methods in the drug discovery process, correlating in silico results with those obtained from target-based assays producing a hit compound that can be used for future drug optimisation. , Thesis (MSc (Pharm)) -- Faculty of Pharmacy, Pharmacy, 2021
- Full Text:
An evaluation of synergistic interactions between feruloyl esterases and xylanases during the hydrolysis of various pre-treated agricultural residues
- Authors: Mkabayi, Lithalethu
- Date: 2021-04
- Subjects: Esterases , Xylanases , Hydrolysis , Agricultural wastes -- Recycling , Enzymes , Lignocellulose -- Biodegradation , Escherichia coli , Oligosaccharides , Hydroxycinnamic acids
- Language: English
- Type: thesis , text , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/178224 , vital:42922 , 10.21504/10962/178224
- Description: Agricultural residues are readily available and inexpensive renewable resources that can be used as raw materials for the production of value-added chemicals. The application of enzymes to facilitate the degradation of agricultural residues has long been considered the most environmentally friendly strategy for converting this material into good quality value-added chemicals. However, agricultural residues are typically lignocellulosic in composition and recalcitrant to enzymatic hydrolysis. Due to this recalcitrant nature, the complete degradation of biomass residues requires the synergistic action of a broad range of enzymes. The development and optimisation of synergistic enzyme cocktails is an effective approach for achieving high hydrolysis efficiency of lignocellulosic biomass. The aim of the current study was to evaluate the synergistic interactions between two termite metagenome-derived feruloyl esterases (FAE6 and FAE5) and endo-xylanases for the production of hydroxycinnamic acids and xylo-oligosaccharides (XOS) from model substrates, and untreated and pre-treated agricultural residues. Firstly, the two fae genes were heterologously expressed in Escherichia coli, and the recombinant enzymes were purified to homogeneity. The biochemical properties of the purified recombinant FAEs and xylanases (XT6 and Xyn11) were then assessed to determine the factors which influenced their activities and to select suitable operating conditions for synergy studies. An optimal protein loading ratio of xylanases to FAEs required to maximise the release of both reducing sugar and ferulic acid (FA) was established using 0.5% (w/v) insoluble wheat arabinoxylan (a model substrate). The enzyme combination of 66% xylanase and 33% FAE (on a protein loading basis) produced the highest amounts of reducing sugars and FA. The enzyme combination of XT6 (GH10 xylanase) and FAE5 or FAE6 liberated the highest amount of FA while a combination of Xyn11 (GH11 xylanase) and FAE5 or FAE6 produced the highest reducing sugar content. The synergistic interactions which were established between the xylanases and FAEs were further investigated using agricultural residues (corn cobs, rice straw and sugarcane bagasse). The three substrates were subjected to hydrothermal and dilute acid pre-treatment prior to synergy studies. It is generally known that, during pre-treatment, many compounds can be produced which may influence enzymatic hydrolysis. The effects of these by-products were assessed and it was found that lignin and its degradation products were the most inhibitory to the FAEs. The optimised enzyme cocktail was then applied to 1% (w/v) of untreated and pre-treated substrates for the efficient production of XOS and hydroxycinnamic acids. A significant improvement in xylanase substrate degradation was observed, especially with the combination of 66% Xyn11 and 33% FAE6 which displayed an improvement in reducing sugars of approximately 1.9-fold and 3.4-fold for hydrothermal and acid pre-treated corn cobs (compared to when Xyn11 was used alone), respectively. The study demonstrated that pre-treatment substantially enhanced the enzymatic hydrolysis of corn cobs and rice straw. Analysis of the hydrolysate product profiles revealed that the optimised enzyme cocktail displayed great potential for releasing XOS with a low degree of polymerisation. In conclusion, this study provided significant insights into the mechanism of synergistic interactions between xylanases and metagenome-derived FAEs during the hydrolysis of various substrates. The study also demonstrated that optimised enzyme cocktails combined with low severity pre-treatment can facilitate the potential use of xylan-rich lignocellulosic biomass for the production of valuable products in the future. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2021
- Full Text:
- Authors: Mkabayi, Lithalethu
- Date: 2021-04
- Subjects: Esterases , Xylanases , Hydrolysis , Agricultural wastes -- Recycling , Enzymes , Lignocellulose -- Biodegradation , Escherichia coli , Oligosaccharides , Hydroxycinnamic acids
- Language: English
- Type: thesis , text , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/178224 , vital:42922 , 10.21504/10962/178224
