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if can use spray drying, do we need to concentrate enzyme after chromatography process before entering the spray dryer?
most of the research gives information on lab-scale operation conditions like temperature and volume. so may know any resources to scale up these parameters into larger-scale/ industrial scale?
thank you for your response.
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Yes it's possible to use spray drying to make alpha-amylase in powder form
- Use maltodextrin as carrier
- In order to retain the activity of enzyme after drying: optimize the feed rate and inlet/outlet temperature - higher value of feed ratio speed and Lower outlet air temperature should be kept in order to obtain high active dried alpha-amylase.
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Normally bacterial cellulose produced from fermentation was in the form of gelatinous layer. Almost 98% of their structure was water. If this gelatinous layer is dry in oven it will become a very thin layer film thus hard to be grind.
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I have tried two methods:
1. freeze dying BC followed by dipping in liquid nitrogen for a few seconds and grinding in a ball mill.
2. freeze dying followed BC by keeping in the refrigerator overnight and grinding in a blender. The powder is sieved and bigger particles are put back in the blender for another round. (modified method of Munair Badshah )
Both of the methods gave BC in powder or particle form.
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What are your thoughts about the equipment? Also, what was the average cost of the equipment?
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I agree with you that there are many self-appointed "experts" that pretend calculating the BMP of mixtures with tables, or doubling the production by adding "magic powders" of whatever nature. But BMP tests can be used indeed to improve the plant's performance. I developed a method and condensed 10 years of experience in true plant optimization in over 100 plants in Spain and Italy using AMPTS Light , anybody can buy and learn: https://www.routledge.com/Managing-Biogas-Plants-A-Practical-Guide/Rosato/p/book/9781138626614 . Anyway , changing one motor every 2 years or so is unacceptable for most customers, at least in Europe, that's why Bioprocess Control abandoned the DC motors and switched to brushless motors, which are virtually eternal. With brushless motors the stirring is exactly the same in all reactors, because the speed is controlled by an inverter and furthermore you can actually measure the unit power, which is impossible with DC motors. Anyway, as long as there is some stirring, and as long as the inoculum is not overstirred, there's no relevant difference in the BMP. Of course, the final word belongs to the purchase department. Unfortunately, thousands of researchers still do tests with eudiometers, syringes, manometers and similar self-built tools, others shake the bottles manually twice a day, just because their universities cannot afford anything better.
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My current research is in the carbon flux distribution of L. rhamnosus in a biofilm reactor and I am struggling to find research pertaining to the continuous metabolic analysis in literature. Any help would be highly appreciated.
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Dear Hendrik,
maybe these recent publications on 13C metabolic flux analysis in biofilm-forming P. aeruginosa and agar plate-grown E. coli might be of help:
Best,
Michael
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in general batch anaerobic digestion process which is the right optimum substrate/ inoculum ratio 0.5 or 1.0
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The Italian norm on BMP test (UNI/TS 11703:2018) states that the inoculum/substrate ratio must be comprised between 2 and 3 for general biomass digestion (i.e. 0.5 to 0.33 if you define as substrate/inoculum). Nevertheless, it also states that some substrates (glycerol, fats, industrial effluents containing inhibitors like olive mill wastewater, etc.) may require teh test to be performmed with I/S >= 5. This must be established by the lab operator and stated in the report of the test. For a discussion on the pros and cons of the Italian , German and IWA draft on BMP assay, please see chapter 6 of my book https://www.crcpress.com/Managing-Biogas-Plants-A-Practical-Guide/Rosato/p/book/9781138626614
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I know one kit but I am looking preferentially for some colorimetric analysis or quantification using fluorescence spectroscopy. Help regarding any of the methods would be appreciated.
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We have just established this method in our study group, see the paper attached.
Protocol:
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Hi guys,
I have a P. pastoris MutS phenotype which express very well in shaking flasks. I was trying to scale up the culture in a fermentor.
The culture was behaving normally for most of the fermentation process, but as soon as I started the methanol feeding (after a DO spike to make sure all the glycerol was consumed) the pH started to rise and the acid pump worked for the whole methanol feeding process.
I took samples at different time intervals and check them for protein expression. What I noticed is that the protein started to express as soon as 3 hours after the induction, then the expression increased in the next hours to a peak expression at 9 hours of induction. After that the cells were expressing less and less until the end of the fermentation. 
Comparing the expression level of the peak in the fermentor (9 hours) and the expression in flasks for 48h, in the fermentor I anyway got 5x less protein than flasks.
Do you have any idea how to improve the expression?
Why the pH of  the culture was increasing during methanol feeding?
Decrease in protein expression, could possibly due to no more nitrogen source in the medium?
The medium I used was the "Basal salt medium" as described by the invitrogen guidelines and the only nitrogen source was the ammonium hydroxide also used to keep the pH at 5.0
Thank you very much for your help,
Marco
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At any point of time during induction phase, the methanol concentration needs to be maintained at a sub optimal level & not optimal level. This is because, as most experts have mentioned, excess accumulation of methanol (above 3-5%) leads to toxicity for the cells, more than its utilization by the cells towards protein expression. The pH control herein becomes crucial as cells then begin to die (a possible reason of pH rise during induction). Solution could be, to feed optimal glycerol during growth phase along with a judicious supply of nitrogen source (NH4OH or any such other) till the point of reaching adaptation stage. This ensures reaching culture OD to at least 250-300. During adaptation, have a dual control by acid-alkali addition, by monitoring which way the pH is shifting towards. Have at least 6-8 hours of adaptation, before finally switching to 100% methanol. However, this time period needs to be established experimentally. During induction, keep temperature low (23-25 C). After adaptation, when one sees the normalization of set parameters (pH & DO) go for about 40-45 hrs induction, taking care to check the expression regularly along with the wet cell weight estimation. At one point of time, the expression is bound to dip. That is where take a call to harvest the culture.
