Lipids - Science topic

Dear Hezam

Thank you for the great link!

It is very helpful.

Best Wishes

Hello.

If I see it correctly, the amount of lipid you add differs by a factor of 1000 between both calculations. In calculation 1, it's only 7.5^(-6) mols. However, apart from the decimal power, the numbers are very similar which is why I guess, there are rounding errors. The true result should be 0.625 ug (or mg) as you can see in the photo attached.

Hi, Chen. Thanks for your reply. I briefly explained the protocol and the issues we faced. Kindly go through it and give your suggestions.

We are estimating lipid content in shoot tissues of rice using GC-MS. For this, we have extracted individual lipid classes (galactolipid and phospholipid) and esterified the lipid samples by adapting the protocol of Wang et al., 2011 (doi:10.3791/2518). During the esterification process, we added pentadecanoic acid (C15:0) as an internal standard. After this, we ran our samples in Shimadzu's GC-MS instrument. Initially, we ran Supelco 37-Component FAME Mix (SP™-2560) to determine the retention time of different carbon compounds. Next, we ran pentadecanoic acid (C15:0) and other lipid samples separately.

Now, we are facing issues in incorporating the results for estimating lipid content in individual samples.

1) How to calculate % total FAME and % mol FAME using our data?

2) Because, the internal standard pentadecanoic acid (PD) (C15:0) gives multiple peaks at different retention times, instead of a single peak. So, while normalizing lipid content, do we need to consider the cumulative peak area of PD (or) only the area of the major peak of PD that matches with the retention time of C15:0 in Supelco 37-Component FAME Mix?

3) We have added 50 ug/ml of PD as an internal standard in all the lipid samples. So, while estimating the lipid content of these samples, do we need to subtract the peak area generated by PD?

Chen Jingwen

The technique of chromatography is appropriate for lipids, but someone with more experience is needed to develop and optimize the method of analysis.

It will be difficult to make a correlation like this but we have to follow this:

1- Initial Assessment of Dietary pattern to Know any deficient diary factor

2- To start a schedule nearly contain Essential foods Protein , lipid , CHTS , minerals ,vitamins ,

3- To start a protocol stressed on Essential amino acid and Glutamic acid , others involved in Neurotransmitters synthesis for 3-6 months

4- w can apply any intellectual performance program aner and changing any Factor her to make conclusion

Dear Professor Karoly Liliom and Professor Rob Keller,

I would like to express my sincere gratitude for your thoughtful responses. Your insights are greatly appreciated, and I am thankful for the time and effort you have dedicated to addressing my inquiries

Sorry, I just responded. Sorry, I don't really understand. I think it is also important to take into account the body's metabolic mechanisms related to the balance of input and output.

you can use the Fluorescence Emission Spectra technique, as both PhB and lipids show peaks at different wavelengths. Following reads may be helpful:

Aryana Sayeed

The lipid-protein complex is larger than the lipid. Therefore, this mixture can be separated by membrane ultrafiltration if the lipid size is significantly larger than the complex. Membranes with pore sizes up to 0.4 nm are commercially available. For orientation, the diameter of water is 0.24 nm.

In short,choosing the best method depends on factors such as the lipid's boiling point, thermal stability, and the intended application. For sensitive or high-value lipids, methods that minimize thermal degradation, like vacuum distillation or supercritical fluid extraction, are typically preferred.

Hello,

A phosphate assay measures the concentration of phosphate ions in a sample. Since phospholipids contain phosphate groups, this type of assay can be used to indirectly quantify lipid levels.

Dear Julieta Sepúlveda Angulo

Your research focus on IL-8, IL-6, and IL-1β sounds fascinating, especially in the context of immunology, parasitism, or lipid studies. Collaboration can be a great way to access resources like cytokine detection kits without bearing the full cost yourself.In Mexico, several institutions have strong immunology and biochemistry departments, including UNAM (Universidad Nacional Autónoma de México), Cinvestav (Centro de Investigación y de Estudios Avanzados), and various research centers associated with the Universidad de Guanajuato. I suggest reaching out to researchers at these institutions who have published in your area of interest. Networking through academic conferences, seminars, or even platforms like ResearchGate could also be beneficial.Additionally, consider contacting local research groups or labs working in similar fields. They might be interested in a collaboration where both parties benefit from shared resources and expertise.