- Description: Agricultural residues are readily available and inexpensive renewable resources that can be used as raw materials for the production of value-added chemicals. The application of enzymes to facilitate the degradation of agricultural residues has long been considered the most environmentally friendly strategy for converting this material into good quality value-added chemicals. However, agricultural residues are typically lignocellulosic in composition and recalcitrant to enzymatic hydrolysis. Due to this recalcitrant nature, the complete degradation of biomass residues requires the synergistic action of a broad range of enzymes. The development and optimisation of synergistic enzyme cocktails is an effective approach for achieving high hydrolysis efficiency of lignocellulosic biomass. The aim of the current study was to evaluate the synergistic interactions between two termite metagenome-derived feruloyl esterases (FAE6 and FAE5) and endo-xylanases for the production of hydroxycinnamic acids and xylo-oligosaccharides (XOS) from model substrates, and untreated and pre-treated agricultural residues. Firstly, the two fae genes were heterologously expressed in Escherichia coli, and the recombinant enzymes were purified to homogeneity. The biochemical properties of the purified recombinant FAEs and xylanases (XT6 and Xyn11) were then assessed to determine the factors which influenced their activities and to select suitable operating conditions for synergy studies. An optimal protein loading ratio of xylanases to FAEs required to maximise the release of both reducing sugar and ferulic acid (FA) was established using 0.5% (w/v) insoluble wheat arabinoxylan (a model substrate). The enzyme combination of 66% xylanase and 33% FAE (on a protein loading basis) produced the highest amounts of reducing sugars and FA. The enzyme combination of XT6 (GH10 xylanase) and FAE5 or FAE6 liberated the highest amount of FA while a combination of Xyn11 (GH11 xylanase) and FAE5 or FAE6 produced the highest reducing sugar content. The synergistic interactions which were established between the xylanases and FAEs were further investigated using agricultural residues (corn cobs, rice straw and sugarcane bagasse). The three substrates were subjected to hydrothermal and dilute acid pre-treatment prior to synergy studies. It is generally known that, during pre-treatment, many compounds can be produced which may influence enzymatic hydrolysis. The effects of these by-products were assessed and it was found that lignin and its degradation products were the most inhibitory to the FAEs. The optimised enzyme cocktail was then applied to 1% (w/v) of untreated and pre-treated substrates for the efficient production of XOS and hydroxycinnamic acids. A significant improvement in xylanase substrate degradation was observed, especially with the combination of 66% Xyn11 and 33% FAE6 which displayed an improvement in reducing sugars of approximately 1.9-fold and 3.4-fold for hydrothermal and acid pre-treated corn cobs (compared to when Xyn11 was used alone), respectively. The study demonstrated that pre-treatment substantially enhanced the enzymatic hydrolysis of corn cobs and rice straw. Analysis of the hydrolysate product profiles revealed that the optimised enzyme cocktail displayed great potential for releasing XOS with a low degree of polymerisation. In conclusion, this study provided significant insights into the mechanism of synergistic interactions between xylanases and metagenome-derived FAEs during the hydrolysis of various substrates. The study also demonstrated that optimised enzyme cocktails combined with low severity pre-treatment can facilitate the potential use of xylan-rich lignocellulosic biomass for the production of valuable products in the future. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2021
- Full Text:
Photodynamic antimicrobial chemotherapy against Staphylococcus aureus and Escherichia coli sensitized using indium (III) cationic porphyrins linked to core-shell magnetic nanoparticles
- Authors: Makola, Lekgowa Collen
- Date: 2021-04
- Subjects: Photochemotherapy , Photosensitizing compounds , Staphylococcus aureus , Escherichia coli , Indium , Porphyrins , Magnetic nanoparticles , Quaternize
- Language: English
- Type: thesis , text , Masters , MSc
- Identifier: http://hdl.handle.net/10962/177225 , vital:42801
- Description: Photodynamic antimicrobial chemotherapy (PACT) is a well-known antimicrobial therapy technique used against multi-drug resistant pathogens. In this study, the syntheses, characterization, photophysicochemical properties, and the applications of symmetrically and asymmetrically substituted cationic indium (III) porphyrins linked to silver/copper ferrite core-shell (Ag/CuFe2O4) magnetic nanoparticles (MNPs) as potential photosensitizers for PACT are reported. The synthesized complexes include axially modified porphyrins quaternized through an axial ligand. All the asymmetrically substituted porphyrins were linked to the NPs via an ester bond and the symmetrically substituted porphyrins were linked (peripherally and /or axially) via self-assembly (Ag-S and/or Ag-N). The impact of axial modification, peripheral substituents, conjugation to the NPs, the number of positive charges, and the chain length of the alkyl halides quaternizing agents on PACT efficacy and photophysicochemical properties of porphyrins were studied. High singlet oxygen quantum yields and antimicrobial log reductions were observed. Lipophilicity and hydrophilicity of the porphyrins were also studies, where the complexes quaternized with methyl iodide showed relatively high hydrophilicity character. Upon in vitro PACT studies, the quaternized complexes were observed to have 0% viable colony, signifying their effectiveness. Moreover, the highest log reductions of 9.27 were observed against S. aureus and 9.58 were observed against E. coli. The findings from this work delineate that singlet oxygen generation alone is not a distinct factor on the PACT efficacy of the porphyrin complexes, since some of the complexes have practically the same singlet oxygen quantum but different PACT activity. However, other contributing factors including water solubility play a significant role. , Thesis (MSc) -- Faculty of Science, Department of Chemistry, 2021
- Full Text:
- Authors: Makola, Lekgowa Collen
- Date: 2021-04
- Subjects: Photochemotherapy , Photosensitizing compounds , Staphylococcus aureus , Escherichia coli , Indium , Porphyrins , Magnetic nanoparticles , Quaternize