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I'm a new researcher who research in Microbial Fuel Cell by adopting yeast (S. cerevisiae) as a biocatalyst. But one of the issue is the reaction mechanism: glucose + H2O --> CO2 + 24H+ + 24 e-  is obtained from anaerobic condition or aerobic condition? some references said from anaerobic condition, but also some references said from aerobic condition. It makes me a little bit confused.
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 Thank you, Mr. Michael. how about the number of electron and hydrogen produced? any big different between aerobic and anaerobic condition?
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Bioprocess engineers and Fermentation technologist. 
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Kla depends primarily on power input and medium. Reactor configuration and viscosity may lead to 'dead' zones reducing overall mass transfer. Also oxygen solubilty is important for mass transfer. So measurements are required or someone's else experience in a very similar reactor may give at least an indication.
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Pichia pastoris is fermented for recombination protein.
I set fermentation condition that DO value 50%.
I want reducing the pure oxygen rate, but maintain or increase DO value.
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In addition to all the answers above...
The DO in your fermentor will depend on the balance between oxygen input (gas-liquid transfer rate) and oxygen consumption by your yeast. If input is high and consumption is low, DO will increase and vise versa. Be aware that oxygen transfer rate is not the same as the volumetric oxygen input, most of the oxygen you will bubble into the fermenter will just espace to the headspace without dissolving into the liquid. Reducing the oxygen gas flow rate might even improve the dissolution of oxygen, if the reduced gas flow rate results in smaller bubbles.... So indeed, like proposed in the notes above, the trick is to improve the oxygen transfer rate QO2 which is a function of kla and the oxygen transfer driving force (=difference in oxygen concentration at liquid/gas surface and your intended 50% DO).
Options are (with increasing order of complexity):
- Reduce bubble size, mechanically by using a adapted gas injector (sintered metal or glass), increasing stirrrer speed, increase residence time of your bubbles in the fermentyer by using baffles, etc.
- Increasing oxygen pressure (this will not improve DO partial pressure in %, but will increase the dissolved oxygen concentration in mg/l)
- Change medium composition to improve specific oxygen transfer rate (kl) and/or bubble size a (e.g. by adding surfactants)
-Add oxygen in another way, e.g. by membrane units (either inserted in the fermentor or as an external unit)
Hope this helps. ;-)
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During Simultaneous Saccharification and Fermentation, Is it possible to know the amount of sugar released into the system by the enzyme from biomass and the amount of sugar consumed by the yeast? If possible, how?
I need to calculate the ethanol yield (in g per g glucose consumed) from an SSF experiment so that I can easily compare the result with that of SHF.
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There is an excellent article published 30 years ago where you can find the answer:
Good luck!
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Understanding algae responses to stress can help to improve our knowledge about algae adaptation strategies which can be used to engineer tolerant strains. Also, it can be used to manipulate culturing condition to yield the higher amount of secondary metabolites like carotenoid. 
In most of the articles that I am reading it is not mentioned why is important understanding adaptation mechanisms. Are there any other applications other than what I stated?
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Thank you for your time and guidance, it is very useful paper. 
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I read an article saying  that 66.07 g of NH4+ will be available in 1 litre of water if  1 mole of ammonium sulfate is introduced to water
But i presume that 36 g/lit of ammonium should be present but the above mentioned reference says that 66 g/lit. kindly clarify me 
1mole of ammonium sulfate = 132 g/lit ( N= 14,H=1,S=32,O=16) 
Ammonium sulfate when dissolved in aqueous solution produces two NH4+ ion i,e 2*18 = 36 gm.
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Hi, Ahamed!
You are correct. I think it is obvious that the "66.07" is a typographical error, as 6 and 3 are next to one another on a computer keypad.
Also, you mean "... if 1 mol of ammonium sulphate ...", not "1 M", just to be clear.
And if I remember correctly, IUPAC switched to spelling sulfur with an "f" in 1990. :D
regards,
Daniel
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I need to selectively isolate fungal cellulase/xylanase producers in a selective medium which supports both bacterial and fungal cellulase/xylanase producing organisms. Tetracycline, a broad spectrum antibiotic, has been shown to inhibit some fungal species. Sodium azide and cyclo-heximide can also prevent both bacterial and fungal growth. This condition is also applicable to some other common antibiotics. I need a more restrictive antibiotic with no known antifungal property so that I will not lose any chance of selecting the best fungal strain.
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streptomycin is good to stop fungus growth ,  I tried.. It should be added in traces.. 
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For 'fatty acid ethyl ester extraction from fish viscera' project, I'm using the enzymatic method to extract and hydrolyse the oil. In this case, I'm producing lipase from solid state fermentation using fungus. Which is later used for hydrolysis and transesterification as well. The solid state fermentation is carried out for 5 days. 
Is there any other method to replace the enzymatic method. I wants to speed up the extraction process? Is there any type catalyst or other method which can be used as alternative. 
I would be glad if anyone would help me in this or send me some reference. Thanks in advance. 
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i agree with all answers above.
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I want to check the interaction between magnetic immobilized enzyme with the nano materials. What are be the reaction parameters should be fixed for this?
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I think two possibilities can occur either the enzyme can increase or decrease in activity. This relate to stereo chemistry between enzyme and substrate. If the magnet can make the stereo binding site of enzyme-subtrate become more "fit", then the enzyme activity increase, and vise-versa.
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First off, eventually what I want to get is a close-packed, monolayer of polystyrene(PS) spheres at the air/water interface (Langmuir-Blodgett method). But unlike those mentioned in many references, my polystyrene spheres are not dispersed in ethanol or butanol, but rather they are dispersed in water. This complicates the method since the spheres in water cannot float upon the water sub-phase for the LB method.
Other than synthesis of polystyrene spheres from scratch, is there a way to replace the water media with ethanol?
Thanks.
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1) Dilute in ethanol, say four times;
2) Centrifuge;
3) Pipette off the excess solvent;
Repeat the cycle until the dispersion has a low enough water content.
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Is there a difference in efficiency when using sodium bicarbonate or sodium carbonate for CO2 gas when growing photosynthetic bacteria in a closed bioreactor?
An article of someone using this for any photosynthetic bacteria or algae would be perfect. 
