Thermo-chemo-sonic pre-digestion of waste activated sludge for yeast cultivation to extract lipids for biodiesel production.

The low cost biosynthesis of microbial lipids are an efficient feedstock to replace plant based oil for biodiesel production. The present study objective is to explore the effect of thermo-chemo-sonic pre-digestion of municipal Waste Activated Sludge (WAS) to cultivate oleaginous L. starkeyi MTCC-1400 as a model organism to produce high yield biomass and lipid. Higher Suspended Solids (SS) reduction (20 and 15.71%) and Chemical Oxygen Demand (COD) solubilization (27.6 and 22.3%) were achieved at a Specific Energy (SE) input of 5569 kJ/kg for WAS digested with NaOH and KOH, respectively. The maximum biomass of 17.52 g L-1 and lipid 64.3% dwt were attained in NaOH pre-digested sample. The analyzed lipid profile exhibited high content of palmitic acid (45.6%) and oleic acid (38.7%) which are more suitable for biofuel production. Thus, these results strongly motivate the use of pre-digested WAS as an efficient and economical substrate for biodiesel production.

In silico approach for enhancing innate lipid content of Yarrowia lipolytica, by blocking the acyl-CoA oxidase-1 enzyme, using various analogous compounds of lipids.

Yarrowia lipolytica is used as a model in this study to screen the potential candidates for inflating the innate lipid content of the cell. This study focuses on reducing the lipid degradation that occurs by the ß-oxidation process and discursively increasing the innate lipid content. Acyl-CoA oxidase-1, the primary and initial enzyme involved in the lipid degradation pathway, was selected as a target and blocked using various lipid analogous compounds. The blocking study was carried out using molecular docking and dynamic studies using computation tools. The largest active site pocket located around the Phe-394 amino acid of the target protein is taken as a site for docking. The molecular docking was performed for the selected compounds (citric acid, Finsolv, lactic acid, oxalic acid, Tween-80 and Triton X-100) and the docking results were compared with the outcome of the standard molecule (octadecatrienoic acid). Citric acid, Finsolv, Tween-80 and Triton X-100 were found to be the potential candidates for blocking the target molecule in the static condition using docking studies, revealing a minimum binding energy requirement than the standard molecule. They were further taken for a dynamics study using GROMACS software. The RMSD, RMSF, number of hydrogen bond interactions and radius of gyration of the complex molecules were studied in a dynamic approach for 100 ns. Citric acid has been found to be the potential hit compound to block acyl-CoA oxidase-1 enzyme with its maximum hydrogen interaction and minimum fluctuations. It also revealed out the minimum total energy requirement for the complex formation.

A review on contemporary approaches in enhancing the innate lipid content of yeast cell.

For the past few decades, industrialization has made a huge environmental hazard to the world with its waste. The approach of waste to wealth in the recent era has made many Eco-economical suggestions for the industries. The valuable products in biorefinery aspects of the eco-economical suggestions include; energy products, high-value drugs and novel materials. Bio-lipids are found to be the major influencing eco-economical products in the process. Production of bio-lipid from microbial sources has paved the way for future research on lipid-bioproducts. The yeast cell is a unique organism with a large unicellular structure capable of accumulating a high amount of lipids. It constitutes 90% of neutral lipids. Various strategies enhance the lipid profile of yeast cells: usage of oleaginous yeast, usage of low cost (or) alternative substrates, developing stress conditions in the growth medium, using genetically modified yeast, altering metabolic pathways of yeast and by using the symbiotic cultures of yeast with other microbes. The metabolic alterations of lipid pathways such as lipid biosynthesis, lipid elongation, lipid accumulation and lipid degradation have been a striking feature of research in lipid-based microbial work. The lipid-bioproducts have also made a strong footprint in the history of alternative energy products. It includes partial acyl glycerol, oleochemicals, phospholipids and biofuels. This report comprises the recent approaches carried out in the yeast cell for enhancing its lipid content. The limitations, challenges and future scope of individual strategies were also highlighted in this article.

