scholarly journals A cell-free synthetic biochemistry platform for raspberry ketone production

2017 ◽  
Author(s):  
Simon J Moore ◽  
Tomasso Tosi ◽  
David Bell ◽  
Yonek B Hleba ◽  
Karen M Polizzi ◽  
...  
Keyword(s):  
Market Price ◽  
Chemical Extraction ◽  
Industrial Processing ◽  
Raspberry Ketone ◽  
Sports Industry ◽  
A Cell ◽  
Cost Efficient ◽  
Green Solution ◽  
Engineering Constraints ◽  
Natural Additive

AbstractCell-free synthetic biochemistry provides a green solution to replace traditional petroleum or agricultural based methods for production of fine chemicals. 4-(4-hydroxyphenyl)-butan-2-one, also known as raspberry ketone, is the major fragrance component of raspberry fruit and is utilised as a natural additive in the food and sports industry. Current industrial processing standards involve chemical extraction with a yield of 1-4 mg per kilo of fruit. As such its market price can fluctuate up to $20,000 per kg. Metabolic engineering approaches to synthesise this molecule by microbial fermentation have only resulted in low yields of up to 5 mg L−1. In contrast, cell-free synthetic biochemistry offers an intriguing compromise to the engineering constraints provided by the living cell. Using purified enzymes or a two-step semisynthetic route, an optimised pathway was formed for raspberry ketone synthesis leading up to 100% yield conversion. The semi-synthetic route is potentially scalable and cost-efficient for industrial synthesis of raspberry ketone.

scholarly journals High-Yield ‘One-Pot’ Biosynthesis of Raspberry Ketone, a High-Value Fine Chemical

2021 ◽  
Author(s):  
Simon J Moore ◽  
Tommaso Tosi ◽  
David Bell ◽  
Yonek B Hleba ◽  
Karen M Polizzi ◽  
...  
Keyword(s):  
Value Added ◽  
Microbial Production ◽  
High Yield ◽  
Chemical Extraction ◽  
One Pot ◽  
Natural Form ◽  
Strong Activity ◽  
Industrial Processing ◽  
Cofactor Specificity ◽  
Raspberry Ketone

Abstract Cell-free extract and purified enzyme-based systems provide an attractive solution to study biosynthetic strategies towards a range of chemicals. 4-(4-hydroxyphenyl)-butan-2-one, also known as raspberry ketone, is the major fragrance component of raspberry fruit and is used as a natural additive in the food and sports industry. Current industrial processing of the natural form of raspberry ketone involves chemical extraction with a yield of ~1-4 mg kg-1 of fruit. Due to toxicity, microbial production provides only low yields of up to 5-100 mg L-1. Herein, we report an efficient cell-free strategy to probe a synthetic enzyme pathway that converts either L-tyrosine or the precursor, 4-(4-hydroxyphenyl)-buten-2-one (HBA), into raspberry ketone at up to 100% conversion. As part of this strategy, it is essential to recycle inexpensive cofactors. Specifically, the final enzyme step in the pathway is catalysed by raspberry ketone/zingerone synthase (RZS1), an NADPH-dependent double bond reductase. To relax cofactor specificity towards NADH, the preferred cofactor for cell-free biosynthesis, we identify a variant (G191D) with strong activity with NADH. We implement the RZS1 G191D variant within a ‘one-pot’ cell-free reaction to produce raspberry ketone at high-yield (61 mg L-1), which provides an alternative route to traditional microbial production. In conclusion, our cell-free strategy complements the growing interest in engineering synthetic enzyme cascades towards industrially relevant value-added chemicals.

scholarly journals Identification of a Second Lipopolysaccharide in Porphyromonas gingivalis W50

2008 ◽  
Vol 190 (8) ◽  
pp. 2920-2932 ◽  
Author(s):  
Minnie Rangarajan ◽  
Joseph Aduse-Opoku ◽  
Nikolay Paramonov ◽  
Ahmed Hashim ◽  
Nagihan Bostanci ◽  
...  
Keyword(s):  
Porphyromonas Gingivalis ◽  
Lipid A ◽  
Fatty Acid Analysis ◽  
Chemical Extraction ◽  
Resonance Spectroscopy ◽  
Cell Integrity ◽  
O Antigen ◽  
Flight Mass Spectrometry ◽  
Anchoring System ◽  
A Cell