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Hi Jared,
To my point, just the same stoichometric differences when use one or another cpturing salt. 
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I have searched dozens of papers with the mentioned methodologies for siderophores. I managed to determine which reagents were used, but lack information about concentrations, and volumes.
I am searching for alternatives to Atkins and Arnow methods (and Neiland's) for hydroxamate and cathecol-type siderophore screening.
Thanks in advance, 
Romeu
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Thank you Sosanka!
Obrigado Isadora!
Sorry for the late answer, I was out of the lab for a few days (sampling) :). Have a nice day/week-end
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Hi,
I am working on Carbon-dioxide sequestration by microalgae using coal-fired actual flue gas in photobioreactors. The flue gas is being used as it is (without any pre-treatment).  I am studying the flue gas introduced heavy metals into the system (medium and/or biomass). Could anyone please throw some light, which heavy metals I should be focusing on when analyzing the samples in ICP-MS. I am looking for Ni, Cr, Cd, As,Pb but Mo,Sn, Rb are also seen. Is it a fine observation? 
Thanks
Geetanjali
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Dear Prof. Karbassi,
Thank you so much for your comments. I have measured Hg and other metals especially Cd,As,Se. Pb etc. and their concentrations were not really high in the biomass when I compared it with maximum permissible limits in drinking water and or plants as set by WHO guidelines. This looks good since my further aim is to assess the potential of biomass as feed. 
Thanks for your feedback.
Geetanjali
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May I know is nanocrystalline cellulose will sink to the bottom? I have studied a paper which stated that unmodiefied NCC will form two layers (NCC and distilled water) and modified NCC will remained in suspension and form a milky suspension
I get two layers after centrifugation.
Btw, how am I going to dry the NCC? I do not have any freeze-drying equipment here. Only oven.
Thanks 
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cellulose is insoluble both in common organic solvents and in water. You can use the azeotropic system "Dean Stark" in reaction to eliminate the amount of water and the reaction is made with a fine condition
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I am working on visual detection of alcohol by using immobilization of AXO (alcohol oxidase) in agarose matrix. The method which I am following is involve reaction of alcohol (source) with AOX in polymer matrix. By which generation of H2O2 occur and react with HRP to produce colour change to green to DARK green. But I am not getting the satisfactory results. Please suggest any other polymer matrix which I can use or any alteration in experiment. I am attaching the reference paper.
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Dear Deepak,
It has been shown that fenugreek hydrogel-agarose matrix with gold nanoparticles is a better film than any of those you mentioned (agarose and etc.). Immobilization of acetylcholinesterase on the membrane resulted in high enzyme retention efficiency (92%) and a significantly prolonged shelf life of the enzyme (half-life, 55 days).
I suspect that your experiments may be were unsuccessful because of the instability of the immobilized enzymes on the agarose matrix.
I suggest that you use enugreek hydrogel-agarose film which consists of  2% fenugreek hydrogel and 2% agarose.
For more on fenugreek hydrogel-agarose matrix with gold nanoparticles, please use the following links:
Hoping this will be helpful,
Rafik
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Hello! We did a small biodigester at home, it had two units: a recipient with 0.25L and a another variable volume recipient for gas storage. The biodigester starts up with 1.5kg of food waste, 1 kg of cow manure and 5L of water. The system seems to be good during adaptation time (we planned 15d for this and then to operate it in semicontinuous mode), pH measurements every three days was nearly to 7. Neverthless, 9 days after the start up we have to burn  the biogas the because the storage recipient was too small, but... it didn't burnt.
We have an hypothesis about what was bad in our design: the (inoculum/substrate ratio) was to small (0.6 aprox.) and for food waste the recommended ISR must be over 2 for preventing the inhibition (food waste are easily biodegradable matter)
We are not sure about adaptation time or if we can burnt the biogas daily, but in this case the pressure maybe won´t be for flow through the tube. Maybe we must change the design or we are doing something bad during the start up?
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Good morning, Lenis, I have just seen your question. Your assertion was correct, in my opinion. However, for start-up and acclimatisation at the same time, even an inoculum/substrate ratio of 2 would be far too high. For example, just to carry out biomethane potential tests using already acclimatised biomass we use a ratio >6 to minimise potential stress. Now, we operate full-scale food waste AD plants in the UK. We have done it for over 10 years now and the volumetric loading to our 16000m3 plant is around 300m3/day => 50 days HRT.  I see that your are probably starting in semi-batch mode, but at that initial loading I am surprised the pH has not dropped further. If it was me, I would start at an inoculum/substrate >25 and monitor biogas composition more regularly, if possible. It CH4>50% increase OLR gradually. I normally control start-up on the basis of biogas CH4 content and flow. Since you are looking at start-up and acclimatisation (i.e., you are not using an inoculum from a food waste digester), I would expect start-up to take a long time, as Rowayda indicated (or longer!). Can you not get some UASB solids to start-up. I hope this is useful.  
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Hello Everyone, 
Am trying to develop an enzyme hydrolysis protocol using bromelain. The specific activity of the bromelain enzyme is 4.5 U/mg protein (provided by the supplier).
My task is to hydrolyze a fish sample using an enzyme sample ratio of 1% (1 g of enzyme powder added to 100 g of fish).
However, I observed from literature that enzyme sample ratio is frequently expressed as U/100 g of sample. 
Hence, how do I convert the 1% enzyme-sample ratio to U/100 g of fish? 
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I agree, you must determine the activity (U) per mass of powdered enzyme.
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I wanted to study the effect of 1 % nitrogen source on the enzyme production via Solid State Fermentation.
Which method is correct?
A). The 1% nitrogen source is determine based on the volume of moisture I provide
or
B). The 1% nitrogen source is determine based on weight of the solid substrate I wanted to use.
Does anyone have experience on this matter? TQ
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I agree with all colegues, I like Rual consideration. I want to add just take into consideration your 5 g as dry basis
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I want to extract cellulose degrading enzymes from the insects salivary glands, guts tissues and gut symbiotic microbes (Like Bacteria and Protozoa). so, can any one will help me to find best protocols for that.