Solubilisation of fruits and vegetable dregs through surfactant mediated sonic disintegration: impact on biomethane potential and energy ratio.

This study investigates the symbiotic effect of cetyltrimethylammonium bromide (CTAB) coupled with sonication of fruits and vegetable dregs (FVD) on disintegration and subsequent energy efficient methane production. The liquefaction of FVD experiments was conducted by varying dosage of surfactant from 0.001to 0.01 g/g SS for 60 min in mechanical shaker. The optimised dosage of surfactant was combined with sonication. Finally, the combined pretreatment and sole pretreatment were assessed using methane potential assay. The results revealed that at optimised conditions (sonication specific energy of 5400 kJ/kg TS, CTAB dosage of 0.006 g/g SS), the maximum liquefiable organics release rate and solids reduction of CTAB mediated sonic disintegration (CSD) were found respectively to be 27% and 17% more than the ultrasonic disintegration (16% and 10%). CSD was noticed to be superior than ultrasonic disintegration (UD) based on highest volatile fatty acid yield (2000 mg/L vs. 1250 mg/L) and biochemical methane potential (203 mL/g COD vs. 144 mL/g COD). CSD achieved energy ratio of 0.9 which is greater than ultrasonic disintegration energy ratio 0.4.

An encapsulated report on enzyme-assisted transesterification with an allusion to lipase.

Biodiesel is a renewable, sulfur-free, toxic-free, and low carbon fuel which possesses enhanced lubricity. Transesterification is the easiest method employed for the production of biodiesel, in which the oil is transformed into biodiesel. Biocatalyst-mediated transesterification is more advantageous than chemical process because of its non-toxic nature, the requirement of mild reaction conditions, absence of saponification, easy product recovery, and production of high-quality biodiesel. Lipases are found to be the primary enzymes in enzyme-mediated transesterification process. Currently, researchers are using lipases as biocatalyst for transesterification. Lipases are extracted from various sources such as plants, microbes, and animals. Biocatalyst-based biodiesel production is not yet commercialized due to high-cost of purified enzymes and higher reaction time for the production process. However, research works are growing in the area of various cost-effective techniques for immobilizing lipase to improve its reusability. And further reduction in the production cost of lipases can be achieved by genetic engineering techniques. The reduction in reaction time can be achieved through ultrasonic-assisted biocatalytic transesterification. Biodiesel production by enzymatic transesterification is affected by many factors. Various methods have been developed to control these factors and improve biodiesel production. This report summarizes the various sources of lipase, various production strategies for lipase and the lipase-mediated transesterification. It is fully focused on the lipase enzyme and its role in biodiesel production. It also covers the detailed explanation of various influencing factors, which affect the lipase-mediated transesterification along with the limitations and scope of lipase in biodiesel production.

Studies on evaluation of surfactant coupled sonication pretreatment on Ulva fasciata (marine macroalgae) for enhanced biohydrogen production.

Biohydrogen production from marine macroalgal biomass by advanced pre-treatment strategies is considered a clean energy technology. The present study focuses on investigating the effects of sonication pre-treatment (SP) and saponin coupled sonic pre-treatment (SSP) on Ulva fasciata for enhancing the production of biohydrogen. The SP and SSP were optimized to improve the hydrolysis process during digestion. The optimized time and sonication power were found respectively as 30 min and 200 W. A high concentration of biopolymer release was noticed in SSP than SP at optimized conditions. The surfactant dosage in SSP was optimized at 0.0036 g/g TS. The effect of SSP process was assessed by estimation of COD (Chemical Oxygen Demand) and SCOD (Soluble Chemical Oxygen Demand) release. The study revealed that, at a specific energy of 36,000 KJ/Kg TS, the SCOD release was higher in SSP (1900 mg/L) than SP (1050 mg/L). The SSP process could improve the COD solubilization to 15 % more than the SP. Carbohydrate and protein release are also more in SSP than SP. The use of biosurfactants significantly reduced the energy utilization in the hydrolysis process. The SSP pre-treated Ulva fasciata biomass has yielded a higher biohydrogen of 91.7 mL/g COD which is higher compared to SP (40.5 mL/g COD) and Control (9 mL/g COD).