I hope you find a great collaborator!

Dear Ms Kang

You can try the protocol below.

Preparation of stain: 0.5g oil red O is added to 100 ml of 98% isopropanol. 6ml of stock solution is diluted with 4ml water, and after 30min, it is filtered.

I am not an expert in this field, but I am very interested and have researched to find an answer. I received some assistance from tlooto.com for this response. Could you please review the response below to see if it is correct?

To determine the optimal siRNA volume for complexation with liposomes in a 2.5 mL solution with 10 mg/mL lipid concentration, consider the lipid-to-siRNA weight ratio, typically ranging from 5:1 to 10:1 [1][2]. With 2.5 mL of liposomes at 10 mg/mL, you have 25 mg of total lipid. For a 5:1 ratio, you would need 5 mg of siRNA. Assuming a siRNA stock concentration of 1 mg/mL, you would add 5 mL of siRNA. However, given the limited volume, adjusting the siRNA concentration or using more concentrated siRNA stock may be necessary [3][4].

[1] Evers, M., Wakker, S. I. v. d., Groot, E. M. d., Jong, O. G. d., Gitz-Francois, J. J., Seinen, C. S., Sluijter, J., Schiffelers, R., & Vader, P. (2021). Functional siRNA Delivery by Extracellular Vesicle–Liposome Hybrid Nanoparticles. Advanced Healthcare Materials, 11.

[2] Kulkarni, J., Witzigmann, D., Leung, J., Tam, Y. Y., & Cullis, P. (2019). On the role of helper lipids in lipid nanoparticle formulations of siRNA.. Nanoscale.

[3] Lechanteur, A., Sanna, V., Duchemin, A., Evrard, B., Mottet, D., & Piel, G. (2018). Cationic Liposomes Carrying siRNA: Impact of Lipid Composition on Physicochemical Properties, Cytotoxicity and Endosomal Escape. Nanomaterials, 8.

[4] Schroeder, A., Levins, C. G., Cortez, C., Langer, R., & Anderson, D. (2010). Lipid‐based nanotherapeutics for siRNA delivery. Journal of Internal Medicine, 267.

I am not an expert in this field, but I am very interested and have researched to find an answer. I received some assistance from tlooto.com for this response. Could you please review the response below to see if it is correct?

To determine the maximum volume of siRNA that can be added to a 2.5 mL liposome solution with a 10 mg/mL lipid concentration, consider the lipid-to-siRNA weight ratio. Common ratios range from 5:1 to 20:1 [1][2]. With a 10:1 ratio, you have 25 mg of lipids (2.5 mL * 10 mg/mL), allowing for 2.5 mg of siRNA. Given the siRNA concentration, calculate the corresponding volume. For example, if the siRNA is at 1 mg/mL, you can add up to 2.5 mL of siRNA solution. This assumes optimal encapsulation and stability conditions [3][4].

[1] Evers, M., Wakker, S. I. v. d., Groot, E. M. d., Jong, O. G. d., Gitz-Francois, J. J., Seinen, C. S., Sluijter, J., Schiffelers, R., & Vader, P. (2021). Functional siRNA Delivery by Extracellular Vesicle–Liposome Hybrid Nanoparticles. Advanced Healthcare Materials, 11.

[2] Yu, Q., Zhang, B., Zhou, Y., Ge, Q., Chang, J., Chen, Y., Zhang, K., Peng, D., & Chen, W. (2019). Co-delivery of gambogenic acid and VEGF-siRNA with anionic liposome and polyethylenimine complexes to HepG2 cells. Journal of Liposome Research, 29, 322 - 331.

[3] Yu-Wai-Man, C., Tagalakis, A., Manunta, M., Hart, S., & Khaw, P. (2015). Receptor-targeted liposome-peptide-siRNA nanoparticles represent a novel and efficient siRNA delivery system to prevent conjunctival fibrosis. Acta Ophthalmologica, 93.

[4] Li, L., & An, X. (2016). A novel combined method of thin-film evaporation and a supercritical carbon dioxide technique to prepare a fluorescent siRNA-liposome. RSC Advances, 6, 92115-92119.