- Language: English
- Type: thesis , text , Masters , MSc
- Identifier: http://hdl.handle.net/10962/177225 , vital:42801
- Description: Photodynamic antimicrobial chemotherapy (PACT) is a well-known antimicrobial therapy technique used against multi-drug resistant pathogens. In this study, the syntheses, characterization, photophysicochemical properties, and the applications of symmetrically and asymmetrically substituted cationic indium (III) porphyrins linked to silver/copper ferrite core-shell (Ag/CuFe2O4) magnetic nanoparticles (MNPs) as potential photosensitizers for PACT are reported. The synthesized complexes include axially modified porphyrins quaternized through an axial ligand. All the asymmetrically substituted porphyrins were linked to the NPs via an ester bond and the symmetrically substituted porphyrins were linked (peripherally and /or axially) via self-assembly (Ag-S and/or Ag-N). The impact of axial modification, peripheral substituents, conjugation to the NPs, the number of positive charges, and the chain length of the alkyl halides quaternizing agents on PACT efficacy and photophysicochemical properties of porphyrins were studied. High singlet oxygen quantum yields and antimicrobial log reductions were observed. Lipophilicity and hydrophilicity of the porphyrins were also studies, where the complexes quaternized with methyl iodide showed relatively high hydrophilicity character. Upon in vitro PACT studies, the quaternized complexes were observed to have 0% viable colony, signifying their effectiveness. Moreover, the highest log reductions of 9.27 were observed against S. aureus and 9.58 were observed against E. coli. The findings from this work delineate that singlet oxygen generation alone is not a distinct factor on the PACT efficacy of the porphyrin complexes, since some of the complexes have practically the same singlet oxygen quantum but different PACT activity. However, other contributing factors including water solubility play a significant role. , Thesis (MSc) -- Faculty of Science, Department of Chemistry, 2021
- Full Text:
Assessment of the microbial quality of various domestic rainwater harvesting systems and the suitability of a nano based treatment method
- Authors: Malema, Mokaba Shirley
- Date: 2020
- Subjects: Escherichia coli , Water harvesting , Microbial contamination , Water Purification , Physicochemical process
- Language: English
- Type: thesis , text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/110218 , vital:33249 , https://dx.doi.org/10.21504/10962/110218
- Description: Thesis (PhD)--Rhodes University, Faculty of Science, Chemistry, 2020. , In most developing countries, people from rural and peri-urban settlements depend on harvested rainwater (HRW) as an alternative water source for drinking and other household purposes. Despite this reliance, there is little monitoring of the microbial quality of HRW in these areas. The most important issue in relation to using untreated harvested rainwater for drinking and other domestic purposes is the potential public health risks associated with microbial pathogens. Unlike chemical contamination, microbial contamination my lead to disease occurring rapidly, hence the need for frequent monitoring. Thus, the current study investigated the microbial quality of various domestic rainwater harvesting systems and the suitability of a nano based treatment method. The first experiments involved determining the microbial (Escherichia coli) and physicochemical quality (pH, turbidity, nitrate and chemical oxygen demand (COD)) of HRW in the Eastern Cape Province, South Africa. Samples were collected from 11 tanks situated at the Rhodes University, Kenton-on-sea (coastal) and in homes in the Grahamstown area on a weekly basis between June and September 2016. The Colilert-18®/Quanti-tray® 2000 system was used for enumeration of E. coli while physicochemical parameters were measured using commercial kits. Results showed that all samples were contaminated with varying concentrations of E. coli ranging from 7 to 1055 MPN/100 mL. Physicochemical analysis revealed that pH ranged from 5.6 to 7.6 and Turbidity values obtained for all tanks were below 5 NTU except for tank 4 (5.12 ± 4.96 NTU) and 7 (5.58 ± 8.19 NTU). Nitrate levels (range: 5.95 to 28.12 mg L-1) and COD (range: 66.53 to 191.12 mg L-1) were higher than the recommended South African drinking water quality guidelines in most of the tanks. In the second experiments, the objective was to determine whether a modified hydrogen sulphide (H2S) test kit with an improved detection rate is an effective preliminary screening qualitative test that can be used for rainwater quality monitoring. The hydrogen sulphide method is a low-cost microbiological field-based test which can be used in areas where water testing facilities are limited. Harvested rainwater samples were collected from various tanks in the Eastern Cape and tested for contaminants of faecal origin using the modified hydrogen sulphide test kit, Colilert-18/Quanti-tray®/2000 and membrane filtration technique. Faecal coliforms were measured using membrane filtration, E. coli was measured using Colilert and correspondence rates were calculated with results of the improved hydrogen sulphide test kit. E. coli results ranged from <1 – >2419.6 MPN/100 mL while the faecal coliforms ranged from 0 – >300 CFU/mL. The agreement rate with hydrogen sulphide test and membrane filtration was 88% while the agreement rate for the Colilert and hydrogen sulphide test was 76%. The third experiments investigated the prevalence of pathogenic E. coli strains and their antimicrobial resistance patterns in HRW tanks in the Eastern Cape, South Africa. E. coli isolates obtained in the first experiments were further