Your vedio is really helpful ❤️ I wish teachers also teach like this in their practical classes 👍 Please make an vedio on how to prepare graph if using standard as Ascorbic acid how toh interpret it.
thank you so much sir ... i am a reasearch scholar in NIPER institute...you have clear all concept very well.very informative video ..plz keep making such type of more video
@@Dr.Rakesh.Bone more help, I did little modification while preparing sample in a serial dilution 5- 1000 ug /ml. For eg. adding 1 ml of sample consist of 500microgram /ml conc. in 2 ml of methanol+ 1ml of DPPH ( 0.1 mM ) = in total 4 ml. Is it right way to evaluate Anti- oxidant ?
the video is really informative... keep going on.. this video helps me alot in understanding the basic of DPPH assay method.... thanks alot sir...................
Very well explained. Keep it up. God bless you dear. Understanding the concepts as important in any segment, is only related to explanation in a lucid and a simple way. And you have done it pretty well. Thank you for the same.
Hello Raki, i have greatly benefitted from your videos so far. Currently doing my Biochemical analysis part of my PhD in black rice. Regarding this DPPH assay, i have read many papers mentioning the use of Trolox as Standard reagent. What other more commonly available antioxidant chemical can i use for standard, other than trolox (which is pretty hard to find besides being costly). Thanks in advance.
Thank you so much Sir. For nicely explaining. One question please do answer 🙏. The concentration u mention 20,40,80 microlitre upto 3 ml each we hv taken. How much concentration in ug/ml for ic 50 calculation we mention them ?
Excellent videos as like your other videos ;-). I hv some doubts in it please clear; we have made blank and diff. conc. of standards but which among these is control absorbance.
Thank you very much You explained very well Please tell me how to convert the microlitres value of plant extract into micrograms/ml while drawing table and also how to calculate IC 50 values
Hello sir your vedios are nicely explained biochemical assay vedios we are so fond of that .sir can you please make a vedio on APX and MAD if possible.
Sir.... which solvent used for make up ascorbic acid........ only with methanol...?..or ....other solvents like ethyl acetate, chloroform etc. that used for extract preparation ?
I had only 1 extract methanolic extract. So I used only methanol. For standard use only methanol. For your extract use the same solvent as your extract u prepared.
thank you for your good video clips. they are really useful. but to some extent please speak slowly. english is not my tongue language. i couldnt understand some of your words. all in all your clips are very good
Not much, methanol rate of evaporation is less. So it's better to do it as fast as possible and store it in air tight bottles and preserve in refrigerator.
Your lecture was very helpful. The theoretical and experimental part was well articulated. Thanks alot!! I just had a doubt regarding the total volume, why the final volume was 4ml. Is it related to the concentration of DPPH and the sample ratio. Can you please clarify this.
Definitely it is the ratio of the both. More of sample and less of dpph or less of sample and more of dpph wouldn't help it so after many trial and errors earlier scientists have formulated this protocol. Nevertheless you can modify slightly. Thanks for those words :)
Sir, can we make our methanolic extracts of plant samples and store/keep them for 1 month @4 degree ? And secondly, do the dpph powder comes in red color also?
Thank you sir for your explanation but sir I am not clear in the result percentages it shows different absorbance 30.50 , 27 , 31 ,24.57 can you please tell..
very helpful video sir thankyu so much , please can you tell that when we say the enzyme SOD ,Catalze activity test as antioxidant so for that we only have to do that dpph test how can we say the SOD activity is?
Very informative video sir, Thank you. Sir If I use polysaccharides crude extract or silver nanoparticle polysaccharides extract, can I use same protocol??
Sir have u analysed hydroxyl radical scavenging assay......? If yes please suggest me the right protocol. I am unable to get the results..pink colour is not developed. Yellow colour is developed
Dear Sir, Thank you so much for the clear presentation. I have a doubt. Why is it necessary to have the DPPH assays read at 517 nm of spectrophotometer, What if it is maintained at 550 to 560 nm across all the samples, is it anything wrong? Please clarify.