ABSTRACT We previously described a cell surface anionic polysaccharide (APS) in Porphyromonas gingivalis that is required for cell integrity and serum resistance. APS is a phosphorylated branched mannan that shares a common epitope with posttranslational additions to some of the Arg-gingipains. This study aimed to determine the mechanism of anchoring of APS to the surface of P. gingivalis. APS was purified on concanavalin A affinity columns to minimize the loss of the anchoring system that occurred during chemical extraction. 1H nuclear magnetic resonance spectroscopy of the lectin-purified APS confirmed the previous structure but also revealed additional signals that suggested the presence of a lipid A. This was confirmed by fatty acid analysis of the APS and matrix-assisted laser desorption ionization-time of flight mass spectrometry of the lipid A released by treatment with sodium acetate buffer (pH 4.5). Hence, P. gingivalis synthesizes two distinct lipopolysaccharide (LPS) macromolecules containing different glycan repeating units: O-LPS (with O-antigen tetrasaccharide repeating units) and A-LPS (with APS repeating units). Nonphosphorylated penta-acylated and nonphosphorylated tetra-acylated species were detected in lipid A from P. gingivalis total LPS and in lipid A from A-LPS. These lipid A species were unique to lipid A derived from A-LPS. Biological assays demonstrated a reduced proinflammatory activity of A-LPS compared to that of total LPS. Inactivation of a putative O-antigen ligase (waaL) at PG1051, which is required for the final step of LPS biosynthesis, abolished the linkage of both the O antigen and APS to the lipid A core of O-LPS and A-LPS, respectively, suggesting that WaaL in P. gingivalis has dual specificity for both O-antigen and APS repeating units.

FORENSIC SPORTS ANALYTICS: DETECTING AND PREDICTING MATCH-FIXING IN TENNIS

2014 ◽  
Vol 8 (1) ◽  
pp. 77-95 ◽  
Author(s):  
Ryan Rodenberg ◽  
Elihu D. Feustel
Keyword(s):  
Law Enforcement ◽  
Predictive Modeling ◽  
Market Price ◽  
Prima Facie ◽  
Fair Trading ◽  
Sports Analytics ◽  
Sports Industry ◽  
Betting Markets ◽  
Sports Leagues ◽  
Tennis Match

Despite some degree of lingering friction between sports leagues and wagering, the eradication of corruption in sports is one issue in which the interests of sports governing bodies, reputable gambling businesses, entities affiliated with the sports industry, and law enforcement are allied.  This paper aims to: (i) detect match-fixing corruption in tennis and (ii) predict such match-fixing before it occurs.  We first compare proxy measures of effort in fair matches versus possibly unfair matches – matches played in the first round, where wagering-induced manipulation is more likely.  The results show that players exert less effort (tanking) in the first round, even when adjusting for the slightly greater difference in skill level in such matches.  We next determine whether the gambling markets were able to identify fixing, tanking, or other types of manipulation before the match was played.  Using two predictive tennis models (ELO, Common-Opponent) to determine the fair trading price of a tennis match, we find prima facie evidence of the betting markets being affected, with an average of 23 matches per year likely being manipulated or outright fixed each year.  Finally, we determine whether fixed matches can be identified before they are played using predictive modeling and by observing real-time market price changes.  We conclude that when the betting price has a large irrational move away from the fair model-predicted price, the move is indicative of a fixed match before it is played. 

scholarly journals Dispersion of Micro Fibrillated Cellulose (MFC) in Poly(lactic acid) (PLA) from Lab-Scale to Semi-Industrial Processing Using Biobased Plasticizers as Dispersing Aids

Chemistry ◽  
2021 ◽  
Vol 3 (3) ◽  
pp. 896-915
Author(s):  
Giovanna Molinari ◽  
Vito Gigante ◽  
Stefano Fiori ◽  
Laura Aliotta ◽  
Andrea Lazzeri
Keyword(s):  
Lactic Acid ◽  
Mechanical Tests ◽  
Poly Lactic Acid ◽  
Scale Production ◽  
Dual Effect ◽  
Industrial Processing ◽  
Twin Screw ◽  
Poly Ethylene ◽  
Green Solution ◽  
Industrial Scale Production