Best regards   
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You can extract cellulase enzyme  fro termite gut very easily . Dissect gut content anf get them in a tube with alcohol. This may be subsequently assessed
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I m working on kinetic study of biodiesel production. my questions are as follows
1. I want to analyse the sample with HPLC so which concentration should i estimate, is it Triglyceride or FAME.
2. what are the standards generally used for the estimation for both TG and FAME.
3. After getting the concentrations how to estimate the rate constant.
4. can we estimate the gibbs free energy from calculated rate constant.
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Dear sumit actually first order kinetic depend on the concentration of reactants or reagents if you need rate you have to do apply spectrophotometric method for the determination of rate constant. 
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I was doing some research on ABE fermentation using bacterium Clostridium acetobutylicum.Before fermentation, deactivated bacteria cultures are stored at -80 °C. The thawed cells are then inoculated into 12 mL synthetic medium containing glucose (30 g/L) and Yeast Extract (5 g/L) in 15 mL Hungate tubes (pre-cultures). Cells are then grown under an aerobic conditions for 48 h at 37 °C, thereafter they are transferred into fermentation bottles.
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Which clostridium species gives the highest concentration of butanol from the broth? 
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Dear all,
While determining the pH stability of a an enzyme. I incubated the enzyme at pH range 3-10 for 72hrs. 
I wish to ask, if it is sufficient to describe an enzyme which retains its activity at alkaline pH 10 as "alkaline stable" even though:
1- I did not test stability up to pH 14
and
2- that its optimum activity was pH 5.3 during assay.
Your kind advice and criticism are welcomed.
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Dear Ajijolakewu Kamoldeen : If your enzyme retains the activity at pH 10 at least for about 12 hr. then you can name as alkaline enzyme.  However, you will have to work out on enzyme half lifeperiod.
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Hi,
I'M working on the soil Enzymes. I need to to check the total reductase enzyme production and its activity produced by certain enzymes especially when the inoculum is converted to liquid media. Can anyone suggest to me a particular method or protocol to analyze the reductase enzyme production by the microbial community in the media?
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I want to study nitrate reductase. I need a specific protocol or some 
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Are there website or scientific paper that have been discussed?
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I am giving below the following reference ; 
PNAS | April 8, 2014 | vol. 111 | no. 14 | 5159–5164
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I need commercial grade Cellobiose dehydrogenase (CDH). Does anyone have the information about any company/organization/supplier who can supply commercial grade CDH?
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At the moment there is no commercial provider of CDH. There are only about 20 research groups that produce it and use it.
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Now I need to design a transfer function for my bioreactor. The controllable parameters pH, DO, Speed, Temperature available in the system. I need to control all the parameters.
Is it possible to add more parameters within a single transfer function?
If yes, Then how i control the parameters separately?
More over i have a plan to design a MRAC for that system.
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Yes. The Stirrer Speed in RPM
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I'm designing a bioprocess using Nicotiana Tabacum BY-2 plant cells, I need it's kinetic parameters, thank you.
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You can find the contents of "Biochemical Engineering & Biotechnology" from added file here.
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I am currently running an MFC at OCV and I want to start applying loads on it. What really determines the starting resistance to be applied to the cell? I read a lot of literature that start with 1kohms and some as low as 10 ohms and I have done this in this past but is there another specific criteria?
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Generally, the external resistance should be comparable to the internal resistance. However, during the starting stage, the internal resistance is supposed to gradually decrease due to the development of functional microbial film on electrodes. So there is no rule, and you may yield different MFCs when apply different resistors. 
Here are some tips: The internal resistance depends on the surface area of the electrodes and the distances between the electrodes. If the area is huge and the distance is close, choose a resistor less than 1 k ohm, If not, choose more than 1k ohm. If you have no idea, just use any resistor and monitor the CCV for several days. For a conventional MFC, the OCV varies from 0.5 to 0.8. The CCV will decrease dramatically when loaded and slowly recover. If not recover, use bigger resistor; if no significant decrease, use smaller resistor.
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In bioreactors, I maintain a low pH via combination of cell metabolism and base addition. In shake flask, base addition is not possible, and I've been looking for a suitable, biologically compatible buffer system for a pH between 3 and 4. Most biological buffers are designed for much higher pH ranges.
I've tried sodium citrate/citric acid, but my cells grow poorly and lyse.
Any suggestions? I plan to try a glycine-HCl system, but I'm not too confident about it, the HCl doesn't sound cell-culture friendly.
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As an alternative to adding a buffer, you might try using urea rather than ammonium salts as the nitrogen source. This simple replacement strongly decreases acidification of yeast cultures in defined media and, in most yeast strains, has little impact on growth rates. 
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Hi, everybody
I am working on phytase production by a B. subtilis strain. We are going to scale up the production using a NBS BioFlo 310 Bioreactor. We are going to use NBS EX-2000 Gas Analyzer for measuring O2/CO2 from the bioreactor exhaust, but unfortunately, we are having difficulty working with this equipment. Although, the air enters and exits from the machine, but its flow-meter doesn't work and the results aren't satisfactory! Also, how can I make sure that the machine is working properly?
Thank you in advance!
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Hello Karim.
I would contact the manufacturer (Eppendorf) and ask for advice.
Perhaps your exhaust gas flowrate is out of range (0.5 to 1.0 L/min) or the pressure is too high (0.5psig max).
I would also calibrate it before each use, you can buy specific gas mixtures from compressed gas suppliers.
Good luck.
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My enzyme is cellulase from A. niger
please guide me how can be used and how much needed for fermentation?
is it based on volume of fermentation medium or substrate (biomass)?
if the enzyme inoculation related to FPU how can convert it to gram/liter?
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Hi Shahabaldin
In addition to the above guidelines, I should say that you can consult with your provider and get the correct instructions for how to assay and the range of activity of your products.
However, usually you should prepare a solution in the suitable buffer with exact activity so you will be able to do your calculation whatever you would like to report.