@@Dr.Rakesh.B Thank you sir, I have one more doubt, it might sound silly, but please bear with me as I am just learning. I performed the DPPH assay with Ascorbic Acid and found that the absorbance value in spectrophotometer was low when compared with the plant extracts. So, is the lower absorbance value corroborating with the high antioxidant activity? Please clarify my doubt sir. Thanks in advance.
very helpful video sir, i am currently performing work in my ph.D on it please can you tell ..what if we have to determine the SOD ,Peroxidase enzyme activity ?
Thank you Sir. Your video is really well explained.Sir I have a doubt. Actually Sir I am using water extract of sample(water as solvent for the sample) . In that case my blank will be water? And for making the volume of sample upto 3 ml should I add water or methanol?
Hi sir, if I am using the leaf itself and not drying it, how would do assay be modified? This is because I understand you instructed us to dry it right. Thanks!
@@Dr.Rakesh.B Just one last question, for the standard, I used ascorbic acid, but I did not get any significant values, what can I do to modify this experiment?
@@Dr.Rakesh.B And a very last question, supposed I were to just take one reading of my leaf extract, and not require a serial dilution, how would the volumes of reagents used change?
Its all trial and error method. You have to work out in many possible ways and concentrations. If u have tried with 10, 20, 30, 40 50 and not getting good results increase or decrease it. For eg. 25, 50, 75, 100 or 100, 200, 300, 400,.500 like that
The volume of reagents will not change. They are same. Only thing if ur taking different dilutions using methanol u will make up to 2/3 ml. If ur doing with just 1 dilution, make it up to 2/3 thats all
Hello sir, ur videos are so helpful thank you. I have a doubt sir, In DPPH or phenolics/ flavanoid assays, you have used methanolic extracts. My extract is aqueous, can i follow same protocol, including blanks, dilutions? 1.) For dilution and blank can I use methanol too ( will that makes any difference, as my extract is aqueous extract and not methanolic extract) in DPPH assay. 2.) In phenolics- FC method, you have used water for dilution, though the extract is methanolic, so can i use methanol for dilution for aqueous extracts of mine ( in flavanoid-Al chloride method, where you have used methanol for dilution ) . Thank you in advance , sir
Yes you can use water to dilute in both the cases. It's not going to affect water or methanol bcoz we are just diluting it that's all. You can follow same procedure
Thank you Sir, nice video. But I have a doubt, if our compound donate electron to radical, our resulting compound will also become single electron species...so how we can ensure that radical is not formed again.
Helped me a lot during my PhD work
Your vedio is really helpful ❤️ I wish teachers also teach like this in their practical classes 👍 Please make an vedio on how to prepare graph if using standard as Ascorbic acid how toh interpret it.
thank you so much sir ... i am a reasearch scholar in NIPER institute...you have clear all concept very well.very informative video ..plz keep making such type of more video
One of the finest lecture I have ever seen God bless you sir 🤲🤲
Thank you ❤
You don’t know how much your detail explanation and video help me! Thank you so much for making this useful video!☺️❤️
Happy it helped you :)
@@Dr.Rakesh.B Sir, could you please send me the reference which u refered for DPPH activity.
Pls do watch a video just made for all the references in my channel.
@@Dr.Rakesh.Bone more help, I did little modification while preparing sample in a serial dilution 5- 1000 ug /ml.
For eg. adding 1 ml of sample consist of 500microgram /ml conc. in 2 ml of methanol+ 1ml of DPPH ( 0.1 mM ) = in total 4 ml.
Is it right way to evaluate Anti- oxidant ?
Thank god, i found this channel. Your explanation is superb.....
Thank you
Very informative sir...... U made each n every part clear....... God bless u
Thanks alot to u and my friend pavan for suggesting this video. Beacuse tmw I have my project presentation. Thank you ❤
Sir. Love you from Sri Lanka. I wish I could meet you one day in my life. Hats off
Thank you sir 🙏 Will come to Srilanka one day ❤️
Video is very helpful .... Nice teacher nd way of talking is good
Thank you so much brother.i got your video at right time it really help me a lot and you explain in very easy method and langauge ...thanks again
Happy to help
the video is really informative... keep going on.. this video helps me alot in understanding the basic of DPPH assay method.... thanks alot sir...................
Thank you so much, sir. It improved my understanding of DPPH a lot.