In the present study, two commercial typologies of microfibrillated cellulose (MFC) (Exilva and Celish) with 2% wt % were firstly melt-compounded at the laboratory scale into polylactic acid (PLA) by a microcompounder. To reach an MFC proper dispersion and avoid the well-known aglomeration problems, the use of two kinds of biobased plasticisers (poly(ethylene glycol) (PEG) and lactic acid oligomer (OLA)) were investigated. The plasticizers had the dual effect of dispersing the MFC, and at the same time, they counterbalanced the excessive stiffness caused by the addition of MFC to the PLA matrix. Several preliminaries dilution tests, with different aqueous cellulose suspension/plasticizer weight ratios were carried out. These tests were accompanied by SEM observations and IR and mechanical tests on compression-molded films in order to select the best plasticizer content. The best formulation was then scaled up in a semi-industrial twin-screw extruder, feeding the solution by a peristaltic pump, to optimize the industrial-scale production of commercial MFC-based composites with a solvent-free method. From this study, it can be seen that the use of plasticisers as dispersing aids is a biobased and green solution that can be easily used in conventional extrusion techniques.

scholarly journals Returns from pulses in different regions of rajasthan at alternative price scenarios

2019 ◽  
Vol 39 (1) ◽  
Author(s):  
Latika Sharma ◽  
Bharat Bhushan
Keyword(s):  
Market Price ◽  
Uttar Pradesh ◽  
New Delhi ◽  
Net Return ◽  
Net Returns ◽  
Level Data ◽  
Large Scope ◽  
Major Producer ◽  
Cost Efficient ◽  
Economic Price

Rajasthan is a major producer of pulses along with Madhya Pradesh, Maharashtra, Karnataka and Uttar Pradesh. In 2013-14 Rajasthan had total area of 4197.7 thousand hectares under pulses with production of 2490.9 thousand tones and productivity of 593 kgs/ha. Moong and moth in kharif season and gram in rabi season are grown extensively in Rajasthan. The study is primarily based on plot-level data collected from the 600 representative households of 60 tehsils for block period (2008-09 to 2010-11) for Rajasthan under CCS scheme of Directorate of Economics and Statistics, Ministry of Agriculture, Govt. of India, New Delhi. The study examined the nutrient consumption level, nutrient and irrigation subsidies availed by the pulses per hectare. The net returns of various pulses to the farmer, to the society and to the environment were estimated across zones. Pea, arhar, lentil and black gram were the pulses providing highest net return to the farmer at market price. After deducting the cost of subsidies which is a cost to the society, moong and gram in arid and semiarid zones and pea and arhar in central and south eastern humid zones gave highest net return at economic price. The estimated results for technical, allocative and cost efficiency indicated that the farmers were not operating at optimal scale and there is large scope for increasing output through factors which are under control of farmers. Lentil and arhar were most technical and cost efficient pulse crops while moong is the least technical and cost efficient pulse. In most of the pulses, on an average, the overwhelming cause of inefficiency is technical rather than allocative.

Effect of Niacin on Mitochondria of Allium Cepa Root Cells

1967 ◽  
Vol 25 ◽  
pp. 78-79
Author(s):  
M. Arif Hayat
Keyword(s):  
Nicotinamide Adenine Dinucleotide ◽  
Hydrogen Transfer ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Nicotinamide Adenine ◽  
Root Tips ◽  
Root Cells ◽  
Transfer Processes ◽  
Standard Procedures ◽  
A Cell ◽  
Critical Interest

Although it is recognized that niacin (pyridine-3-carboxylic acid), incorporated as the amide in nicotinamide adenine dinucleotide (NAD) or in nicotinamide adenine dinucleotide phosphate (NADP), is a cofactor in hydrogen transfer in numerous enzyme reactions in all organisms studied, virtually no information is available on the effect of this vitamin on a cell at the submicroscopic level. Since mitochondria act as sites for many hydrogen transfer processes, the possible response of mitochondria to niacin treatment is, therefore, of critical interest.Onion bulbs were placed on vials filled with double distilled water in the dark at 25°C. After two days the bulbs and newly developed root system were transferred to vials containing 0.1% niacin. Root tips were collected at ¼, ½, 1, 2, 4, and 8 hr. intervals after treatment. The tissues were fixed in glutaraldehyde-OsO4 as well as in 2% KMnO4 according to standard procedures. In both cases, the tissues were dehydrated in an acetone series and embedded in Reynolds' lead citrate for 3-10 minutes.

Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

1978 ◽  
Vol 36 (2) ◽  
pp. 650-651
Author(s):  
Raul I. Garcia ◽  
Evelyn A. Flynn ◽  
George Szabo
Keyword(s):  
Direct Injection ◽  
Skin Pigmentation ◽  
Freeze Fracture ◽  
Pigment Granule ◽  
Time Lapse ◽  
Ultrastructural Level ◽  
Lanthanum Tracer ◽  
A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

Dendritic cells of the human epidermis: immuno-electron microscopic studies of la antigen reactivity

1978 ◽  
Vol 36 (2) ◽  
pp. 644-645
Author(s):  
G. Rowden ◽  
M. G. Lewis ◽  
T. M. Phillips
Keyword(s):  
Dendritic Cells ◽  
Langerhans Cells ◽  
Cell Types ◽  
Cell Type ◽  
Electron Microscopic ◽  
La Antigen ◽  
Microscopic Studies ◽  
A Cell ◽  
C3 Receptors ◽  
Antigen Reactivity

Langerhans cells of mammalian stratified squamous epithelial have proven to be an enigma since their discovery in 1868. These dendritic suprabasal cells have been considered as related to melanocytes either as effete cells, or as post divisional products. Although grafting experiments seemed to demonstrate the independence of the cell types, much confusion still exists. The presence in the epidermis of a cell type with morphological features seemingly shared by melanocytes and Langerhans cells has been especially troublesome. This so called "indeterminate", or " -dendritic cell" lacks both Langerhans cells granules and melanosomes, yet it is clearly not a keratinocyte. Suggestions have been made that it is related to either Langerhans cells or melanocyte. Recent studies have unequivocally demonstrated that Langerhans cells are independent cells with immune function. They display Fc and C3 receptors on their surface as well as la (immune region associated) antigens.

Presence of antibody in virus-infected brain cells of SSPE ferrets

1983 ◽  
Vol 41 ◽  
pp. 804-805
Author(s):  
Hannah R. Brown ◽  
Tammy L. Donato ◽  
Halldor Thormar
Keyword(s):  
Measles Virus ◽  
Plasma Cells ◽  
Subacute Sclerosing Panencephalitis ◽  
Protein A ◽  
Neutralizing Antibody ◽  
Brain Cells ◽  
Antibody Titers ◽  
A Cell ◽  
The Central Nervous System ◽  
The Brain

Measles virus specific immunoglobulin G (IgG) has been found in the brains of patients with subacute sclerosing panencephalitis (SSPE), a slowly progressing disease of the central nervous system (CNS) in children. IgG/albumin ratios indicate that the antibodies are synthesized within the CNS. Using the ferret as an animal model to study the disease, we have been attempting to localize the Ig's in the brains of animals inoculated with a cell associated strain of SSPE. In an earlier report, preliminary results using Protein A conjugated to horseradish peroxidase (PrAPx) (Dynatech Diagnostics Inc., South Windham, ME.) to detect antibodies revealed the presence of immunoglobulin mainly in antibody-producing plasma cells in inflammatory lesions and not in infected brain cells.In the present experiment we studied the brain of an SSPE ferret with neutralizing antibody titers of 1:1024 in serum and 1:512 in CSF at time of sacrifice 7 months after i.c. inoculation with SSPE measles virus-infected cells. The animal was perfused with saline and portions of the brain and spinal cord were immersed in periodate-lysine-paraformaldehyde (P-L-P) fixative. The ferret was not perfused with fixative because parts of the brain were used for virus isolation.

Aberrant Type C Virus Production in a Cell Line Derived from Balb/3T3

1972 ◽  
Vol 30 ◽  
pp. 286-287
Author(s):  
N. Savage ◽  
A. Hackett
Keyword(s):  
Electron Microscopy ◽  
Cell Line ◽  
Leukemia Virus ◽  
Morphological Characteristics ◽  
Virus Production ◽  
Oncogenic Viruses ◽  
Endogenous Virus ◽  
Virus Particles ◽  
Moloney Leukemia Virus ◽  
A Cell

A cell line, UC1-B, which was derived from Balb/3T3 cells, maintains the same morphological characteristics of the non-transformed parental culture, and shows no evidence of spontaneous virus production. Survey by electron microscopy shows that the cell line consists of spindle-shaped cells with no unusual features and no endogenous virus particles.UC1-B cells respond to Moloney leukemia virus (MLV) infection by a change in morphology and growth pattern which is typical of cells transformed by sarcoma virus. Electron microscopy shows that the cells are now variable in shape (rounded, rhomboid, and spindle), and each cell type has some microvilli. Virtually all (90%) of the cells show virus particles developing at the cell surface and within the cytoplasm. Maturing viruses, typical of the oncogenic viruses, are found along with atypical tubular forms in the same cell.

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