Enhancement of ultrasound assisted aqueous extraction of polyphenols from waste fruit peel using dimethyl sulfoxide as surfactant: Assessment of kinetic models.

Pomegranate peel, a major waste from the food processing industries containing biologically active compounds, could be converted into value-added products having medicinal properties. Present study deals with the ultrasound-assisted surfactant, namely dimethyl sulfoxide (DMSO) aided polyphenolics extraction from pomegranate peel waste using double distilled water (DDW) as a solvent. Maximum total yield of extraction and total polyphenolic content (TPC) were found respectively to be 43.58 ± 1.0 and 49.55 ± 0.8%, at optimized sonication parameters viz. temperature 50 °C, power density 1.2 W/mL and time 40 min followed by surfactant aided extraction under optimum conditions 0.6% DMSO, 50 °C and 150 rpm for 90 min. Kinetic models were developed to determine the polyphenolics concentration and validated. GC-MS analysis of the extract revealed 22 phenolic compounds. Thus, the acquired results have ensured the significance of ultrasound pre-treated surfactant aided extraction of polyphenolic compounds and this process can be developed for commercial production.

A novel energetically efficient combinative microwave pretreatment for achieving profitable hydrogen production from marine macro algae (Ulva reticulate).

This study aims to enhance the hydrogen (H2) production from marine macro algae (Ulva Reticulate) by microwave combined with hydrogen peroxide (H2O2) under alkaline condition. Microwave (domestic type) (M) pretreatment of algal biomass at its optimal power (40%) resulted in 27.9% COD solubilization at 15 min time interval. When this optimal microwave power was combined with H2O2 (MH) an increment in COD solubilization was achieved at 24 mg H2O2/g macroalgae dosage. Under alkaline condition (pH 7-12), microwave and H2O2 combination (MHA) yielded better result than MH. At optimal alkaline condition (pH 10), MHA pretreatment shows a COD solubilization of 34%. Microwave in alkaline condition induces decomposition of H2O2 and more OH radical synthesis. This synergistically promotes solubilization. The MHA process considerably diminish time and specific energy required for biomass disintegration. Among the samples, highest H2 yield of 87.5 mL H2/g COD was observed for MHA.

Disperser coupled rhamnolipid disintegration of pulp and paper mill waste biosolid: Characterisation, methane production, energy assessment and cost analysis.

In this study, the disintegration potential of disperser coupled rhamnolipid (RLD) was investigated on pulp and paper mill secondary sedimentation tank (PPST) sludge. Initially, RLD dosage and pH were optimized for liquefied organic content release. Maximal of liquefied organic content release of 2158 mg/L was attained at an optimized RLD dosage (0.009 g/g TS (Total solids), pH 10). To augment liquefaction of PPST sludge further, disperser and disperser coupled RLD methods were carried out. Disperser coupled RLD method has achieved maximal liquefaction rate (27%) and total suspended solids (TSS) reduction (20%) at 5128 kJ/kg TS when compared to disperser method. Subsequently, methane assay was performed in which disperser coupled RLD method yielded higher methane production of 295 mL/g VS (Volatile solids). Then, cost analysis was performed in which disperser coupled RLD method achieved a net profit of 134 $/ ton of PPST sludge.

Investigation on ultrasonication mediated biosurfactant disintegration method in sludge flocs for enhancing hydrolytic enzymes activity and polyhydroxyalkanoates.