Very well explained. Keep it up. God bless you dear. Understanding the concepts as important in any segment, is only related to explanation in a lucid and a simple way. And you have done it pretty well. Thank you for the same.
Thank you so much means a lot:)
I dont have words to express my gesture of thanks to u extremely well explained!
Thank you means a lot 🙏
Thank you very much... Brilliant way of explaining the things❤
thank you so much for explaining in simple manner ..really helpful
Your videos are a boon to researchers and students sir....
Your explanations are so elaborate. Thank you for sharing.
Very helpful 👍👍 and nicely elaborated 💯👍👍
Thank you :)
hatts off... thanks for such a helpful video
Thanks for this amazing clarification. Do you have any published papers on this topic. I want to use them as references in my paper
Yes you can download my papers. I have made a separate video in my channel how to access my papers and cite references pls watch that
thank you so much sir, keep doing these kind of videos it will be helpful for research students
Sure :)
Great explanation...Rakesh. Best wishes...
Thank you mam 🙏
Thank you so so much sir..very sincere effort 🙏
Means a lot :)
Thank you it was really helpful for my project work
21:07
Very fast
Well done sir! Beautifully explained
Means a lot :) 🙏
Very well explained 👍👍👍
Thank you 🙏
Beautiful explanation in a very nice and easy way.
Thank you. Means a lot :)
Excellent explanation sir keep going this is very helpful for us..👍
Thank you:)
THANK YOU SO MUCH, well explained and so easy to understand 👍🏼👍🏼👍🏼
Means a lot :)
VERY VERY INFORMATIVE, USEFUL THANK YOU
nice video sir. Plz give the information about anti inflammatory activity.
Sir your videos are really very informative. Could you please provide link of the research paper you followed for this experiment
Do watch the video in my channel i made for all references
Hello Raki, i have greatly benefitted from your videos so far. Currently doing my Biochemical analysis part of my PhD in black rice. Regarding this DPPH assay, i have read many papers mentioning the use of Trolox as Standard reagent. What other more commonly available antioxidant chemical can i use for standard, other than trolox (which is pretty hard to find besides being costly). Thanks in advance.
Ascorbic acid
Butylated hydroxy toluene
Butylated hydroxy anisole
@@Dr.Rakesh.B thanks can i also use Quercetin as standard?
I haven't read any article where they used quercetin. If any article mentions it you can proceed
Sir shall we use Ascorbic Acid ?
@therebel5378 yes u can
Thank you very much for such a very informative video presentation.
Hi sir. thank you for this video it is VERY helpful.
Thank you so much Sir. For nicely explaining. One question please do answer 🙏. The concentration u mention 20,40,80 microlitre upto 3 ml each we hv taken. How much concentration in ug/ml for ic 50 calculation we mention them ?
Thank you, It is really very helpful.
Excellent videos as like your other videos ;-). I hv some doubts in it please clear; we have made blank and diff. conc. of standards but which among these is control absorbance.
You need to make a separate control without your extracts jst all chemicals
Absolutely fantastic! Just what I needed!
TYSMMM SIR your video helps me a lot!!!
Sir this vdo is really helpful . Also I could you please mention the paper that you have referred to for this procedure. Thank you
Please watch another video from Mt channel for the references
Ok sir . thank you
Very helpful thaaanku
Plz anti inflammatory activity bhi btain using bovine serum albumin
Amazing lecture sir... thanks a ton
Your welcome:)
Thank you very much
You explained very well
Please tell me how to convert the microlitres value of plant extract into micrograms/ml while drawing table and also how to calculate IC 50 values
Very informative video sir... kindly tell me one thing about the standard. It didn't understand that. How to prepare standard?
I have made a separate video on standard graph preparation. You can watch that from my channel for more information and better understanding
Thanks a lot for video... very helpful
It helps me lot thank for sharing
Hello sir your vedios are nicely explained biochemical assay vedios we are so fond of that .sir can you please make a vedio on APX and MAD if possible.
Sir.... which solvent used for make up ascorbic acid........ only with methanol...?..or ....other solvents like ethyl acetate, chloroform etc. that used for extract preparation ?
I had only 1 extract methanolic extract. So I used only methanol. For standard use only methanol. For your extract use the same solvent as your extract u prepared.