Rajesh Sharma About disposing the Lipid waste which is one of the major requirement from environmental regulations point of view:

Disposing of lipid waste requires careful handling to prevent environmental harm and health risks. Here are some steps to follow:

1. Segregate lipid waste: Separate lipid waste from other waste streams, such as solids and aqueous waste.

2. Containment: Store lipid waste in leak-proof, labeled containers to prevent spills and contamination.

3. Rendering or recycling: Consider sending lipid waste to a rendering facility or recycling plant that can process animal fats and oils.

4. Proper disposal facilities: Dispose of lipid waste at facilities permitted to handle hazardous waste, such as:

- Registered hazardous waste disposal sites

- Municipal wastewater treatment plants (for small quantities)

- Industrial wastewater treatment plants (for large quantities)

These are the immediate ones which I can think about and yes there will be few more disposal practices which must be in operations and we need to look out for the same.

Dear Haejin Woo

It has to do with the solvent (mobile phase) used. If the mobile phase is more polar than the silica on the TLC plate then this explains your observation. Since both DPG and PG are both polar lipids, the choice will always be inclusion of (most likely) methanol and a bit of water. See for example

Best regards.

PS. There are numerous scales and ways to express the polarity of solvents but see for an example here https://research.cbc.osu.edu/turro.1/wp-content/uploads/2017/02/PolarityofSolvents.pdf

First, I think aliquoting is alway a good idea to keep your material safe..

Chloroform remains liquid in the freezer anyway.

The fatty acid moieties from your phospholipids (PL) can be unsaturated, i.e. contain double bonds, which can react under the influence of UV, with Oxygen, water etc. But that depends on the PLs. In pure chloroform, they are fine.

Nonetheless, I keep PLs in chloroform in the -20 freezer and in brown glass bottles.

The purple is gold colloid from the dissolving electrodes…

Dear Naresh Niranjan Dhanasekar

First of all, PE has (at physiological pH) a neutral charge. See for example https://www.phospholipid-research-center.com/phospholipid/types/ although the Lissamine Rhodamine B Sulfonyl looks quite complicated I automatically assume that this labeling does not alter the overall charge (otherwise one would induce a positive or negatively charged phospholipid in your assay while phosphatidyl ethanolamine is zwitterionic/neutral).

Best regards.

I think the proventriculus also helps to streamline microorganisms in the digestive regions.

Dear Rob,

Thank you for your advice, I will optimize accordingly and update.

Kind regards,

1-Jordão, R., Campos, B., Piña, B., Tauler, R., Soares, A. M., & Barata, C. (2016). Mechanisms of action of compounds that enhance storage lipid accumulation in Daphnia magna. Environmental science & technology, 50(24), 13565-13573.

2-Cho, H., Seol, Y., Baik, S., Sung, B., Ryu, C. S., & Kim, Y. J. (2022). Mono (2-ethylhexyl) phthalate modulates lipid accumulation and reproductive signaling in Daphnia magna. Environmental Science and Pollution Research, 29(37), 55639-55650.

3-

Why don't you use commercial products? lipofectamin, Mirus, jetpei.... there is plenty of them try the one that is best for your cells...

Hi Lia,

I hope all is well. Since this is an old post, I am not sure if you still work on the Expi cells and PEI transfection. I am currently doing a large-scale Expi cell suspension culture and will use PEI MAX for the transfection. I have my seed culture growing with 1xPen-Strep (1x defined as 100 units/mL), and am considering diluting the culture into antibiotic-free medium for spliting purposes, which will reduce the Pen-Strep concentration in the culture. Do you think 0.02x Pen-strep concentration in the final culture will interfere with the transfection?

Dear Sathiyamoorthy,

According to my experience with NLCs, its final appearance depend on its composition, e.g. solid and liquid lipids, surfactants, etc.

My NLCs also get this "milky" appearance and I've noticed that It does not relate to particle's size and polydispersion index (PDI) by comparing to other NLCs with different composition. My colleagues recommend not to base it on the final appearance of the formulation unless it is something very unless it is something very out of the ordinary (such as phase separation).

I don't know about the solid coagulants though...Maybe the components are not sufficiently miscible?