screened for their virulence potentials using polymerase chain reaction (PCR) and subsequently tested for antibiotic resistance using the disc-diffusion method against 11 antibiotics. The pathotype most detected was the neonatal meningitis E. coli (NMEC) (ibeA 28%) while pathotype enteroaggregative E. coli (EAEC) was not detected. The highest resistance of the E. coli isolates was observed against Cephalothin (76%). All tested pathotypes were susceptible to Gentamicin, and 52% demonstrated multiple-antibiotic resistance (MAR). The fourth experiments shed light on the occurrence of Legionella, zoonotic and fungal pathogens in the rainwater harvesting systems (RWHS) situated in different regions of South Africa. Rainwater samples were collected in urban and semi-urban areas from tanks situated in various areas in South Africa (Johannesburg, Pretoria and Grahamstown). Pathogenic organisms investigated were Salmonella, Shigella, Vibrio cholerae, Legionella and fungal isolates. Pure isolates were obtained and screened using PCR. Results revealed the presence of pathogenic bacteria and fungi in all the tested RWHS. In Grahamstown the most detected pathogen was Salmonella (73%) while Vibrio Cholerae was not detected. All the tested pathogens were present from the RWHS situated in Pretoria. Shigella was not detected from the RWHS in Johannesburg while others were detected. Identification of fungal isolates from HRW showed the presence of pathogenic fungi such as Aspergillus fumigatus, Cryptococcus laurentii, Aureobasidium pullulans and Mucor circinelloides. The last experiments, focussed on exploring a suitable treatment method for HRW where a nano compound quaternary imidazolium modified montmorillonite (MMT) was used as a potential household rainwater treatment option. Harvested rainwater samples were collected from the RWHS situated at the Council for Scientific and Industrial Research (CSIR), Pretoria South Africa. River and borehole water samples were included in the study to check the efficiency of the treatment method on various water sources. River water samples were collected from Olifants River, Witbank, South Africa while borehole water was collected from a privately-owned borehole in Pretoria. For inoculation studies, all the water sources were sterilised in batches of 1 and 2 L and inoculated with approximately 107 CFU/mL of overnight E. coli. Approximately 200 mg of the quaternary imidazolium modified MMT was added to the inoculated water and samples collected immediately after inoculation (time 0) and thereafter every hour for 5 hrs. The analyses were further conducted using unsterilised water samples (total bacterial count) and 500 mg of the treatment material. Complete inactivation of E. coli in sterilised HRW was achieved in 2 hrs for the 2 L water samples and 3 hrs for the 1 L water samples. Sterilised river water achieved complete E. coli inactivation in 4 hrs for the 1 L and 5 hrs for the 2 L samples while borehole water samples achieved complete E. coli inactivation in 5 hrs (2 L) and 6 hrs for the 1 L samples. In the unsterilised water sources (total bacteria), complete bacterial inactivation was observed in 5 hrs for both the 1 and 2 L harvested rainwater samples, 6 hrs in river water samples (both 1 and 2 L) and 8 hrs for borehole water samples (1 and 2 L). The results suggest that the treatment option was more efficient in harvested rainwater (required less time for bacterial inactivation compared to river and borehole water). The results of the current study are of public health concern since the use of untreated HRW for potable purposes may pose a risk of transmission of pathogenic and antimicrobial-resistant E. coli and other pathogenic organisms such as Salmonella, Shigella and Vibrio cholerae. It is therefore recommended that in cases where the tested harvested rainwater is used for potable purposes, simple treatment methods such as boiling and SODIS be applied so the harvested rainwater is fit for human consumption.
- Full Text: false
- Authors: Malema, Mokaba Shirley
- Date: 2020
- Subjects: Escherichia coli , Water harvesting , Microbial contamination , Water Purification , Physicochemical process
- Language: English
- Type: thesis , text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/110218 , vital:33249 , https://dx.doi.org/10.21504/10962/110218
- Description: Thesis (PhD)--Rhodes University, Faculty of Science, Chemistry, 2020. , In most developing countries, people from rural and peri-urban settlements depend on harvested rainwater (HRW) as an alternative water source for drinking and other household purposes. Despite this reliance, there is little monitoring of the microbial quality of HRW in these areas. The most important issue in relation to using untreated harvested rainwater for drinking and other domestic purposes is the potential public health risks associated with microbial pathogens. Unlike chemical contamination, microbial contamination my lead to disease occurring rapidly, hence the need for frequent monitoring. Thus, the current study investigated the microbial quality of various domestic rainwater harvesting systems and the suitability of a nano based treatment method. The first experiments involved determining the microbial (Escherichia coli) and physicochemical quality (pH, turbidity, nitrate and chemical oxygen demand (COD)) of HRW in the Eastern Cape Province, South Africa. Samples were collected from 11 tanks situated at the Rhodes University, Kenton-on-sea (coastal) and in homes in the Grahamstown area on a weekly basis between June and September 2016. The Colilert-18®/Quanti-tray® 2000 system was used for enumeration of E. coli while physicochemical parameters were measured using commercial kits. Results showed that all samples were