Sir, do we have to take the sample and the stabdard in triplicates?? Plz ans
Yes all should be done in triplicate or atleast minimum in duplicated
Hi if our extract is using water as solvent. So the blank is using water right?
thank you for your good video clips. they are really useful. but to some extent please speak slowly. english is not my tongue language. i couldnt understand some of your words. all in all your clips are very good
Great work man
Excellent explanation. Thank you very much.
Pls make a vdo on antioxidant activity by ABTS assay...
As we r making up volume with methanol it may evoporate and may cause error right
Not much, methanol rate of evaporation is less. So it's better to do it as fast as possible and store it in air tight bottles and preserve in refrigerator.
Very informative. Thank you. Is it okay if I use water infused plant extract instead of methanolic extract. Can I use the same protocol for DPPH ?
Yes you can use any extract.
Your lecture was very helpful. The theoretical and experimental part was well articulated. Thanks alot!! I just had a doubt regarding the total volume, why the final volume was 4ml. Is it related to the concentration of DPPH and the sample ratio. Can you please clarify this.
Definitely it is the ratio of the both.
More of sample and less of dpph or less of sample and more of dpph wouldn't help it so after many trial and errors earlier scientists have formulated this protocol.
Nevertheless you can modify slightly.
Thanks for those words :)
@@Dr.Rakesh.B can i get your email.. Need help in research
Rakeshb96@yahoo.com
Thanks for the video, could you tell me the relationship with DPPH and the Beer lambert equation is this needed for all DPPH assays?
Sir, can we make our methanolic extracts of plant samples and store/keep them for 1 month @4 degree ?
And secondly, do the dpph powder comes in red color also?
Yes you can store.
Dpph comes in 2 forms as per my knowledge red/orange crystals and pink
Thank you sir for your explanation but sir I am not clear in the result percentages it shows different absorbance 30.50 , 27 , 31 ,24.57 can you please tell..
Hello sir,
If plant extract is ethanolic then can we use methanol for DPPH solution preparation????
Check for papers on ethanol extracts what they have used for DPPH. I haven't tried .
Very good demonstration
Can you please provide the research paper for this protocol. It would be great help. Thankyou
Pls watch my video in my channel for references
This is very helpful..but please can you share the paper you have followed...
Mail me rakeshb96@yahoo.com
very helpful video sir thankyu so much , please can you tell that when we say the enzyme SOD ,Catalze activity test as antioxidant so for that we only have to do that dpph test how can we say the SOD activity is?
If possible kindly do a video on how to do statistical analysis......
Sure
Well explained sir
Sir u have taken reading on single beam uv spectrophotometer or double beans? Which one is best for this assay?
I have taken in single beam both is good
Thanks alot sir stay blessed
Very informative video sir, Thank you. Sir If I use polysaccharides crude extract or silver nanoparticle polysaccharides extract, can I use same protocol??
Yes you can use. We tried with nanoparticles it works.
For DPPH assay by Nanoparticles does depend on morphology of nanoparticles?
Your video is very helpful. but i have a question, can i just use DPPH as control? means i will not use methanol to add up to 3ml
It will be too concentrated and leads to wrong values (high values)
Sir have u analysed hydroxyl radical scavenging assay......?
If yes please suggest me the right protocol. I am unable to get the results..pink colour is not developed. Yellow colour is developed
It depends on the concentration. More concentration would reduce all dpph and make yellow colour. Reduce the concentration.
@@Dr.Rakesh.B sir i have done the dpph scavenging assay
Now I am telling about the hydroxyl radical scavenging assay...
Dear Sir,
Thank you so much for the clear presentation. I have a doubt. Why is it necessary to have the DPPH assays read at 517 nm of spectrophotometer, What if it is maintained at 550 to 560 nm across all the samples, is it anything wrong? Please clarify.
Many papers give different wavelength you can follow that wavelength given in your paper
@@Dr.Rakesh.B Thank you sir, I have one more doubt, it might sound silly, but please bear with me as I am just learning. I performed the DPPH assay with Ascorbic Acid and found that the absorbance value in spectrophotometer was low when compared with the plant extracts. So, is the lower absorbance value corroborating with the high antioxidant activity? Please clarify my doubt sir. Thanks in advance.