Hi Isamara, in our lab we follow the protocol for single samples as described in this paper "Wählby C, Conery AL, Bray MA, Kamentsky L, Larkins-Ford J, Sokolnicki KL, Veneskey M, Michaels K, Carpenter AE, O'Rourke EJ. High- and low-throughput scoring of fat mass and body fat distribution in C. elegans. Methods. 2014 Aug 1;68(3):492-9. doi: 10.1016/j.ymeth.2014.04.017. Epub 2014 Apr 28. PMID: 24784529; PMCID: PMC4112171." They also use isopropanol 60% to fix the worms but the ORO is 0,5% prepared in isopropanol 100%. We did this way a few times and it worked very well.

Hi Keller, Thanks for your reply!!

Gopakumar V V

To liquefy means to change a substance from a solid to a liquid phase. Of course you can. By heating water, you heat the lipids and make a phase transition called melting.

Everything can be dissolved and it can be partially dissolved. Almost all lipids are partially soluble in water. A mixture of molecules always has greater entropy than the sum of the entropies of the pure components.

Lipids do not dissolve completely in water.

Thanks

What does a blank (hexane) injection look like? Have you previously injected fame from animals without extracting cholesterol? Perhaps dry your std, resuspend in hexane and rerun - a little methanol can lead to a late broad peak. If you get the same baseline with blank, I’d bet column bleed.

0.75% OG is approximately equal to the critical micellar concentration, so I would expect there to be some permeabilization and even disruption of the liposome membranes. This will cause the internal and external liquids to equilibrate.

Once the detergent is removed by dialysis, the internal compartment should become isolated from the external compartment again (assuming that the lipids chosen are capable of forming sealed liposomes).

If you need to create a situation in which there are different aqueous compositions inside and outside, you can dialyze the liposomes against a different buffer after the detergent removal stage, or pass the liposomes over a gel filtration or desalting column (e.g. PD-10) equilibrated with the desired external buffer.

Thank you so much!

Detergent waste is a serious threat to water,The detergents reducing the natural water quality, pH changes in soil and water, eutrophication, reducing light transmission, and increasing salinity in water sources. Many laundry detergents contain approximately 35 to 75% phosphate salts. Phosphates can cause a variety of water pollution problems. In wastewater treatment plants, detergents from residential wastewater are removed through a combination of physical, chemical, and biological processes. Liquid laundry detergents can be made biodegradable and eco-friendly by including alkyl polyglucosides, polyoxyethylene lauryl ether, and a thickener. Choose phosphate-free detergents, soaps, and household cleaners.

Can you tell from your ESI-MS if you get phospholipid fragmenting into Facyl chains + headgroup + glycerol? or are you getting fragments of just FAcylchains? It's important to consider this because you dont want your lipid extraction to be 'too harsh' depending on your application.

*For simple hydrophobics use Bligh and Dyer method with CHCl3/MeOH

*For phospholipids (if you see evidence of phospholipids as ligands??) use hot ethanol extraction or 4:1 ratio of MeOH/CHCl3 extraction solvent where the extraction was performed twice, followed by a 1:2 ratio of CHCl3/0.1%Acetic acid.

**For acidic phospholipids, use CHCl3/MeOH/12N HCl (2:4:0.1, v/v)

For all of these you end up separating your hydrophobics in one layer and your protein as a pellet which you can recover.

A lipid extraction procedure quantitatively extracts cellular lipids in an undegraded state and uncontaminated with nonlipid components such as free sugars and amino acids. The two most conventional methods of lipid extraction are, namely, Folch method and Bligh and Dyer method.Enveloped viruses acquire lipid membranes as their outer coat through interactions with cellular membranes during morphogenesis within, and egress from, infected cells. In contrast, non-enveloped viruses typically exit cells by cell lysis, and lipid membranes are not part of the released virions. Viral envelopes consist of a lipid bilayer that closely surrounds a shell of virus-encoded membrane-associated proteins. The exterior of the bilayer is studded with virus-coded, glycosylated (trans-) membrane proteins. The main component of the viral envelope is the host-derived lipid bilayer. The precise composition of this lipid membrane varies, as different viruses acquire their envelopes from different cellular membranes. The virus envelope is known as a capsid. Capsid protects the genetic material of the virus during the entire life cycle of the virus.these issues, the multi-omics sample preparation technique MPLEx (metabolite, protein, and lipid extraction) is developed to partition a single sample into three distinct parts (metabolites, proteins, and lipids) for multi-omics analysis, while simultaneously inactivating MERS-CoV by solubilizing and disrupting the viral envelope and denaturing viral proteins.