In this study, a novel biosurfactant potential bacterial strain Pseudomonas pachastrellae RW43 was isolated from pulp and paper sludge and the biosurfactant namely rhamnolipid produced by Pseudomonas pachastrellae RW43 was investigated by varying pH and incubation time in batch liquid fermentation process. The maximal yield of rhamnolipid was found to be 12.1 g/L at an optimized condition of pH 7 and incubation time of 168 h. NMR analysis was performed for identification of molecular structure of produced rhamnolipid and its results concluded that the product was identified as di rhamnolipid. Then, statistically the global optimum conditions for hydrolytic enzymes extraction parameters (sonication power (100 W), extraction time (15 min) and rhamnolipid dosage (2% v/v)) were established. At 30,456 kJ/kg TS specific energy, ultrasonication with rhamnolipid disintegration method extracted maximal consortium activity of hydrolytic enzymes from mixed sludge (municipal and pulp & paper sludge) and the maximum observed were found to be 42.22, 51.75, 34.26, 24.21, 11.35 Units/g VSS respectively for protease, α-amylase, cellulase, lipase and α-glucosidase. Polyhydroxyalkanoates was recovered from enzymes extracted sludge using various solvents namely chloroform, sodium hypochlorite with chloroform and sodium lauryl sulfate with sodium hypochlorite. The maximum recovery was found to be 74 g/kg using sodium hypochlorite and chloroform extraction solvents.

Study on Lipid Accumulation in Novel Oleaginous Yeast Naganishia liquefaciens NITTS2 Utilizing Pre-digested Municipal Waste Activated Sludge: a Low-cost Feedstock for Biodiesel Production.

The economical production of lipids is considered as an appropriate renewable alternative feedstock for biodiesel production because of the contemporary concerns on fuel crisis, climate change and food security. In this study, lipid accumulation potential of a novel oleaginous yeast isolate Naganishia liquefaciens NITTS2 by utilizing pre-digested municipal waste activated sludge (PWAS) was explored. Optimization of culture conditions was performed using response surface methodology coupled with genetic algorithm and maximum lipid content of 55.7% was obtained. The presence of lipid was visually confirmed by fluorescence microscopy and its characteristic profile was determined by GC-MS. The yeast lipid was recovered and converted into biodiesel by garbage lipase with the efficiency of 88.34 ± 1.2%, which was further analyzed by proton nuclear magnetic resonance spectroscopy. Hence, the results of this study strongly suggest the possibility of using PWAS as an efficient and low-cost resource for the production of biodiesel from the oleaginous yeast.

Effect of surfactant assisted sonic pretreatment on liquefaction of fruits and vegetable residue: Characterization, acidogenesis, biomethane yield and energy ratio.

The present study explored the disintegration potential of fruits and vegetable residue through sodium dodecyl sulphate (SDS) assisted sonic pretreatment (SSP). In SSP method, initially the biomass barrier (lignin) was removed using SDS at different dosage, subsequently it was sonically disintegrated. The effect of SSP were assessed based on dissolved organic release (DOR) of fruits and vegetable waste and specific energy input. SSP method achieved higher DOR rate and suspended solids reduction (26% and 16%) at optimum SDS dosage of 0.035 g/g SS with least specific energy input of 5400 kJ/kg TS compared to ultrasonic pretreatment (UP) (16% and 10%). The impact of fermentation and biomethane potential assay revealed highest production of volatile fatty acid and methane yield in SSP (1950 mg/L, 0.6 g/g COD) than UP. The energy ratio obtained was 0.9 for SSP, indicating proposed method is energetically efficient.

Green synthesis of silver nanoparticles using leaf extract of Acalypha hispida and its application in blood compatibility.

The blood compatibility of AgNPs is of great relevance as it has good antifungal, antibacterial and anti-inflammatory properties and the toxicological information of their effects on cells need to be analyzed before using it as drug carriers in the biomedical field. The present study deals with the synthesis of AgNPs from an aqueous solution of silver nitrate using Acalypha hispida leaf extract as the reducing and capping agent. The presence of AgNPs in the reaction mixture was confirmed by visual observation of color change and subsequently identified using UV-Visible Spectroscopy. XRD results revealed the crystalline nature of synthesized AgNPs. The shape and size of particles were characterized by TEM. These results revealed the elemental status of nanopowder. The components present in leaf extract were identified by GC-MS and functional groups present in the sample when treated with silver nitrate were obtained from FT-IR results. The surface of synthesized AgNPs was modified using four different compounds such as CTAB, PEG, PEI, and APTMS to evaluate the blood compatibility. The results showed that 50 µg/mL CTAB coated AgNPs and 50 and 100 µg/mL PEG coated AgNPs had non-hemolytic property and considered as more blood compatible surface modified AgNPs. This investigation gives an idea of using surface modified AgNPs in the field of biomedicine and therapeutic applications.