Yes low absorbance indicates high antioxidant activity
Dpph for essential oils? What should be solvent used?
very helpful video sir, i am currently performing work in my ph.D on it please can you tell ..what if we have to determine the SOD ,Peroxidase enzyme activity ?
Extremely helpful sir ..🙏🏻 what variations can be done in DPPH assay?
You can use different parts of plant, different extractions using different solvents, you can do in different concentrations too
Sir thank you so much. But sir I am unable to get the reference from your video can you please say for easy use
th-cam.com/video/S_hxotaRhwQ/w-d-xo.html
Sir, Very much Informative video. Can you provide the link of research paper for this procedure?
Please find video in my channel which gives details about the references
Hello i want the research article reference from which you follow this method.
Pls do watch a separate video in my channel i have uploaded for references
Thank you it was very helping. Can you please tell or share the link of the paper whose protocol you are following. Please
You can get access to my published paper. Find details in one of the video from my channel for references
Which method suitable for broth extracts of fungi and bacteria.
There are so many antioxidant assays. You can perform anything according to the chemicals you have
Thank you Sir. Your video is really well explained.Sir I have a doubt. Actually Sir I am using water extract of sample(water as solvent for the sample) . In that case my blank will be water? And for making the volume of sample upto 3 ml should I add water or methanol?
Yes your correct. Your solvent will be your blank so water is blank and you can use water to make up.
Ok Sir. Thank you.
Thanku Sir
Sir can we use gallic acid as a standard?
Yes
Hi sir, if I am using the leaf itself and not drying it, how would do assay be modified? This is because I understand you instructed us to dry it right. Thanks!
You can take 1g of fresh leaf and grind with 10 ml solvent. Filter and use the filtrate as extract. The protocol won't change.
@@Dr.Rakesh.B Just one last question, for the standard, I used ascorbic acid, but I did not get any significant values, what can I do to modify this experiment?
@@Dr.Rakesh.B And a very last question, supposed I were to just take one reading of my leaf extract, and not require a serial dilution, how would the volumes of reagents used change?
Its all trial and error method. You have to work out in many possible ways and concentrations. If u have tried with 10, 20, 30, 40 50 and not getting good results increase or decrease it. For eg. 25, 50, 75, 100 or 100, 200, 300, 400,.500 like that
The volume of reagents will not change. They are same. Only thing if ur taking different dilutions using methanol u will make up to 2/3 ml. If ur doing with just 1 dilution, make it up to 2/3 thats all
Hlo sir...Do this procedure can be followed for testing the antioxidant activity of emulgel? If so how?
Very helpful 🥰
superb explanation
Sir .are you from Kerala...?....very helpful channel 🙏🙏
No Bangalore. Thank you
Well explained. Please explain a bit slow. Thank you
Hello sir, ur videos are so helpful thank you. I have a doubt sir,
In DPPH or phenolics/ flavanoid assays, you have used methanolic extracts. My extract is aqueous, can i follow same protocol, including blanks, dilutions?
1.) For dilution and blank can I use methanol too ( will that makes any difference, as my extract is aqueous extract and not methanolic extract) in DPPH assay.
2.) In phenolics- FC method, you have used water for dilution, though the extract is methanolic, so can i use methanol for dilution for aqueous extracts of mine ( in flavanoid-Al chloride method, where you have used methanol for dilution ) .
Thank you in advance , sir
Yes you can use water to dilute in both the cases. It's not going to affect water or methanol bcoz we are just diluting it that's all.
You can follow same procedure
Can we follow the same protocol with Lactobacillis Cell Free Supernatant?
Excellent!
THANK YOU
Thank you Sir, nice video. But I have a doubt, if our compound donate electron to radical, our resulting compound will also become single electron species...so how we can ensure that radical is not formed again.
is it necessary to have methanolic antioxidant sample for scavenging test using method showing in video
No any extract can be used. But methanolic is advisable and many papers report the same
Can u make videos on copper reducing and ABTS radical scavenging assay as well pls
Sir can you please tell me which paper you prefer for DPPH assay, as i need it for citation purpose