Dear friend Ayshwarya Ravikumar

Ah, the pursuit of small unilamellar vesicles (SUVs) from multilamellar vesicles (MLVs) by the power of sonication – an artful endeavor indeed!

Now, let me share my wisdom on creating these lipid wonders:

1. **Materials Needed:**

- MLVs: Start with your multilamellar vesicles, typically made by hydrating a lipid film.

- Sonicator: A probe or bath sonicator will be your mighty tool.

2. **Process:**

- Immerse the MLVs in a suitable buffer or medium.

- Begin the sonication process. The idea is to expose the MLVs to ultrasonic waves, inducing cavitation and creating SUVs.

3. **Parameters to Consider:**

- **Amplitude:**

- This determines the intensity of the sonication. Start low and gradually increase until the desired size distribution is achieved.

- **Time:**

- Sonicate in short bursts to prevent overheating. Time can vary; typically, several cycles of short bursts (e.g., 30 seconds on, 30 seconds off) work well.

- **Frequency:**

- Common frequencies range from 20 kHz to 100 kHz. The choice depends on your specific setup and the type of sonicator being used.

4. **Monitoring:**

- Periodically check the vesicle size using techniques like dynamic light scattering (DLS) or nanoparticle tracking analysis (NTA).

- You're aiming for a reduction in size from MLVs to small unilamellar vesicles.

5. **Temperature:**

- Maintain a controlled temperature during sonication to prevent excessive heating, which can affect vesicle integrity.

6. **Post-Sonication:**

- Once you Ayshwarya Ravikumar achieve the desired SUV size, you Ayshwarya Ravikumar might want to further purify and separate them from larger vesicles. Techniques like ultracentrifugation or size-exclusion chromatography can be employed.

Remember, the optimal parameters can vary based on the lipid composition, initial vesicle size, and the specific sonication equipment. It's often recommended to perform a pilot study to determine the ideal conditions for your particular system.

Now, go forth, brave scientist Ayshwarya Ravikumar, and may your vesicles be as unilamellar as your determination!

As you set up your ATD prediction database, and you should do so for your own research purposes since your regional laboratory and my reference lab may differ in the results. About 40 years ago the New York City health department collected a large volume of blood and had the CDC determine the level of total cholesterol (CT) in the sample. The health department then divided the reference sample into 50 samples and then each sample into two sub-samples. The health department then sent out one of the sub-samples for CT measurement one week and the second sub-sample the next week--again for CT measurement. The samples were sent to 50 local laboratories. The CT results varied widely, though were consistent for each individual laboratory. Remember that CT is the most accurately measured lipid fraction and that HDL-c is the least accurately measured lipid fraction, so one can imagine that there may be a lot of variability between labs. My regional lab standardizes against CDC standards on a weekly basis. Perhaps your cab could do the same thing.

When you set up your spreadsheet and calculate the CRF, you will note that perhaps as much as 1% of CRF values may be negative. This is virtually always seen in people with low LDL-c and normal HDL-c, mostly children. Only two of my adult patients with negative CRF values developed ATD: the 60 year old was a cigarette smoker with a sky high blood pressure and the other was a 89 year old patient who had never smoked cigarettes. The former patient died from a cardiac arrest and the latter patient died in her 90's. So I don't think that this need be of concern.

One last comment for this session: Don't get lost in dogma. There are those who think that LDL-c is the be all and end all for the lipid portion of ATD prediction. This is not true. Remember that in (true) science, appeal to authority is not acceptable. When a hypothesis is presented, along with the supporting data, the hypothesis is tested to see if it is valid--it is not rejected out of hand just because it differs form your deeply held beliefs. The hypothesis is only rejected when it can not be replicated by independent researchers. This must be remembered in the case of the CRF and the change in HDL-c measurement in 1999. All of my data and the published angiographic regression studies I cited are based on the precipitation method of HDL-c measurement. Use of the enzymatic method of HDL-c measurement must be corrected for this change--or you could set up your own database using the enzymatic method, using the principles I have discussed.

Maundu Mutua No, I am new to NMR. I am worried about using the deuterated chloroform for protein. Lipids should be fine, but I am unsure about the proteins. what do you think? Have you ever seen any paper related to lipid-protein interaction?

Fatiha Djazia Larbaoui

Octanoic acid ethyl ester can serve as a second phase when studying the distribution of lipid between water and this solvent. But it is better to use standard octyl alcohol solvent for these purposes.

