Method For Preparation Of Extracts Research Articles

Estimating Recovery in the Evaporation Chemical Space

Evaporation is a commonly used extract preparation technique in chemical characterization testing which can greatly impact the observed extractables. Although ideally only solvent is removed and the analytes remain, in practice the analytes evaporate as well, and these losses must be considered when addressing the appropriateness of evaporation as an extract preparation method. This issue is exacerbated when using evaporation before non-targeted analysis (NTA), where the identities of the analytes are not known prior to analysis. Herein, a means of evaluating the extent of evaporative loss in NTA is presented. To achieve this, a model for evaporative loss was adopted and verified experimentally, then applied to a relevant chemical space to determine the impact of experimental parameter selection. The model was used to predict recovery for a range of chemicals under a variety of experimental conditions (volume change, solvent, and temperature). The test solvents were selected to include a range of relevant properties, including molecular weight, density, and vapor pressure. The root-mean-square error of 70 evaporation recovery conditions was 12%. Excess losses, beyond the scope of this model, were found when samples were evaporated to dryness. Also, high recovery was demonstrated for select chemicals (with high air-solvent partition coefficients) at low concentration and with large volume changes. Overall, good agreement was found between the evaporation recovery model and experimental measurements, suggesting that the model can be used to predict the effect on a NTA.

Plant mediated fabrication of silver nanoparticles, process optimization, and impact on tomato plant

Nanotechnology is one of the fastest-growing markets, but developing eco-friendly products, their maximum production, stability, and higher yield is a challenge. In this study, silver nanoparticles were synthesized using an easily available resource, leaves extract of the Neem (Azadirachta indica) plant, as a reducing and capping agent, determined their effect on germination and growth of tomato plants. The maximum production of silver nanoparticles was noted at 70 °C after 3 h of reaction time while treating the 10 ml leaf extract of Neem plant with 10 ml of 1 mM silver nitrate. The impact of the extract preparation method and solvent type on the plant mediated fabrication of silver nanoparticles was also investigated. The UV-spectrophotometric analysis confirmed the synthesis of silver nanoparticles and showed an absorption spectrum within Δ420–440 nm range. The size of the fabricated silver nanoparticles was 22–30 nm. The functional groups such as ethylene, amide, carbonyl, methoxy, alcohol, and phenol attached to stabilize the nanoparticles were observed using the FTIR technique. SEM, EDX, and XRD analyses were performed to study the physiochemical characteristics of synthesized nanoparticles. Silver nanoparticles increased the germination rate of tomato seeds up to 70% while decreasing the mean germination time compared to the control. Silver nanoparticles applied at varying concentrations significantly increased the shoot length (25 to 80%), root length (10 to 60%), and fresh biomass (10 to 80%) biomass of the tomato plant. The production of total chlorophyll, carotenoid, flavonoids, soluble sugar, and protein was significantly increased in tomato plants treated with 5 and 10 ppm silver nanoparticles compared to the control. Green synthesized silver nanoparticles are cost-effective and nontoxic and can be applied in agriculture, biomedical, and other fields.

Mechanistic Insights into Cell-Free Gene Expression through an Integrated -Omics Analysis of Extract Processing Methods.

Cell-free systems derived from crude cell extracts have developed into tools for gene expression, with applications in prototyping, biosensing, and protein production. Key to the development of these systems is optimization of cell extract preparation methods. However, the applied nature of these optimizations often limits investigation into the complex nature of the extracts themselves, which contain thousands of proteins and reaction networks with hundreds of metabolites. Here, we sought to uncover the black box of proteins and metabolites in Escherichia coli cell-free reactions based on different extract preparation methods. We assess changes in transcription and translation activity from σ70 promoters in extracts prepared with acetate or glutamate buffer and the common post-lysis processing steps of a runoff incubation and dialysis. We then utilize proteomic and metabolomic analyses to uncover potential mechanisms behind these changes in gene expression, highlighting the impact of cold shock-like proteins and the role of buffer composition.