Good luck
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ammonium is nitrified mostly to nitrate and most phosphates are removed
during the aerobic period (aeration), where the accumulated nitrate
is completely denitrified during the anoxic period (non-aeration), and phosphorus (P) is taken up.
P is found in wastewater as phosphates (orthophosphates, condensed
phosphates, organic phosphate fractions), and it can be eliminated either by
precipitation and/or adsorption, or by luxury uptake. Luxury uptake of P is accomplished by the introduction of an anaerobic phase in the wastewater treatment line ahead of the aerobic phase and recycling of sludge through the anaerobic and aerobic phase. Exposing mixed liquor to an anaerobic/aerobic sequence selects phosphate accumulating microorganisms (PAOs) due to a competition between PAOs and other aerobic organisms. This competition mechanism is based on a complete anaerobic uptake of the lower fatty acids by the polyP organisms (i.e., PAOs), which assures that in the aerobic phase, no fatty acids are left. The polyP organisms use the stored internal substrate during aerobic conditions while other aerobic organisms are lacking substrate. This process is usually referred to as the enhanced biological phosphorus removal (EBPR) process. EBPR process can be established in MBR treatment unit by operating it in intermittent aeration mode. 
Intermittently aerated MBR can achieve nitrogen and phosphorus removal
by a simultaneous nitrification and denitrification, P-uptake and P-release in
the same reactor in accordance with time cycle of aeration and non-aeration.
However, even though intermittent aeration was successful in removing nitrogen, P removal was difficult to achieve at a higher level  This is probably due to the inhibition by nitrate. In the anaerobic stage, nitrate reduces phosphate release, and in the aerobic stage it diminishes its uptake.
Denitrification has more capability than phosphorus release with respect to
the competition of substrate. This is because nitrate will be utilized as a final electron acceptor in the growth of non-polyP heterotrophs. Thereby, the amount of substrate available for polyP organisms is reduced and hence the removal of phosphorus is lowered. There are some studies that confirm the ability of polyP organisms for denitrification, however, not all PAOs can use nitrate as an electron acceptor.
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Please find attached this chapter... Hope it would be able to solve most of your queries.. Good luck... Best regards-Deepti
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Looking to make a bio-fuel cell with GOx as the anode enzyme and BOx as the cathode enzyme. 
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What are your current collectors (i.e. gold, carbon paper, etc.)? Are you operating in DET or MET mode? SAMs are frequently used for modifying on gold, but adsorption and entrapment are more comment for carbon. Entrapment techniques are very common if you are operating in MET mode with a redox polymer as mediator.
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I'm looking for some help in setting up a Bioreactor system for cultures of Cupriavidus necator which will be gas-fed growths. My problem comes in determining the flow rates that will be required for the gases. Without some idea of the necessary flow rates suitable to feed a culture, I cannot determine which Mass Flow Controller's I will need to purchase.
So far my search for information has only revealed various ratios of gases used previously, and that the rates must be varied in order to support the growths. What I need to know is the ratio of gas flow to culture size, so I can calculate the range of flow control I will need for my system in relation to the vessel size, for Air, Oxygen, Nitrogen and Carbon dioxide.
Does anyone know this information? Or can direct me to some papers/journals/books etc that can give me some idea? Or even have experience in Cupriavidus necator cultures?
Any assistance will be very welcome. Thank you.
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Thanks everyone.
Shane, to answer your question the CO2 is to be used as the carbon source as the projects are designed around using C.necator feeding off of syngas for biofuel production. So it will be aerobic growths, but as the projects have not been fully designed yet I couldn't say much more. It is possible that there will also be anaerobic growths as well, just one of the joys of a research centre, need to be prepared for everything! 
Assuming that I will have to cascade for the DO control and include Oxygen enrichment, would you have a suggestion about what vvm of O2 I might need as a maximum? It's literally just a matter of deciding on suitable MFC's right now to cover the most likely range of use.
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We are interested to use the Aber Futura Biomass probe (Dielectric spectroscopy) to measure the capacitance data of E.Coli and also scanning data of E.coli cell parameters (Cole-Cole alpha,Cell Diameter,Biovolume, Conductivity etc...). We are using bacterial mode(Default: 1.1MHz) with Polarisation OFF but we couldn't able to measure the capacitance data using default mode. Our offline data suggested that cells are growing well. Anyone can help us to solve this issue.
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Hi,
Aber Futura, work with two types of on-line biomass measurements, optical and capacitance. The second one, takes a measurement of available cells in the medium. You should ask them, the range of capacitance depending of your type of cells and the type of medium (sometimes there a lot of nutrients that can affect the capacitance measurements).
In the attached file, you can have an idea of this.
Best Regards.
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I just began working with E. coli K-12 for use as an oxygen scavenger in my reactors. I'm growing them on glucose, but since glucose is fermented I expect it to be quickly depleted in my reactor since I cannot reinject for reasons I won't go into. Once their food source is depleted, they'll no longer be able to reduce the O2 that periodically enters. I'd like to give E. coli a non-fermentable substrate that requires O2 as an electron acceptor. This will maintain a low level of continuous growth as O2 slowly leaks into my reactor since the cells can only grow when O2 is present. Can someone give me a short list of electron donors that E. coli can grow on with O2, but not without?
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lactate, succinate, ethanol, acetate should work as they are all products of fermentation in E. coli.
Yoram
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i am currently fermenting pichia and I am having issues with contamination. A red film keeps appearing in my solution after i inoculate it. instead of going acidic overtime the solution becomes basic. I have made new seed stocks which were grown in zeocion. Using the new seed stocks still leads to contamination.
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 I would suggest the following strategies: 1. You may want to streak plate your culture and inoculate a single colony for seed culture to rule out contamination in the seed culture itself. 2. Increase the inoculum volume 3. Increase the amount of methanol in the culture media. This apart, you may like to re-ensure that all the steps are being carried out asceptically as well as check for any leak in the fermentation vessel as well as its proper sterilization. Hope this helps....All the best!