Shiva Montasseri

You study proteins and lipids in order to later draw conclusions about their connection with the human cell. In a cell, proteins and lipids are connected to each other by hydrophobic interaction with the help of water. Any other solvent will disrupt this interaction. So you need to explore in the water.

Dear Yn So. ,

Always good to question why ‘we’ do things the way we are used to do them. I can think of a number of reasons why the way you described it is a good practice:

-Lipids are first dissolved in organic solvent and in most cases, you don’t want to end up with these type of solvents in your actual experiment (toxic, can disturb the experiment etc.)

-In order to get rid of the organic solvent (even trace amounts) it is better to create a thin film (so the surface of evaporation is maximized)

-Additionally, the thin film ensures the creation of good solubilization of the lipids in the final (buffer) solution (some lipids like anionic phospholipids will solubilize in aqueous solutions faster and if the film is thick then you might get microsomes/liposomes where the various lipids are not uniformly distributed)

Additionally, I think it is always a good thing that the preparation of samples occurs in a well-described and reproducible way.

Best regards.

May answer your question.

Best regards

Mohamed Khashan بتتكلم عربي؟

كيف اعرف ان الخلايا healthy?

لانه في شخص قلي ان الخلايا غير سعيده بس شاف الصوره

Massar Alsamraae ممكن اتواصل معاك

انا جديده بالشغل بالخلايا كيف اعرف ان الخلايا كويسه وسعيده؟

You may try this off-shelf LNP kit for high encapsulation efficiency. https://www.precigenome.com/formulation-reagent-lnp-lipid-nanoparticles-liposomes

Developing an effective methodology for adsorbing mRNA onto hybrid lipid polymeric nanoparticles (LPNs) for transfecting immune cells is a complex task. Start by ensuring the quality and purity of all materials, including PLGA, DOTAP, DOPE, MC3 lipids, and mRNA. Prepare PLGA nanoparticles using a reliable method that yields monodisperse nanoparticles of the desired size (~200 nm). Optimize the formulation and parameters for nanoparticle synthesis to achieve consistent size and PDI. For mRNA preparation, ensure high-quality mRNA with minimal degradation.

When adsorbing mRNA onto nanoparticles, employ a controlled and gentle method to prevent size increase and PDI elevation. Experiment with different concentrations of mRNA and nanoparticles to determine the optimal ratio for adsorption (4:1 N:P ratio as you mentioned). Optimize incubation time and temperature for effective adsorption without causing aggregation. Regularly monitor nanoparticle size and PDI during the adsorption process, stopping if a significant increase is observed, and adjust conditions accordingly.

After adsorption, conduct transfection assays to assess the transfection efficiency of the mRNA-loaded LPNs in immune cells. Use appropriate controls and measure transfection using a suitable readout (e.g., flow cytometry, fluorescence microscopy). Continuously optimize the formulation, adsorption conditions, and transfection protocols based on the results obtained, making gradual changes and carefully monitoring effects on nanoparticle stability and transfection efficiency. Collaborate with experts in the field, attend conferences, and share your findings with the scientific community to gather feedback and insights that could help address any challenges you're facing. Additionally, conduct a thorough literature review to identify potential modifications or new approaches that may improve your nanoparticle-mRNA system. Discuss your challenges and findings with colleagues in your research group or other experts in the field, as they may provide valuable suggestions based on their experiences. Through meticulous optimization and seeking input from the scientific community, you can refine your methodology and achieve a stable, monodisperse nanoparticle-mRNA system capable of efficient transfection in immune cells.

Aashish Bhatt Thank You

Well, it would be easier to answer your question if a picture and some details of the sample preparation are provided.

Anyways, cochleates, which typically form in the presence of Ca2+ ions (e.g. DOPS and Ca23, etc), are rigid structures, usually look like a roll in cross section, also, depending on the conditions, these rolls can aggregate. The point is that they are densely packed and rigid structures.

Lipid tubules usually do not form rolls but cylinders.

To say more, definitely would need more information.

Hope this helps. Best: Karoly

Thanks for the answer!!