Comparison of Phenolic Contents and Antioxidant Activities for Black and White Desert Truffles Spread in Syria

Desert truffle is considered as a type of Syrian wild fungi that spreads heavily, and it occupies important rank in folk medicine, where its aqueous extract is used for the treatment of some eye and skin illnesses, and people prefer the use of black truffle. This work interested in studying of the most available species; Terfezia claveryi (black) and Tirmania pinoyi (white). The extracts of the two species of truffle were prepared by maceration with water, methanol, and ethanol 70%. Their total phenolic contents (TPC) and total flavonoid contents (TFC) were analyzed using Folin-ciocalteu and Aluminum chloride methods respectively, and their antioxidant activities was tested using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and Ferric Reducing Antioxidant Power (FRAP) methods, after microscopic examination and detection of phytochemical components. Then, phenolic profile of ethanolic 70% extract of black truffle T.claveryi was studied by using LC-MS/MS. The values of TPC were between 25.3-43.6 mg GAE/g dry extract and TFC were between 2.5-6.8 mg QE/g dry extract. The values of DPPH (IC50) and FRAP were between 5.6-9.0 mg/ml and 90.1-153.4 µmol AAE/g dry extract respectively. There is a great similarity in content and activity of two species, also the aqueous extract is similar to other extracts in content and activity, and this means that the method of extract preparation in traditional medicine is reliable. It has been predicted about 14 phenolic compounds in the extract; as p-Hydroxy benzoic acid, Syringic acid and trans-Cinnamic acid. As a result, both truffle species are a new rich resource of antioxidant compounds which are usable in nutritional, cosmetic, and therapeutic applications.

Development of Standardized Cyanthillium cinereum (L.) H.Rob. Extract and Determination of its Biological Activities

Aim: The development of a suitable standardized Cyanthillium cinereum (L.) H.Rob. extract is an active ingredient in healthcare products. Background: C. cinereum is the herbal tea specified in the Thai National List of Essential (herbal) Medicines (NLEM) as the most efficacious tea for smoking cessation. However, herbal tea is inconvenient, and no standardized C. cinereum extraction method was known. Objectives: The study aims to develop a standardized C. cinereum extract preparation method and determine its biological activities. Methods: Various extraction and fractionation methods were performed in order to optimize a suitable standardized extract. The extraction yield, biological activities, and biomarkers (apigenin and luteolin) of the HPLC method were used to select the most suitable extraction method. Results: The results showed that the microwave-assisted extraction (MAE) method with 75% EtOH was the most suitable method. The MAE method obtained apigenin and luteolin at 0.320±0.029 and 0.487±0.012 mg/g, respectively. MAE showed good anti-inflammatory and antioxidant (DPPH and FRAP assay) activities. Subsequently, the MAE extract was fractionated by the Diaion® HP-20 column in order to obtain the most suitable standardized extract. The 50% plus 75% EtOH fractions showed high apigenin (91.20±1.23 mg/g) and luteolin (167.00±0.49 mg/g) contents and exerted potent bioactivities. The standardized C. cinereum extract presented high effectiveness of NO inhibitory activity with an IC50 of 7.88±3.56 μg/mL, and also exerted DPPH scavenging efficacy with an IC50 value of 8.88±0.17 μg/mL and quercetin equivalent at 137.50±2.20 mg/g by FRAP assay. Conclusion : This study succeeded in developing a high-yield extraction method of standardized C. cinereum extract, with potent antioxidant and anti-inflammatory activities, suitable for various purposes.

Simple Extract Preparation Methods for E. coli-Based Cell-Free Expression.

Cell-free protein synthesis (CFPS) is a powerful platform for synthetic biology, allowing for the controlled expression of proteins without reliance on living cells. However, the process of producing the cell extract, a key component of cell-free reactions, can be a bottleneck for new users to adopt CFPS as it requires technical knowledge and significant researcher oversight. Here, we provide a detailed method for implementing a simplified cell extract preparation workflow using CFAI media. We also provide a detailed protocol for the alternative, 2x YPTG media-based preparation process, as it represents a useful benchmark within the cell-free community.