contaminated with varying concentrations of E. coli ranging from 7 to 1055 MPN/100 mL. Physicochemical analysis revealed that pH ranged from 5.6 to 7.6 and Turbidity values obtained for all tanks were below 5 NTU except for tank 4 (5.12 ± 4.96 NTU) and 7 (5.58 ± 8.19 NTU). Nitrate levels (range: 5.95 to 28.12 mg L-1) and COD (range: 66.53 to 191.12 mg L-1) were higher than the recommended South African drinking water quality guidelines in most of the tanks. In the second experiments, the objective was to determine whether a modified hydrogen sulphide (H2S) test kit with an improved detection rate is an effective preliminary screening qualitative test that can be used for rainwater quality monitoring. The hydrogen sulphide method is a low-cost microbiological field-based test which can be used in areas where water testing facilities are limited. Harvested rainwater samples were collected from various tanks in the Eastern Cape and tested for contaminants of faecal origin using the modified hydrogen sulphide test kit, Colilert-18/Quanti-tray®/2000 and membrane filtration technique. Faecal coliforms were measured using membrane filtration, E. coli was measured using Colilert and correspondence rates were calculated with results of the improved hydrogen sulphide test kit. E. coli results ranged from <1 – >2419.6 MPN/100 mL while the faecal coliforms ranged from 0 – >300 CFU/mL. The agreement rate with hydrogen sulphide test and membrane filtration was 88% while the agreement rate for the Colilert and hydrogen sulphide test was 76%. The third experiments investigated the prevalence of pathogenic E. coli strains and their antimicrobial resistance patterns in HRW tanks in the Eastern Cape, South Africa. E. coli isolates obtained in the first experiments were further screened for their virulence potentials using polymerase chain reaction (PCR) and subsequently tested for antibiotic resistance using the disc-diffusion method against 11 antibiotics. The pathotype most detected was the neonatal meningitis E. coli (NMEC) (ibeA 28%) while pathotype enteroaggregative E. coli (EAEC) was not detected. The highest resistance of the E. coli isolates was observed against Cephalothin (76%). All tested pathotypes were susceptible to Gentamicin, and 52% demonstrated multiple-antibiotic resistance (MAR). The fourth experiments shed light on the occurrence of Legionella, zoonotic and fungal pathogens in the rainwater harvesting systems (RWHS) situated in different regions of South Africa. Rainwater samples were collected in urban and semi-urban areas from tanks situated in various areas in South Africa (Johannesburg, Pretoria and Grahamstown). Pathogenic organisms investigated were Salmonella, Shigella, Vibrio cholerae, Legionella and fungal isolates. Pure isolates were obtained and screened using PCR. Results revealed the presence of pathogenic bacteria and fungi in all the tested RWHS. In Grahamstown the most detected pathogen was Salmonella (73%) while Vibrio Cholerae was not detected. All the tested pathogens were present from the RWHS situated in Pretoria. Shigella was not detected from the RWHS in Johannesburg while others were detected. Identification of fungal isolates from HRW showed the presence of pathogenic fungi such as Aspergillus fumigatus, Cryptococcus laurentii, Aureobasidium pullulans and Mucor circinelloides. The last experiments, focussed on exploring a suitable treatment method for HRW where a nano compound quaternary imidazolium modified montmorillonite (MMT) was used as a potential household rainwater treatment option. Harvested rainwater samples were collected from the RWHS situated at the Council for Scientific and Industrial Research (CSIR), Pretoria South Africa. River and borehole water samples were included in the study to check the efficiency of the treatment method on various water sources. River water samples were collected from Olifants River, Witbank, South Africa while borehole water was collected from a privately-owned borehole in Pretoria. For inoculation studies, all the water sources were sterilised in batches of 1 and 2 L and inoculated with approximately 107 CFU/mL of overnight E. coli. Approximately 200 mg of the quaternary imidazolium modified MMT was added to the inoculated water and samples collected immediately after inoculation (time 0) and thereafter every hour for 5 hrs. The analyses were further conducted using unsterilised water samples (total bacterial count) and 500 mg of the treatment material. Complete inactivation of E. coli in sterilised HRW was achieved in 2 hrs for the 2 L water samples and 3 hrs for the 1 L water samples. Sterilised river water achieved complete E. coli inactivation in 4 hrs for the 1 L and 5 hrs for the 2 L samples while borehole water samples achieved complete E. coli inactivation in 5 hrs (2 L) and 6 hrs for the 1 L samples. In the unsterilised water sources (total bacteria), complete bacterial inactivation was observed in 5 hrs for both the 1 and 2 L harvested rainwater samples, 6 hrs in river water samples (both 1 and 2 L) and 8 hrs for borehole water samples (1 and 2 L). The results suggest that the treatment option was more efficient in harvested rainwater (required less time for bacterial inactivation compared to river and borehole water). The results of the current study are of public health concern since the use of untreated HRW for potable purposes may pose a risk of transmission of pathogenic and antimicrobial-resistant E. coli and other pathogenic organisms such as Salmonella, Shigella and Vibrio cholerae. It is therefore recommended that in cases where the tested harvested rainwater is used for potable purposes, simple treatment methods such as boiling and SODIS be applied so the harvested rainwater is fit for human consumption.