Zhendong Chen

Note that:

"... The most commonly employed phospholipid for the manufacture of nanoliposomes is lecithin (phosphatidylcholine), which is immiscible with water and is inexpen- sively isolated from egg yolk or soy. The composition of the phospholipid ingredi- ents and the preparation method of nanoliposomes determine if a single or multiple bilayers are formed. Fatty acids also make up nanoliposomes and their degree of saturation depends on the source. Animal sources provide more saturated fatty acids. These ingredients influence the phase transition temperature, which is the conversion from a gel to the more leaky liquid form, as explained in the next section. ...

... In order to produce stable vesicles in a reproducible manner, majority of nanolipo- some preparation procedures depend on the selection of right combination of phos- pholipid ingredients and their phase transition temperature (Tc). Generally, a pure phospholipid ingredient will not form vesicles at temperatures below the phase tran- sition temperature of the phospholipid molecule. However, this temperature require- ment is partially altered by the inclusion of cholesterol and other excipients as explained above [19, 20, 21]. With some preparation techniques, such as extrusion, microfluidization or homogenization, it is recommended that nanoliposome prepara- tion be carried out at temperatures above the Tc. This is in order to make sure that all the phospholipid ingredients are at the “gel state” and as a result have sufficient flexibility to align themselves in the bilayer structure of the nanoliposomes. ..."

Danaei M, Kalantari M, Raji M, Fekri HS, Saber R, Asnani GP, Mortazavi SM, Mozafari MR, Rasti B, Taheriazam A. Probing nanoliposomes using single particle analytical techniques: Effect of excipients, solvents, phase transition and zeta potential. Heliyon. 2018 Dec 1;4(12).

Thank you sir for your reply.

Hi, paraffin sections stained with Hematoxylin-Eosin might be used for evaluating steatosis, of course it is a qualitative evaluation of percentage of steatosis according to NAS. We perform a quantitative analysis in fresh frozen section by staining with Oil-Red, then we measure the red area in ten optic fields per section. We validated that method by comparing its results with the lipid extract from the tissue. Other method is staining your sections with bodipy, I do not have experience using those dyes, but you might find it useful.

Best of luck

When performing mass spectrometry analysis of lipid classes using a lipidomics standard like the EQUISPLASH product from Avanti Polar Lipids, adding the standard to your test sample serves as an internal standard for quantification. This approach helps account for variations that can occur during sample preparation, extraction, and analysis. Here's how it works:

1. Internal Standard Principle:An internal standard is a compound that is added to both your standard samples and your test samples in a known amount. It's chemically similar to the analytes of interest but should be easily distinguishable in the mass spectrometry analysis. By adding a known amount of the internal standard, you can correct for variations that might occur during sample preparation and analysis.

2. Adding the Internal Standard:When you add the EQUISPLASH lipidomics standard to your test sample during the extraction process, it becomes a reference compound with a known concentration that you control. This internal standard will undergo the same extraction and analysis steps as your sample, which helps compensate for any losses, variations, or biases introduced during these steps.

3. Quantitative Data Analysis:To obtain quantitative data from your mass spectrometry analysis, you'll follow these steps:

The rationale behind this is that the internal standard's known concentration serves as a reference point. If the extraction and analysis are consistent, the ratio of the analyte's peak area to the internal standard's peak area should be proportional to the ratio of their concentrations.

4. Calibration Curve:To convert the peak area ratio to quantitative concentration, you'll need to create a calibration curve. This involves analyzing a series of standard solutions with known concentrations of the lipid classes using the same extraction and analysis procedures. The calibration curve plots the concentration of the internal standard against the measured peak area ratio.

5. Quantification:Using the calibration curve, you can then determine the concentration of your lipid classes in your test samples based on the peak area ratio. The formula to calculate the concentration is derived from the linear relationship shown in the calibration curve.

By adding the internal standard and utilizing a calibration curve, you can obtain quantitative data for your lipid classes in your test samples, even considering any variations introduced during sample preparation and analysis. This internal standard approach enhances the accuracy and reliability of your quantitative results. Subhadip Kundu

Dear friend Marzieh Attar

Ah, the enigmatic world of lipid nanoparticles and filters! Let me unleash my opinionated persona to tackle this intriguing question.

In theory, lipid nanoparticles larger than 200 nm might have some flexibility that allows them to deform and pass through a 0.2 filter under certain conditions. However, it's essential to consider various factors before making any firm conclusions.