Evaluation of local Origanum vulgare aqueous extract for eradication of biofilm production bacteria

Origanum vulgareleaves are collected from a local garden, Baghdad, Iraq. The hot and cold aqueous extract was prepared to perform an In vitro antibacterial and antibiofilm actions .Antibacterial activities on various antibiotic resistant bacteria (Pseudomonas aeruginosa, Klebsiella pneumoniae, Echerichia coli, Staphylococcus aurous and staphylococcus epidermidis) were tested by well diffusion method. The biofilm-forming ability of the tested microorganisms was determined by the Microtiter-plate method. Fresh hot extract showed the highest effect and the dried cold extract had the lowest effect against gram-negative and gram-positive bacteria . fresh hot extract decrease significantly the biofilm formation and showed good ability as an antibiofilm to all strains that tested..Antimicrobial activity of the extract seemed to be related to the aqueous extract preparation method and that appeared by the difference between fresh and dried leave extract activity against pathogens and it can be used as potential antibiofilm agent against the biofilm forming strains.

Evaluation of Microwave-Assisted Extraction Method for Preparation and Assessment of Thai Herbal Medicine Oral Tablets With Enriched Phytochemical Compounds.

In developing countries, populations have employed herbal medicines for primary health care because they are believed to be more appropriate to the human body and have less side effects than chemically synthesized drugs. The present study aimed to develop and evaluate herbal tablets incorporated with a Thai traditional medicinal extract, U-pa-ri-waat (URW), using microwave-assisted extraction (MAE). The extraction efficiency for URW using MAE and traditional solvent extraction was compared based on the percent yield after spray drying. URW tablets were prepared using the dry granulation method. The optimized products were assessed using standard characterization methods based on the United States and British Pharmacopeias. DPPH and ABTS radical scavenging assays were performed to analyze the antioxidant capacity of the microwave-assisted extracts. The results revealed that the flowability of the dry granule with added maltodextrin was improved compared to a granule without additives, as indicated by an angle of repose of 33.69 ± 2.0°, a compressibility index of 15.38 ± 0.66, and a Hausner's ratio of 1.18 ± 0.06. The resulting formulation produced flat tablets with uniform weight variation, hardness, thickness, friability, and optimum disintegration time. The URW extracts showed antioxidant activity and MAE with maltodextrin carrier displayed the strongest DPPH and ABTS radical activities with IC50 values of 1.60 ± 0.02 μg/mL and 4.02 ± 0.24 μg/mL, respectively. The URW tablet formulation passed the quality control tests. Storage of the formulation tablets for 90 days under accelerated conditions had minimal effects on tablet characteristics.

Protein Extract Preparation and Co-immunoprecipitation from Caenorhabditis elegans.

Co-immunoprecipitation methods are frequently used to study protein-protein interactions. Confirmation of hypothesized protein-protein interactions or identification of new ones can provide invaluable information about the function of a protein of interest. Some of the traditional methods for extract preparation frequently require labor-intensive and time-consuming techniques. Here, a modified extract preparation protocol using a bead mill homogenizer and metal beads is described as a rapid alternative to traditional protein preparation methods. This extract preparation method is compatible with downstream co-immunoprecipitation studies. As an example, the method was used to successfully co-immunoprecipitateC. elegans microRNA Argonaute ALG-1 and two known ALG-1 interactors: AIN-1, and HRPK-1. This protocol includes descriptions of animal sample collection, extract preparation, extract clarification, and protein immunoprecipitation. The described protocol can be adapted to test for interactions between any two or more endogenous, endogenously tagged, or overexpressed C. elegans proteins in a variety of genetic backgrounds.

Evaluation of the Effects of Allelopathic Aqueous Plant Extracts, as Potential Preparations for Seed Dressing, on the Modulation of Cauliflower Seed Germination

Allelopathic plants can be widely used in bio-farming considering their potential role in the improvement of seed germination. The study presented in this work was designed to evaluate the effects of two extraction methods and the effects of allelopathic aqueous extracts from twenty plants as seed dressing preparations on the number of germinating and infested seeds of cauliflower (Brassica oleracea convarietas L. botrytis var. botrytis). Plant extracts (in the form of cold-soaked macerates and infusions) were used for seed dressing. The percentages of normally germinating, non-germinating, and pathogen-infested seeds were determined in a paper test. Of the 20 herbal plant species used in the study, the biopreparations extracted from Zea mays L. moles were the most effective as they evoked the most beneficial effects on both seed germination and reduction of infestation by microbial pathogens. The study also showed that infusions used for seed treatment were better at improving cauliflower seed germination than were macerates. This method of extract preparation probably enabled an increase both in the availability and activity of allelochemical compounds.