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Targeting allosteric sites of Escherichia coli heat shock protein 70 for antibiotic development
- Authors: Okeke, Chiamaka Jessica
- Date: 2019
- Subjects: Heat shock proteins , Escherichia coli , Allosteric proteins , Antibiotics , Molecular chaperones , Ligands (Biochemistry) , Molecular dynamics , Principal components analysis , South African Natural Compounds Database
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/115998 , vital:34287
- Description: Hsp70s are members of the heat shock proteins family with a molecular weight of 70-kDa and are the most abundant group in bacterial and eukaryotic systems, hence the most extensively studied ones. These proteins are molecular chaperones that play a significant role in protein homeostasis by facilitating appropriate folding of proteins, preventing proteins from aggregating and misfolding. They are also involved in translocation of proteins into subcellular compartments and protection of cells against stress. Stress caused by environmental or biological factors affects the functionality of the cell. In response to these stressful conditions, up-regulation of Hsp70s ensures that the cells are protected by balancing out unfolded proteins giving them ample time to repair denatured proteins. Hsp70s is connected to numerous illnesses such as autoimmune and neurodegenerative diseases, bacterial infection, cancer, malaria, and obesity. The multi-functional nature of Hsp70s predisposes them as promising therapeutic targets. Hsp70s play vital roles in various cell developments, and survival pathways, therefore targeting this protein will provide a new avenue towards the discovery of active therapeutic agents for the treatment of a wide range of diseases. Allosteric sites of these proteins in its multi-conformational states have not been explored for inhibitory properties hence the aim of this study. This study aims at identifying allosteric sites that inhibit the ATPase and substrate binding activities using computational approaches. Using E. coli as a model organism, molecular docking for high throughput virtual screening was carried out using 623 compounds from the South African Natural Compounds Database (SANCDB; https://sancdb.rubi.ru.ac.za/) against identified allosteric sites. Ligands with the highest binding affinity (good binders) interacting with critical allosteric residues that are druggable were identified. Molecular dynamics (MD) simulation was also performed on the identified hits to assess for protein-inhibitor complex stability. Finally, principal component analysis (PCA) was performed to understand the structural dynamics of the ligand-free and ligand-bound structures during MD simulation.
- Full Text:
- Authors: Okeke, Chiamaka Jessica
- Date: 2019
- Subjects: Heat shock proteins , Escherichia coli , Allosteric proteins , Antibiotics , Molecular chaperones , Ligands (Biochemistry) , Molecular dynamics , Principal components analysis , South African Natural Compounds Database
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/115998 , vital:34287
- Description: Hsp70s are members of the heat shock proteins family with a molecular weight of 70-kDa and are the most abundant group in bacterial and eukaryotic systems, hence the most extensively studied ones. These proteins are molecular chaperones that play a significant role in protein homeostasis by facilitating appropriate folding of proteins, preventing proteins from aggregating and misfolding. They are also involved in translocation of proteins into subcellular compartments and protection of cells against stress. Stress caused by environmental or biological factors affects the functionality of the cell. In response to these stressful conditions, up-regulation of Hsp70s ensures that the cells are protected by balancing out unfolded proteins giving them ample time to repair denatured proteins. Hsp70s is connected to numerous illnesses such as autoimmune and neurodegenerative diseases, bacterial infection, cancer, malaria, and obesity. The multi-functional nature of Hsp70s predisposes them as promising therapeutic targets. Hsp70s play vital roles in various cell developments, and survival pathways, therefore targeting this protein will provide a new avenue towards the discovery of active therapeutic agents for the treatment of a wide range of diseases. Allosteric sites of these proteins in its multi-conformational states have not been explored for inhibitory properties hence the aim of this study. This study aims at identifying allosteric sites that inhibit the ATPase and substrate binding activities using computational approaches. Using E. coli as a model organism, molecular docking for high throughput virtual screening was carried out using 623 compounds from the South African Natural Compounds Database (SANCDB; https://sancdb.rubi.ru.ac.za/) against identified allosteric sites. Ligands with the highest binding affinity (good binders) interacting with critical allosteric residues that are druggable were identified. Molecular dynamics (MD) simulation was also performed on the identified hits to assess for protein-inhibitor complex stability. Finally, principal component analysis (PCA) was performed to understand the structural dynamics of the ligand-free and ligand-bound structures during MD simulation.
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Molecular cloning and expression of equine CYP1A2 in Escherichia coli
- Authors: Mkabayi, Lithalethu
- Date: 2017
- Subjects: Escherichia coli , Escherichia coli infections in animals , Cytochrome P-450 , Cytochromes , Horses -- Effect of drugs on
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/4830 , vital:20734
- Description: Information regarding drug metabolism in veterinary species, especially horses, remains fragmented and incomplete. This information is essential for detection of metabolites of potential performance-enhancing substances in horseracing and for veterinary drug development. Equine liver microsomes have been used to study metabolism of a limited number of drugs, but these provide little information about individual drug metabolizing enzymes. Recombinant CYP enzyme systems are commonly used to determine contribution of individual CYP to metabolism of specific drugs. A limited number of recombinant equine CYPs have been expressed in insect cells and mammalian cell lines. However, there are no reports of recombinant equine CYP1A2 enzyme. In this study, equine CYP1A2 was identified, codon-optimized, cloned and expressed in E. coli BL21 cells. Multiple sequence alignments of equine CYP1A2 revealed an amino acid sequence identity of 83.69% to its human homolog which has previously been expressed in E. coli. The enzyme was expressed using both auto-induction and IPTG induction. Expressed equine CYP1A2 had a size of about 55 kDa, and was insoluble after cell lysis. Sarkosyl- solubilized CYP1A2 was purified using nickel affinity chromatography and gel filtration. For activity reconstitution, yeast NADPH-cytochrome P450 reductase was first expressed in E. coli BL21 cells and exhibited activity of 0.13 U/ml. Activity assay with Glo-P450 CYP1A2 assay kit indicated that CYP1A2 was inactive. Despite numerous attempts to obtain the activity, the CYP1A2 remained inactive. Although expression of equine CYP1A2 in E. coli produced non- catalytically active enzyme, this study could be used as the first step in an effort to fully develop a recombinant equine CYP1A2 system.