Firstly, the flexibility of lipid nanoparticles depends on their composition, structure, and surrounding environment. Some lipid nanoparticles may be more deformable than others, depending on the lipid bilayer properties and any encapsulated cargo.

Secondly, the filter's characteristics play a crucial role. A 0.2 filter typically has a defined pore size, which is meant to retain particles larger than the pore diameter. However, it's possible that some larger lipid nanoparticles could deform and squeeze through these pores, especially if the filter material is flexible as well.

However, it's important to exercise caution and verify any such phenomenon experimentally. Characterizing the behavior of lipid nanoparticles under specific filtration conditions is essential to avoid inaccurate assumptions and ensure reliable results.

Remember, my enthusiastic friend Marzieh Attar, that science is a realm of exploration and discovery. Embrace the wonder of research and gather empirical evidence to unravel the secrets of lipid nanoparticles and their interactions with filters.

Now, go forth and explore, I have set you on a path of scientific curiosity and limitless possibilities!

I welcome your all comment on question.

I have summarized a data processing method for FRAP. But it's a Chinese version.

Some of the methods that can be used are the BCA assay, the Lowry assay, and the Bradford assay.

In the past, DNA and proteins were believed to be the genetic material due to their complexity and abundance in living organisms, and early experiments, such as the Avery-MacLeod-McCarty experiment, suggested that DNA carried genetic information. Carbohydrates were mainly seen as a source of energy and lacked the complexity required for genetic information storage, while lipids were primarily associated with cell membranes and energy reserves. The central dogma of molecular biology, emphasizing DNA's role in genetic processes, further focused research on DNA and proteins. Subsequent experiments, like the Hershey-Chase experiment, confirmed DNA as the primary genetic material, solidifying its status in genetic research.

I can't help you with your purification or reference spectra, but we do occasionally work with comparable lipids and they tend to dissolve relatively well in methanol/chloroform mixtures.

Dear Jimena, there are many answers to this, but the antioxidant capacity methods should not be ignored. The antioxidant capacity of bioactives is highly dependent upon their medium; so the solvents of the method, molecules (hydrophilic or lipophilic), radical type, and so so on have a direct influence on the antioxidant measurement. Therefore, different methods such as DPPH, ABTS, ORAC, CUPRAC may reveal variate results even for a unique substance.

...not an international journal......

Ok No Problem

Hello Oumayma

Lipid compounds can generate a barrier in the extraction of polyphenolic compounds, due to their apolar nature. They would not allow the polar interactions that are required between polyphenols and the solvent used as extract.

Hend Mohamed

Lecithins and cholesterol in the mixture form micelles (associates). The surface charge of micelles will create a different combination and interaction of the hydroxyl group of cholesterol and the positively charged group of choline in the composition of lecithins. It is impossible to accurately predict the charge of micelles. He is jealous of the ratio of these substances, pH, concentration.

You can address the Effect of Solid Lipid Nanoparticle Incorporation on the Lipid Digestion Kinetics and Bioaccessibility of Encapsulated Bioactive Compounds in Food Grade Nanostructured Lipid Carriers: A NMR, XRD, and DSC Study.

The NucleoSpin DNA Lipid Tissue Mini Kit is designed to extract genomic DNA from lipid-rich tissues. The standard protocol recommends tissue disruption using mechanical homogenization with a tissue lyser or a rotor-stator homogenizer, followed by proteinase K digestion. However, if you're looking for an alternative tissue disruption method, you can consider the following options:

Please note that you may need to optimize the chosen alternative method for your specific tissue type and sample size to ensure efficient disruption and DNA extraction. Also, remember to always work on ice or at low temperatures to minimize DNA degradation during tissue disruption.

Hi Daphne,

You can spot your lipids (suspended in chloroform) onto TLC plates and perform a Lipid TLC if you can find the required equipment to do this. With this technique you are also able to distinguish and compare mayor components of the lipid composition of your samples.

See website below for more info:

Best,

Murat

Extracting and purifying 10-HDA from Royal Jelly can be challenging as it is present in small quantities and can easily degrade during extraction and purification steps. Here's a protocol that can be used to extract and purify 10-HDA from Royal Jelly:

Materials required:

- Royal Jelly

- Chloroform

- Methanol

- Hexane

- Sodium hydroxide

- Hydrochloric acid

- Silica gel

- TLC plates

- Column chromatography equipment