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I like to study the barotolerance of few bacterial strains by applying high pressure. In the Bioreactor pneumatic pressure is applied. Can anyone suggest me high pressure resistive materials to inoculate cultures and incubate in the reactor? Similar studies has used sterile plastic bags, hungate tubes, blood pouches were they have applied hydraulic pressure. The material should withstand high pneumatic pressure.
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I have used saline pouches similar to blood bag for inoculation and the pouches were tested till 50 bar pressure. There is no leakage and cross contamination..hope it could be tested at more pressure condition. Thanks everyone.
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We are trying to find the best stop solution for in vitro microbiota fermentation that does not interfere with SCFA readings. So far I've seen KOH, CuSO4, Na3PO4 used as stop solution. If you have used any of these or any other stop solutions, what is your opinion? Thank you very much.
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Filter sterilize 0.45 uM, you don't add any contaminant.
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I have tried to increase molecular weight of poly lactic acid of lower molecular weight by chain extension with di dissociates . It give me weight average molecular weight of 200000 Da. I would like to know does there is any other method to get ultra high molecular weight PLA.
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Helena is completely right with her answer. In order to obtain high Mw PLA, you should use ROP from lactides.
cheers,
Miguel
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I need to know the conversion of density to mg/l and how much ml needed to prepare 100mg/l of solution of tert. butanol in water. The density of tert.butanol is 0.775g/ml, can anyone suggest this calculation.
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Hii..Sakti Sharmila..
As the density is 0.775g/ml = 775mg/ml
Required conc. 100mg/L = 100ng/ml
Then N1V1=N2V2
Say V2= 100ml then V1= (100ng*100ml)/775mg= 12.90ul
Add 12.90 ul in water to make final volume 100ml
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I'm currently conducting research on the production of xylanase, pectinase and protease via solid state fermentation. The crude enzyme I harvested from each set of SSF is my primary source of each enzyme I mentioned earlier.
From my literature review it is okay to use sodium acetate buffer for the extraction of xylanase and pectinase. How about protease? does sodium acetate buffer suitable for the leaching out of protease from the SSF?
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Since most of the pectinase work in acidic pH therefore the extraction buffer used is generally citrate or acetate. Protease can be neutral, acidic or alkaline. Based on type of protease secreted by your fungi during SSF, you need to choose the buffer system. When I was doing my Ph.D. even physiological saline or 1% Tween-80/20 used to be sufficient for leaching out protease from the SSF... 
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Nicotiana Tabacum BY-2
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I have no information on this. Sorry.
Ben
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Currently I'm doing research on production of hydrolytic enzymes via SFF. Does anyone know the best way to store crude enzyme while maintaining its freshness in term of enzyme activity. And how long can it maintain?
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First point: it is impossible to generalise and you need to play around with conditions to find out what your enzyme likes and does not like!
Second point, although it is a bit boring, that initial exploration will give rich rewards and could save you much grief. People often persist with conditions in which their enzyme is terribly unstable when a few experiments could eliminate the problem.
Third, you haven't told us what kind of hydrolytic enzyme. If it is a proteinase then this is the toughest challenge, because, given enough time a proteinase will digest itself. Even if your enzyme is not a proteinase, a crude extract is bound to contain protepytic activity and so a wise precaution is to use a cocktail of proteinase inhibitors. There are cocktails commercially available to take account of the fact that a proteinase might be a serine proteinase, a cysteine proteinase, a metalloproteinase, an aspartate proteinase.
Fourth you should try some simple experiments to explore pH, temperature and choice of buffer substance in terms of how they affect stability over time. Sample and assay e.g. every hour and you will soon find out:
if it is a good idea to keep on ice (some enzymes are actually cold-labile!);
if the enzyme is perhaps stable in Tris but not in phosphate, or vice versa;
if the enzyme is really happy at pH6 but terribly unhappy at pH8;
Finally, if you find that after these preliminary basic experiments you still have a problem, there a number of well-tried additives to explore - glycerol, bovine serum albumin, sucrose, trehalose, PEG, salt etc.
Above all, remember that each protein is unique and what works for one protein may be quite different from what works for the next.
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I am unable to have a definite peak in the chromatogram of the five monomeric sugar and cellobiose analysis using HPLC. All five monomeric sugars, except Glucose, forms tailing peaks while Cellobiose is not detected. This tailing peaks caused overlapping when all sugar standards were run. My sugars and cellobiose were either from Merck or Sigma. 
 HPLC conditions are well stated. Other details are:
The brand and model of HPLC used: -HPLC Type: Fisher Scientific Thermofisher
-Brand: Shidmazu
-Model: RID-10A
Size of column: -Product name: APS-2 Hypersil
Diameter(mm): 250 x 4.6
-Particle size: (micron-N-)
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@ Dr Henrik Romar, in case of further works and a possible column blocking, which alternative mobile phase would you recommend ?
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We are trying to make anaerobic fermentation media from scratch. However, we noticed that hemin precipitated in the bottom of the media. Is this normal? If not, is there any way to dissolve hemin? Thanks!
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Hi Tung
Do you use the Hemin with the two branched carbonic acids?
I have no experience in Anaerobic culturing, buth this  structure looks a lot like you have many points for complex or ionic interaction.
The solubility and availability of Iron can be a generalized problem, maybe you have an interaction to phosphate molecules or others in your media, that lowers the solubility.
If your pH is far below 7, it is not surprising that two not dissociated carbonic acids also lower the solubility.....may be they form an internal salt or complex.
Did you try to dissolve the molecule with a higher pH and use enough time? Some trace elements need time to dissolve clearly.
For fermentation you have the chance to feed Hemin i a dissolved form, so you can make sure that you always have it present in solution.  For Iron (II) and (III) the problems are similar, sometimes you must feed it because you do not get stable conditions for the complete reaction time   
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I am currently conducting a research into fungal enzymes involved in glycosyl hydrolysis. I am to select one of three organisms for enzymes production. The activity of the enzyme produced by the best of these three organisms is twice the second best and three times the least. However, when I used the same enzyme units (30FPU) for the three organism in a target hydrolysis, enzymes from the other two organisms (those with lower activities) had  higher sugar yields (10%  total sugar yield more than the best organism). Of course, I thought that the other two organisms might posses some other complementary enzymes which act in synergy to produce a higher sugar yield but I am concerned that they however require a higher volume of crude enzymes or some expensive enzyme concentration mechanisms that will rather make the hydrolytic process non cost effective. I urgently need a decisive guide about this. 