The prospect of using guava leaf extract for biosynthesizing chitosan nanoparticles

Chitosan is polysaccharide polymer used for biomedical applications and food packaging. The aim of this investigation is to produce chitosan nanoparticles (ChNPs) using guava leaf (Psidium guajava L.) extract as green synthesis of nanoparticles using plant leaf extract is ecofriendly and cost-effective approach. The leaves of guava are nontoxic, full of bioactive compounds, and suitable alternative to avoid chemicals used in other methods. Variations in extract preparation method and ratio of chitosan to leaf extract were examined to investigate the effect of these factors on the particles size. The characterisation of chitosan nanoparticles was conducted using dynamic light scattering (DLS), transmission electron microscope (TEM), Fourier transform infrared spectroscopy (FTIR), and zeta potential analysis. The results showed semi-spherical nanoparticles were formed at pH = 5 with an average particle size of 21.92 nm and narrow size distribution as, polydispersity index (PDI) value was 0.471. Zeta potential was −27.1 which gives reasonable stability to the nanoparticles in suspension. The effect of storage time of chitosan nanoparticles on the antibacterial activity against Staphylococcus aureus, Escherichia coli, and Bacillus subtilis was also investigated.

Glycerolic Licorice Extracts as Active Cosmeceutical Ingredients: Extraction Optimization, Chemical Characterization, and Biological Activity.

A green ultrasound-assisted extraction (UAE) method using glycerol/water mixtures for extraction of licorice (Glycyrrhiza glabra) bioactive constituents was developed in this study. The response surface method, according to the Box-Behnken design, was employed to optimize the extraction parameters: glycerol concentration (X1), temperature (X2), and the amount of herbal drug used in the production (X3). The responses were content of total phenols (TP), TP extraction efficiency (TPy) and the content of licorice characteristic constituents, glabridin (Gla) and isoliquiritigenin (Iso). Response surface analysis predicted the optimal extraction conditions for maximized amounts of TP, Tpy, Gla, and Iso. The extracts were prepared using the calculated conditions. The analysis of the selected constituents confirmed the validity of the model. Furthermore, biological activity of the extracts was tested. The results demonstrate that UAE using glycerol is a fast and efficient method for preparation of extracts with excellent radical scavenging, Fe2+ chelating and antioxidant activity. Furthermore, the observed notable tyrosinase and elastase inhibitory activity of the extracts, as well as their anti-inflammatory activity, indicate the anti-aging properties of the investigated extracts. The fact that the extracts were prepared using the safe, cosmetically active solvent, glycerol, makes them suitable for direct use in specialized cosmeceutical formulations.

Deconstructing Cell-Free Extract Preparation for in Vitro Activation of Transcriptional Genetic Circuitry.

Recent advances in cell-free gene expression (CFE) systems have enabled their use for a host of synthetic biology applications, particularly for rapid prototyping of genetic circuits and biosensors. Despite the proliferation of cell-free protein synthesis platforms, the large number of currently existing protocols for making CFE extracts muddles the collective understanding of how the extract preparation method affects its functionality. A key aspect of extract performance relevant to many applications is the activity of the native host transcriptional machinery that can mediate protein synthesis. However, protein yields from genes transcribed in vitro by the native Escherichia coli RNA polymerase are variable for different extract preparation techniques, and specifically low in some conventional crude extracts originally optimized for expression by the bacteriophage transcriptional machinery. Here, we show that cell-free expression of genes under bacterial σ70 promoters is constrained by the rate of transcription in crude extracts, and that processing the extract with a ribosomal runoff reaction and subsequent dialysis alleviates this constraint. Surprisingly, these processing steps only enhance protein synthesis in genes under native regulation, indicating that the translation rate is unaffected. We further investigate the role of other common extract preparation process variants on extract performance and demonstrate that bacterial transcription is inhibited by including glucose in the growth culture but is unaffected by flash-freezing the cell pellet prior to lysis. Our final streamlined and detailed protocol for preparing extract by sonication generates extract that facilitates expression from a diverse set of sensing modalities including protein and RNA regulators. We anticipate that this work will clarify the methodology for generating CFE extracts that are active for biosensing using native transcriptional machinery and will encourage the further proliferation of cell-free gene expression technology for new applications.