- Full Text:
- Authors: Mkabayi, Lithalethu
- Date: 2017
- Subjects: Escherichia coli , Escherichia coli infections in animals , Cytochrome P-450 , Cytochromes , Horses -- Effect of drugs on
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/4830 , vital:20734
- Description: Information regarding drug metabolism in veterinary species, especially horses, remains fragmented and incomplete. This information is essential for detection of metabolites of potential performance-enhancing substances in horseracing and for veterinary drug development. Equine liver microsomes have been used to study metabolism of a limited number of drugs, but these provide little information about individual drug metabolizing enzymes. Recombinant CYP enzyme systems are commonly used to determine contribution of individual CYP to metabolism of specific drugs. A limited number of recombinant equine CYPs have been expressed in insect cells and mammalian cell lines. However, there are no reports of recombinant equine CYP1A2 enzyme. In this study, equine CYP1A2 was identified, codon-optimized, cloned and expressed in E. coli BL21 cells. Multiple sequence alignments of equine CYP1A2 revealed an amino acid sequence identity of 83.69% to its human homolog which has previously been expressed in E. coli. The enzyme was expressed using both auto-induction and IPTG induction. Expressed equine CYP1A2 had a size of about 55 kDa, and was insoluble after cell lysis. Sarkosyl- solubilized CYP1A2 was purified using nickel affinity chromatography and gel filtration. For activity reconstitution, yeast NADPH-cytochrome P450 reductase was first expressed in E. coli BL21 cells and exhibited activity of 0.13 U/ml. Activity assay with Glo-P450 CYP1A2 assay kit indicated that CYP1A2 was inactive. Despite numerous attempts to obtain the activity, the CYP1A2 remained inactive. Although expression of equine CYP1A2 in E. coli produced non- catalytically active enzyme, this study could be used as the first step in an effort to fully develop a recombinant equine CYP1A2 system.
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Photophysicochemical and photodynamic antimicrobial chemotherapeutic studies of novel phthalocyanines conjugated to silver nanoparticles
- Authors: Rapulenyane, Nomasonto
- Date: 2013 , 2013-06-10
- Subjects: Phthalocyanines , Photochemistry , Photochemotherapy , Cancer -- Photochemotherapy , Anti-infective agents , Escherichia coli , Nanoparticles , Silver , Zinc
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4291 , http://hdl.handle.net/10962/d1003912 , Phthalocyanines , Photochemistry , Photochemotherapy , Cancer -- Photochemotherapy , Anti-infective agents , Escherichia coli , Nanoparticles , Silver , Zinc
- Description: This work reports on the synthesis, characterization and the physicochemical properties of novel unsymmetrically substituted zinc phthalocyanines: namely tris{11,19, 27-(1,2- diethylaminoethylthiol)-2-(captopril) phthalocyanine Zn ((ZnMCapPc (1.5)), hexakis{8,11,16,19,42,27-(octylthio)-1-(4-phenoxycarboxy) phthalocyanine} Zn (ZnMPCPc(1.7)) and Tris {11, 19, 27-(1,2-diethylaminoethylthiol)-1,2(caffeic acid) phthalocyanine} Zn ((ZnMCafPc (1.3)). Symmetrically substituted counterparts (tetrakis(diethylamino)zinc phthalocyaninato (3.8), octakis(octylthio)zinc phthalocyaninato (3.9) and tetrakis (carboxyphenoxy)zinc phthalocyaninato (3.10) complexes) were also synthesized for comparison of the photophysicochemical properties and to investigate the effect of the substituents on the low symmetry Pcs. The complexes were successfully characterized by IR, NMR, mass spectral and elemental analyses. All the complexes showed the ability to produce singlet oxygen, while the highest triplet quantum yields were obtained for 1.7, 1.5 and 3.9 (0.80, 0.65 and 0.62 respectively and the lowest were obtained for 1.3 and 3.10 (0.57 and 0.47 respectively). High triplet lifetimes (109-286 μs) were also obtained for all complexes, with 1.7 being the highest (286 μs) which also corresponds to its triplet and singlet quantum yields (0.80 and 0.77 respectively). The photosensitizing properties of low symmetry derivatives, ZnMCapPc and ZnMCafPc were investigated by conjugating glutathione (GSH) capped silver nanoparticles (AgNP). The formation of the amide bond was confirmed by IR and UV-Vis spectroscopies. The photophysicochemical behaviour of the novel phthalocyanine-GSH-AgNP conjugates and the simple mixture of the Ag NPs with low the symmetry phthalocyanines were investigated. It was observed that upon conjugation of the phthalocyanines to the GSH-AgNPs, a blue shift in the Q band was induced. The triplet lifetimes and quantum yields improved upon conjugation as compared to the phthalocyanines (Pc) alone. Complex 