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Enzyme activity by set protocol and applying these enzymes for particular target reaction are totally different things. Factors influencing the target reaction might have significant effect on the enzyme from your three organisms. So I suggest choose your organism based on the actual performance in your target reaction and then further investigate the parameters influencing the enzyme activity such as temperature, pH, solvents, buffer etc. 
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Basically I work on solid state fermentation of fungus and find it really difficult to confirm if my seed cultures are pure.
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You can do centrifugation of fermenter sample around 1000 g for 5 to 10 min , which will first remove the fungal growth and followed by supernatant smear  gram staining to check bacterial contamination.
Also the fermenter sample may strike plated on SCDM agar ( Soyabean Casein Digest Medium)  and need to incubate at 35-37 deg. for 24 hr. If any bacterial  contamination is present ,it will grow faster and  will be helpful prove present or absent of bacterial contamination .
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the pattern of consumption of amino acid is depend on yeast strain. 
but is there any one knows the availability of oxygen also cause to consumed more amino acid or not?
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Yes, you are correct. The alcoholic fermentation is conducted by yeast of the genus Saccharomyces. The two common species involved are S. cerevisiae and S. bayanus. These two species are closely related, and the subject of a continuing debate among taxonomists as to whether they constitute separate species or races of the same species. Saccharomyces converts the glucose, fructose and sucrose found in grape must and juice into ethanol via the process of fermentation. In fermentation, an organic
compound, in this case acetaldehyde, serves as terminal electron acceptor. This leads
to the production of ethanol. In aerobic respiration an exhaustive consumption of amino acids occur, but in anaerobic respiration fewer amino acids are consumed. 
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I may not know enough to know if this is even possible, but if it were: who, what industry, where would be likely to be working on something like this?  I am sure there would be more interest in this kind of thing in countries who acknowledge the importance of LAB fermented foods for physical and mental wellness.
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Yes, I was referring to that.  I will check them out!  Thanks!
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I want to use temporary immersion for differentiation phase in somatic embryogenesis.
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Also available at: Application of bioreactor systems for large scale productio...
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I need to observe the viscosity in a bioreactor on-line. Does anybody know how to do so? Is there any device/ any probe to measure viscosity?
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The bioreactor agitator may possibly be adapted to provide real-time torque measurements, which could be correlated with off-line viscosity data obtained trough laboratory essays.  A small level of sinusoidal excitation of the agitation speed can help to obtain on-line viscosity estimates through the recursive least squares algorithm with forgetting factor. For a study based at a related estimation strategy, applied for the energetic optimization and adaptive control of aerobic fermenters, you can check: C.M.G.A. Queirós, "Controlo do Oxigénio Dissolvido em Fermentadores para Minimização de Energia Consumida", MSc Thesis, Instituto Superior Técnico, Universidade Técnica de Lisboa, 1997 (in Portuguese); https://www.researchgate.net/publication/233799574_Controlo_do_Oxignio_Dissolvido_em_Fermentadores_para_Minimizao_de_Energia_Consumida?ev=prf_pub
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I deal with production of Lipase from Candida sps. (CALB). The fermentation has been recently scaled up to a 3L fermentor. The problem that I face now is the fluctuating O.D values of the broth measured through a period of 48 hours. I cannot seem to point out the actual cause of the decline in O.D. 
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It's easy: Bible OLD TESTAMENT Genesis 3:19 You will eat bread by the sweat of your brow until you return to the ground, since you were taken from it. For you are dust, and you will return to dust." Candida makes the same. OD of dust is negligible.
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I need to extract this compounds from my biodiesel sample in order to characterize them in HPLC tests. Any suggestions are welcome.
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Thank you Lorena
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Obtaining microalgal oils for biodiesel research.
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The  BETA Lab of Texas A&M University  is doing researches of producing bio-oil from microalgae. However, Im not sure if they sale bio-oil. Collaborative research might  be more interesting to them.
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I am working on separation of biosurfactant from fermentation broth by using the three phase partitioning method. In this method biosurfactant gets partitioned in t-butanol phase.
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If you want to check the biosurfactant production in the fermentation media then you can check the surface tension and emulsification activity.  and for analysis consult the paper attached..I am sure you will find many solutions.. :):)
Good luck,
Garima
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I want to bind plant cells together with biopolymer (biocompatible) with a very simple protocol in order to not induce too much stress to the cells.
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That's an interesting problem. It depends on what your starting material is and how tightly bound you want them to be.  Are they suspension-cultured cells?  You might consider something like alginate, (available from FMC biopolymers or Sigma) which can be gelled with Ca++.  You could centrifuge them through an alginate layer, maybe 1-2%, and then gel them in place when you get the degree of proximity you need.  There are also biocompatible alginate/methacrylate polymers, some of which can be catalysed with low dosage UV light. 
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I am working on enzyme production and want to study its kinetics. To stop an enzymatic reaction I need to add KCN. Due to potential toxicity, I don't want to use KCN, so please suggest some other enzyme inhibitors that have little or no toxicity..
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You can use sodium azide as its a known inhibitor for cytochrome oxidase instead of KCN. Hope it serves your purpose.... Best regards
Deepti
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My shake flask experiment when lactobacillus delbreuckii lactis ATCC4797 grown in media containing lactose and casein hydrolysate, in which trace quantities of calcium carbonate added as neutralizing agent. We observed increase in pH from 6.72 to 8.73 after 46 hours. Cell OD is also found to increasing ironically. I would like to put before this forum to what would be the possible cause of increasing pH whereas the with the same organism and media components without calcium carbonate were showing decreased pH as is expected.