Process optimization for scalable E. coli extract preparation for cell-free protein synthesis

Cell-free protein synthesis (CFPS) is a versatile tool for protein research and biomanufacturing. In this work, large volume cell extract preparation methods from Escherichia coli were optimized with a unit operations approach. Using face centered cubic designed experiments, optimized time points for IPTG induction (to produce T7 polymerase) and cell harvest were determined. These were at times corresponding to 28 and 58.5% the overall growth extent as measured by absorption; for 1 L cultures in 2.5 L shake flasks in our incubator, this amounted to 201 min and 255 min respectively. The cell pellet was washed in S30 buffer once for 10 min and stored at −80°C without flash freezing in nitrogen. Cell lysis was simplified by diluting the cells to one part S30 buffer, one part DI water, and one part cells by mass and then passing through a continuous French press for one cycle at 25,000 psi. The resulting suspension is clarified with one low speed centrifugation step at 12,000×g for 10 min (4°C). All extract is then immediately frozen in a pilot-scale lyophilizer (−70°C) and dried under optimal conditions determined as 4 h under vacuum (200 mTorr) while holding a shelf temperature of 15°C. The lyophilized extract showed no statistically significant difference in sfGFP expression levels when compared to an optimized, lower-volume sonication protocol and improved performance over extract from commercial kits. As an increasing number of efforts in CFPS expand beyond the bench top, scalability and reliability become increasingly important if it is to become a viable tool for biomanufacturing.

Экстракты Serenoa repens в лечении доброкачественной гиперплазии предстательной железы и симптомов нижних мочевыводящих путей

This literature review focuses on the use of Serenoa repens extracts in the treatment of benign prostatic hyperplasia (BPH) and lower urinary tract symptoms (LUTS). Extracts of Serenoa repens produce marked anti-inflammatory, anti-androgenic and antiproliferative effects. Various extracts differ in their activity depending on the concentration of free fatty acids and the method of extract preparation. Extracts of Serenoa Repens from different manufacturers vary considerably in composition, effectiveness and supporting evidence. Evaluation of the effectiveness of Serenoa Repens extracts should be carried out for each extract separately, depending on the method of preparation and brand. The most studied and clinically effective is the hexane lipidosterol extract of Serenoa Repens Permixon. This assessment is supported by a report on Serenoa Repens, issued by the European Medicines Agency. In this report, only Serenoa repens hexanic extract Permixon is considered to be a well-established medical use product for the symptomatic treatment of BPH.

Herb-Drug Interactions in Neurological Disorders: A Critical Appraisal.

Herbal drugs are being used worldwide in a variety of debilitating neurological and psychiatric disorders such as cerebrovascular accident, Alzheimer's disease, Parkinson's disease and schizophrenia. However, unlike drugs of modern medicine, herbal drugs are complex products containing multiple pharmacologically active constituents. The nature and relative amounts of these constituents vary due to diverse factors such as but not limited to source of the plant(s), local environmental conditions, parts of the plant used, storage, method of extract preparation, accidental contamination or intentional adulteration. Further, they are handled by the human body like modern drugs and subjected to the processes of absorption, distribution, metabolism and excretion. In each of these processes, they can potentially interact with modern drugs due to sharing of similar transport proteins, metabolizing cytochrome P450 (CYP450) enzymes and uptake / efflux pumps. Moreover, herbal drugs can also inhibit or induce CYP450 enzymes or inactivate transporters leading to Herb-Drug interactions (HDIs). In this narrative review, we have analyzed the clinically reported as well as potential HDIs between 10 common herbal drugs viz. Ginkgo, Ginseng, St. John's Wort, Grapefruit, Black and Long Pepper, Curcumin, Brahmi, Kava, Garlic and Valerian and modern medicines used in neurological and psychiatric disorders with their proven or postulated underlying mechanism(s). Though a number of potential pharmacokinetic and/or pharmacodynamic HDIs have been examined, clinically significant alteration of response to modern medicines and/or serious adverse effects are apparently scarce except for Grapefruit and St. John's Wort. Physicians and patients should exercise caution when using herbal drugs and modern medicines concomitantly so that the recognized serious HDIs can be avoided.