1.5 triplet lifetimes increased from 109 to 148 and triplet quantum yield from 0.65 to 0.86 upon conjugation. Fluorescence lifetimes and quantum yields decreased for the conjugates compared to the phthalocyanines alone, due to the quenching caused by the Ag NPs. The antimicrobial activity of the zinc phthalocyanines (complexes 1.3 and 1.5) and their conjugates against Escherichia coli was investigated. Only 1.3 and 1.5 complexes were investigated because of the availability of the sample. In general phthalocyanines showed increase in antibacterial activity with the increase in phthalocyanines concentration in the presence and absence of light. The Pc complexes and their Ag NP conjugates showed an increase in antibacterial activity, due to the synergistic effect afforded by Ag NP and Pcs. Improved antibacterial properties were obtained upon irradiation. 1.5-AgNPs had the highest antibacterial activity compared to 1.3-AgNPs conjugate; these results are in agreement with the photophysical behaviour. This work demonstrates improved photophysicochemical properties of low symm
- Full Text:
- Authors: Rapulenyane, Nomasonto
- Date: 2013 , 2013-06-10
- Subjects: Phthalocyanines , Photochemistry , Photochemotherapy , Cancer -- Photochemotherapy , Anti-infective agents , Escherichia coli , Nanoparticles , Silver , Zinc
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4291 , http://hdl.handle.net/10962/d1003912 , Phthalocyanines , Photochemistry , Photochemotherapy , Cancer -- Photochemotherapy , Anti-infective agents , Escherichia coli , Nanoparticles , Silver , Zinc
- Description: This work reports on the synthesis, characterization and the physicochemical properties of novel unsymmetrically substituted zinc phthalocyanines: namely tris{11,19, 27-(1,2- diethylaminoethylthiol)-2-(captopril) phthalocyanine Zn ((ZnMCapPc (1.5)), hexakis{8,11,16,19,42,27-(octylthio)-1-(4-phenoxycarboxy) phthalocyanine} Zn (ZnMPCPc(1.7)) and Tris {11, 19, 27-(1,2-diethylaminoethylthiol)-1,2(caffeic acid) phthalocyanine} Zn ((ZnMCafPc (1.3)). Symmetrically substituted counterparts (tetrakis(diethylamino)zinc phthalocyaninato (3.8), octakis(octylthio)zinc phthalocyaninato (3.9) and tetrakis (carboxyphenoxy)zinc phthalocyaninato (3.10) complexes) were also synthesized for comparison of the photophysicochemical properties and to investigate the effect of the substituents on the low symmetry Pcs. The complexes were successfully characterized by IR, NMR, mass spectral and elemental analyses. All the complexes showed the ability to produce singlet oxygen, while the highest triplet quantum yields were obtained for 1.7, 1.5 and 3.9 (0.80, 0.65 and 0.62 respectively and the lowest were obtained for 1.3 and 3.10 (0.57 and 0.47 respectively). High triplet lifetimes (109-286 μs) were also obtained for all complexes, with 1.7 being the highest (286 μs) which also corresponds to its triplet and singlet quantum yields (0.80 and 0.77 respectively). The photosensitizing properties of low symmetry derivatives, ZnMCapPc and ZnMCafPc were investigated by conjugating glutathione (GSH) capped silver nanoparticles (AgNP). The formation of the amide bond was confirmed by IR and UV-Vis spectroscopies. The photophysicochemical behaviour of the novel phthalocyanine-GSH-AgNP conjugates and the simple mixture of the Ag NPs with low the symmetry phthalocyanines were investigated. It was observed that upon conjugation of the phthalocyanines to the GSH-AgNPs, a blue shift in the Q band was induced. The triplet lifetimes and quantum yields improved upon conjugation as compared to the phthalocyanines (Pc) alone. Complex 1.5 triplet lifetimes increased from 109 to 148 and triplet quantum yield from 0.65 to 0.86 upon conjugation. Fluorescence lifetimes and quantum yields decreased for the conjugates compared to the phthalocyanines alone, due to the quenching caused by the Ag NPs. The antimicrobial activity of the zinc phthalocyanines (complexes 1.3 and 1.5) and their conjugates against Escherichia coli was investigated. Only 1.3 and 1.5 complexes were investigated because of the availability of the sample. In general phthalocyanines showed increase in antibacterial activity with the increase in phthalocyanines concentration in the presence and absence of light. The Pc complexes and their Ag NP conjugates showed an increase in antibacterial activity, due to the synergistic effect afforded by Ag NP and Pcs. Improved antibacterial properties were obtained upon irradiation. 1.5-AgNPs had the highest antibacterial activity compared to 1.3-AgNPs conjugate; these results are in agreement with the photophysical behaviour. This work demonstrates improved photophysicochemical properties of low symm
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