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It may be possible.....without pH control this strain will produce acid and you have found the same...but when added CaCO3, it will react with acid produced by the strain and will produce CO2. In high concentration of CO2, the residual CaCO3 may have the chance to form Calcium bicarbonate that have alkaline pH (around 7-10). This effects seems more pronounced when culture is grown under static condition. Your cell growth will not be hampered since it is not inhibiting normal biochemistry of cell...You may check my hypothesis by adding different concentration of CaCO3, check pH under shaking and static condition, I am sure you will find variations in final pH. Remember to check initial media pH after autoclaving.
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I would like to produce gel filament of cell entrapped in polymer solution. But when I mix the suspension with the polymer solution, a pre-gel is formed and air bubbles are also entrapped in the mixture. I've already tried to removed the air bubbles by vacuuming but it takes very long time and isn't good for cell viability. Any idea how to remove the air bubbles without causing cell damaging?
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Hi, maybe you can use slower stirring speed while mixing the solution and add it gradually, i had same problem with prepare hydrocolloid solution at high concentration and i added the particles gradually to the solution at lower stirring speed.
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Conventional enzyme screening methods are often time consuming and laborious. There must be some alternative way to do this. Can anybody suggest a reliable method for this?
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Thank you Boopathi and Rinkoo
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When we plot glucose consumption, ethanol production or growth curve in a fermentation process, most of the time we can not get a smooth line connecting the data points. I found some articles e.g.:
1. Alfenore et al. (2002) Appl. Microbiol. Biotechnol. 60:67–72
2. da Cruz et al. (2003) J. Inst. Brew. 109(4): 349–355
These authors seems to use trend line on their graph, since I noticed that the line are not exactly connecting the data points.
Is there anyone know that we may do that? if the answer is yes , can anyone suggest what regression used for those curve and refer me to any reference that mentioned this?
Thank you in advance.
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Not required.
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I did the experiment for the determination of reducing power by oyaizu 1986 method and found absorbance values but how to do the calculation to determine reducing power or the extract? do we have any formula? Or the absorbance values corresponds to reducing power? What unit should I use?
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Please refer eplantscience.com for clear-cut calculations
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I have a digital scale that reads in the format 0.00 and in grams. I need to measure 4.5 milligrams. Would I still be able to use this scale? Shouldn't it read 0.45 g for 4.5 mg's? I'd settle for 4 milligrams instead of 4.5.
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When you want to have milligrams you must to multiply grams by 1000, so 4.5 mg equals 0.0045 and you will not see it in your scale.
Good luck
Mustafa
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Does anybody have any ideas about water binding capacity and water holding capacity? Are there any differences between water binding capacity and water holding capacity? And how to measure them?
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Water binding capacity is the tendency of water to associate with hydrophilic substances whereas Water holding capacity is the ability of a matrix of molecules to entrap large amounts of water in a manner such that exudation is prevented. Bound water exists in the vicinity of solutes and other non-aqueous constituents, exhibit reduced mobility and properties differing significantly from “bulk water” in the same system and does not freeze at -40°C.
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I am wondering if a protein disulfide bond shuffling can be started without denaturant in presence.
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Yes. I found this for a fragment of the LDL receptor
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If we want to relate changes in NADH/NAD ratio to increase/decrease in production of certain metabolite in a bacteria, then what range of ratio changes will be considered significant?
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For a specific condition, the redox level might not be same as with other condition. It applies same for the use of different substrate. However, the ratio is directly related with redox and it will effect the product formation. Therefore, doing a back calculation, I think you may get some idea of electron flow. that will give you an answer.
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I am performing the cellulase + cellobiase activity checking.
For cellulase, I managed to check its activity in FPU/ml as described by Ghose, 1987.
Can anyone tell me the rapid, simple method to check the cellobiase activity?
Any paper to refer?
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Thank you Safri
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Bioprocess industries are facing number of challenges like expensive process, new technology and sophistication of instrumentation and control etc.
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Process Analytical Technology (PAT) is a system for designing, analyzing and controlling manufacturing processes based on an understanding of the engineering and scientific principles involved and identification of variables that affect product quality. It is based on the US FDA's belief that: “quality cannot be tested into products; it should be built-in or should be by design.” This article provides the reader with an overview of the evolution of this technology, its key benefits and the challenges faced in its implementation. It then explains how PAT can be applied to a typical biopharmaceutical manufacturing process involving upstream and downstream processing, drug product manufacturing and chemometrics.
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How does the polarity of media change during CHO cell culture and what products/by products might increase/decrease the polarity?
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Thank you very much Dr. Thomson. How can I roughly estimate the polarity changes. I am trying to understand changes occurred to the tryptophan fluorescence along the CHO batch culture.
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I am working on bioethanol production. How can I quantify the reducing sugars percentage (glucose, xylose, arabinose,galactose, etc.) by using HPLC, after pretreatment, and what are the column conditions for the analysis?
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I had something like this problem during my research, for this purpose you can use HPLC with refractive index detection, with folowing parametes:
Mobile phase - 25 min, from 10 to 20 mg/l Ca-EDTA,
flow rate - 1ml/min,
Column - Ultron PS80-H, with Ultron PS 80G predcolumn or equvalent.
By my opinion it will be work.
Do not hesitate to ask questions. Good luck
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Can I use Orcinol Method to determine rhamnose concentration?
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Use bHPLC or GCMS for details
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I´m looking for the best method to concentrate 400L - 800L of fermentation:
- Centrifugation (different industrial machines)
- Lyophilization ?
- Any other suggestion?
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Tangential flow ultrafiltration is the best method for protein concentration and tangetial flow filtration can also help you with the biomass. Perhaps you can visit the sartorious web page to find more information about sizes and prices.
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Process analytical technology is applied in On-line HPLC which is used to measure the purity of biopharmaceuticals. The main keyword used in this article is POOLING. i.e. Online HPLC offers a feasible approach for analysis that can facilitate real-time decisions for column pooling. Feasibility of using ONline HPLC system for REAL TIME POOLING of process chromatography column. In the case of high resolution separation where a part of peak is being collected to pool the product and pool out the impurities.. Does anyone know what does the word pool refers to?
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Thank you very much Gabriel and Amir. Understood.
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