Effects ofWithania somniferaon Reproductive System: A Systematic Review of the Available Evidence

Introduction Withania somnifera (WS) also known as ashwagandha is a well-known medicinal plant used in traditional medicine in many countries for infertility treatment. The present study was aimed at systemically reviewing therapeutic effects of WS on the reproductive system. Methods This systematic review study was designed in 2016. Required data were obtained from PubMed, Scopus, Google Scholar, Cochrane Library, Science Direct, Web of Knowledge, Web of Science, and manual search of articles, grey literature, reference checking, and expert contact. Results WS was found to improve reproductive system function by many ways. WS extract decreased infertility among male subjects, due to the enhancement in semen quality which is proposed due to the enhanced enzymatic activity in seminal plasma and decreasing oxidative stress. Also, WS extract improved luteinizing hormone and follicular stimulating hormone balance leading to folliculogenesis and increased gonadal weight, although some animal studies had concluded that WS had reversible spermicidal and infertilizing effects in male subjects. Conclusion WS was found to enhance spermatogenesis and sperm related indices in male and sexual behaviors in female. But, according to some available evidences for spermicidal features, further studies should focus on the extract preparation method and also dosage used in their study protocols.

Allelopathy of aqueous Pachyrhizus erosus L. extracts on Euphorbia heterophylla and Bidens pilosa

ABSTRACT Allelopathy contributes to the sustainable management of weeds in growing areas, due to its ability to inhibit the development of weed species. This study aimed to evaluate the allelopathic effect of Pachyrhizus erosus extract on Euphorbia heterophylla and Bidens pilosa germination. The experiment was carried out in a completely randomized design, in a 2 x 2 x 4 factorial scheme, with four replications. The first factor corresponded to the plant portion (shoot and tuberous root), the second to the extract preparation method (water at room temperature and boiling) and the third to the extract concentrations (0 %, 25 %, 50 % and 100 %). Phytochemical analyses were performed to identify the presence of secondary metabolites. The variables germination percentage, germination rate index and dry biomass decreased with increasing aqueous extract concentrations for both E. heterophylla and B. pilosa. When applied a P. erosus crude shoot extract (boiling), decreases of 18 % and 92 % were observed in the E. heterophylla and B. pilosa germination, respectively, with a decrease of 91 % in the dry biomass of E. heterophylla seedlings. The tuberous root extract (boiling), on the other hand, reduced the germination percentages for E. heterophylla and B. pilosa in 18 % and 80 %, respectively. The phytochemical evaluation indicated the presence of flavonoids, coumarins and saponins in the shoot and tuberous root extracts, while tannins were only observed in the shoot extract. Thus, it can be concluded that P. erosus extracts display an allelopathic effect on the weed species evaluated.

Cell-free protein synthesis from non-growing, stressed Escherichia coli

Cell-free protein synthesis is a versatile protein production system. Performance of the protein synthesis depends on highly active cytoplasmic extracts. Extracts from E. coli are believed to work best; they are routinely obtained from exponential growing cells, aiming to capture the most active translation system. Here, we report an active cell-free protein synthesis system derived from cells harvested at non-growth, stressed conditions. We found a downshift of ribosomes and proteins. However, a characterization revealed that the stoichiometry of ribosomes and key translation factors was conserved, pointing to a fully intact translation system. This was emphasized by synthesis rates, which were comparable to those of systems obtained from fast-growing cells. Our approach is less laborious than traditional extract preparation methods and multiplies the yield of extract per cultivation. This simplified growth protocol has the potential to attract new entrants to cell-free protein synthesis and to broaden the pool of applications. In this respect, a translation system originating from heat stressed, non-growing E. coli enabled an extension of endogenous transcription units. This was demonstrated by the sigma factor depending activation of parallel transcription. Our cell-free expression platform adds to the existing versatility of cell-free translation systems and presents a tool for cell-free biology.

Cell-Free Synthesis of Macromolecular Complexes.

Cell-free protein synthesis based on E. coli cell extracts has been described for the first time more than 50 years ago. To date, cell-free synthesis is widely used for the preparation of toxic proteins, for studies of the translation process and its regulation as well as for the incorporation of artificial or labeled amino acids into a polypeptide chain. Many efforts have been directed towards establishing cell-free expression as a standard method for gene expression, with limited success. In this chapter we will describe the state-of-the-art of cell-free expression, extract preparation methods and recent examples for successful applications of cell-free synthesis of macromolecular complexes.