Study on optimization of process parameters for enhancing the multi-hydrolytic enzyme activity in garbage enzyme produced from preconsumer organic waste.

The garbage enzymes produced from preconsumer organic waste containing multi hydrolytic enzyme activity which helps to solubilize the waste activated sludge. The continuous production of garbage enzyme and its scaling up process need a globe optimized condition. In present study the effect of fruit peel composition and sonication time on enzyme activity were investigated. Garbage enzyme produced from 6g pineapple peels: 4g citrus peels pre-treated with ultrasound for 20min shows higher hydrolytic enzymes activity. Simultaneously statistical optimization tools were used to model garbage enzyme production with higher activity of amylase, lipase and protease. The maximum activity of amylase, lipase and protease were predicted to be 56.409, 44.039, 74.990U/ml respectively at optimal conditions (pH (6), temperature (37°C), agitation (218 RPM) and fermentation duration (3days)). These optimized conditions can be successfully used for large scale production of garbage enzyme with higher hydrolytic enzyme activity.

Economical synthesis of silver nanoparticles using leaf extract of Acalypha hispida and its application in the detection of Mn(II) ions.

This study was focused on the synthesis of silver nanoparticles using Acalypha hispida leaf extract and the characterization of the particles using UV-Vis spectroscopy, XRD, FT-IR, and TEM. The results showed the formation of silver nanoparticles, crystalline in nature, with an average size of 20-50 nm. The leaf extract components were analyzed with GC-MS and exhibited a high content of Phytol (40.52%), n-Hexadecanoic acid (9.67%), 1,2,3-Benzenetriol (7.04%), α-d-Mannofuranoside methyl (6.22%), and d-Allose (4.45%). The optimization and statistical investigation of reaction parameters were studied and maximum yield with suitable properties of silver nanoparticles was obtained at leaf extract volume (0.5 mL), the concentration of silver nitrate (1.75 mM), and reaction temperature (50 °C). The method of detecting Mn2+ ions using the colloidal silver nanoparticles was discussed. The minimum and maximum detection limit were found to be 50 and 200 µM of Mn(II) ions, respectively. Thus, the obtained results encourage the use of economical synthesis of silver nanoparticles in the development of nanosensors to detect the pollutants present in industrial effluents.

Identification and optimization of parameters for the semi-continuous production of garbage enzyme from pre-consumer organic waste by green RP-HPLC method.

Reuse and management of organic solid waste, reduce the environmental impact on human health and increase the economic status by generating valuable products for current and novel applications. Garbage enzyme is one such product produced from fermentation of organic solid waste and it can be used as liquid fertilizer, antimicrobial agents, treatment of domestic wastewater, municipal and industrial sludge treatment, etc. The semi-continuous production of garbage enzyme in large quantity at minimal time period and at lesser cost is needed to cater for treatment of increasing quantities of industrial waste activated sludge. This necessitates a parameter for monitoring and control for the scaling up of current process on semi-continuous basis. In the present study a RP-HPLC (Reversed Phase-High Performance Liquid Chromatography) method is used for quantification of standard organic acid at optimized condition 30°C column oven temperature, pH 2.7, and 0.7 ml/min flow rate of the mobile phase (potassium dihydrogen phosphate in water) at 50mM concentration. The garbage enzyme solution collected in 15, 30, 45, 60, 75 and 90 days were used as sample to determine the concentration of organic acid. Among these, 90th day sample showed the maximum concentration of 78.14 g/l of acetic acid in garbage enzyme, whereas other organic acids concentration got decreased when compare to the 15th day sample. This result confirms that the matured garbage enzyme contains a higher concentration of acetic acid and thus it can be used as a monitoring parameter for semi-continuous production of garbage enzyme in large scale.

HE2beta and HE2gamma, new members of an epididymis-specific family of androgen-regulated proteins in the human.

HE2 is an epididymis-specific sperm-binding secretory protein. We isolated a family of HE2-related complementary DNAs from a human caput/corpus library. The transcripts code for identical 71-amino acid N-termini and different C-termini, and 5'- and 3'-untranslated regions. Compared with the original HE2, HE2beta and HE2gamma proteins have a 25-amino acid deletion near the C-terminus, and HE2gamma isoforms have a second deletion. These frame-shifting deletions result in C-termini differing in length, amino acid sequence, including number of cysteines, and isoelectric point. Identical sequences and deletion start and stop points indicate the HE2 isoforms are derived from alternative splicing of 8 or more exons of a single gene. Northern hybridization revealed that the 0.9-kb messenger RNA (mRNA) is most abundant in human caput; there is much less of it (20%) in corpus and little (<5%) in cauda. In castrated Macaca mulatta, HE2 mRNA decreased to 10% of sham-operated levels. Testosterone replacement maintained HE2 mRNA 3- to 5-fold higher than castrate levels, indicating its androgen dependence. Immunohistochemical staining revealed that the beta1 form is highly expressed in principal cells of the initial segment and caput. It is secreted into the lumen and binds to the sperm surface in the postacrosomal and neck regions. The beta2 form is expressed in principal cells primarily in efferent ducts.

Primate epididymis-specific proteins: characterization of ESC42, a novel protein containing a trefoil-like motif in monkey and human.

Epididymal secreted proteins promote sperm maturation and fertilizing capacity by interacting with sperm during passage through the epididymis. Here we investigate the molecular basis of sperm maturation by isolating cDNA clones for novel epididymis-specific expressed sequences. Thirty-six novel cDNAs were isolated and sequenced from a subtracted Macaca mulatta epididymis library. The clones encode proteins with a range of motifs characteristic of protein-modifying enzymes, protease inhibitors, hydrophobic ligand-binding and transport proteins, extracellular matrix-interacting proteins, and transcription regulatory factors. The full length coding sequences were obtained for 11 clones representing a range of abundance levels. Expression of each is regionally localized and androgen regulated. The most abundant, ESC42, contains a cysteine-rich region similar to the signature binding domain of the trefoil family of motogenic wound repair proteins. The monkey and human proteins are nearly 90% identical. Immunohistochemical staining revealed that the protein is most abundant in the epithelium of the caput and is also present in the lumen and bound to sperm. The ESC42 gene, located on chromosome 20q11, contains two exons encoding two nearly identical predicted signal peptides and a third exon encoding the rest of the protein.

Cystatin 11: a new member of the cystatin type 2 family.

Cystatin (CST)11, a novel member of the CST type 2 family of cysteine protease inhibitors, was identified in Macaca mulatta epididymis by subtractive hybridization cloning. The human CST11 gene on chromosome 20p11.2 is located near three other CST genes expressed predominantly in the male reproductive tract. The CST11 gene spans three exons, a structure similar to that of other CST family 2 genes. An exon 2-deleted alternative transcript (CST11Delta2) was also identified. CST11 mRNA is expressed only in the epididymis as judged by Northern blot hybridization and is androgen regulated. The protein is most abundant in the initial segment, but is detected throughout the epididymis and on ejaculated human sperm. The calculated tertiary structure of CST11 reveals that the three regions corresponding to the protease inhibitory wedge of CST3 are similarly juxtaposed in CST11, consistent with protease inhibitor function. Intact and exon 2-deleted CST11 recombinant proteins were tested for antibacterial activity. After a 2-h incubation of Escherichia coli with 50 microg/ml recombinant CST11 or CST11Delta2, bacterial colony-forming units were reduced to 30% of control, indicating that both forms have antimicrobial activity.