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Making positive charged polyacrylamide

Making positive charged polyacrylamide


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I am interested in positive charged polyacrylamide to electrophorese molecules I am interested in.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2643323/ http://onlinelibrary.wiley.com/doi/10.1002/bip.360300309/pdf

The articles shown above use allyltriethylammonium bromide (ATAB), but I am not sure which chemical they mention.

Or is there any other chemical to add to acrylamide to get positive charged polyacrylamide copolymer?


ATAB contains a quaternary amine and a terminal carbon-carbon double bond. The polyacrylamide reaction links acrylamide by their double bonds forming the polymer. The double bond on ATAB can be incorporated into the growing polymer chain, and the quaternary amine holds a positive charge, making a positively charged polymer.

The immobilines mentioned are probably just a weak acid or weak base group with a terminal double bond like ATAB or acrlyamide. They get trapped into the polymer and if you pour the gel with a pH gradient, they pick up charges according to the pH. I haven't found a good concrete immobiline example, I can't access that second article you linked.

I am away from my good computer and cannot make chemical structures right now.


1. What is polyacrylamide PAM?
Polyacrylamide is a polymer (-CH2CHCONH2-) formed from acrylamide subunits, it is long-chain polymer (same molecule repeating itself many times) designed to attract either positively charged particles (organic materials, such as carbon or human waste) or negatively charged particles (inert materials, such as sand or clay). The abbreviation of polyacrylamide is PAM, it is a chemical which purchased in dry, emulsion, liquid and tablet form.
Polyacrylamide can be carbonated into black powder at 210οC without oxygen. Polyacrylamide could be made to four series,e.g., non-ion polyacrylamide,zwitterionic polyacrylamide, cationic polyacrylamide CPAM, and anionic polyacrylamide APAM.
Polymers may be purchased in dry, emulsion, liquid and tablet form. These chemical compounds are used to flocculate and coagulate suspended solids in water, wastewater, and soil. They assist in management of the Earth’s soil and water. In the straight-chain form, it is also used as a thickener and suspending agent.

2. How many types of polyacrylamide? What is the difference between them?
There are three types of polyacrylamide: APAM, CPAM, and NPAM.We also provide acrylamide.
Anionic polyacrylamide APAM :
This type of polymer has molecules that carry negative charge. Anionic polyacrylamide can pick up positively charged particles (clay, sand), much like a magnet picks up nails and other metal objects. There are over 100 varieties of this type of polymer. Anionic polyacrylamide has no aquatic toxicity. It is recommended for use in furrow irrigation, dust control, crop dusting, treating wholesale nursery & stormwater runoff, hydroseeding, animal waste treatment, construction projects, sports fields, landscaping, turf & sod, drilling mud, mining, and water & soil conservation.
Cationic polyacrylamide CPAM :
This type of polymer has molecules that carry positive charge. Cationic PAM can pick up negatively charged particles (organic materials like carbon or human waste). There are over 1000 varieties of this type of polymer. It is recommended for use in wastewater plants, animal waste treatment, water clarification, drinking water, and many industrial applications, such as mining and paper processing. Many hours of testing are often required in these applications to determine the correct polymer choice.
Non-ionic polyacrylamide:
This type of polymer has molecules with no charge. Non-ionic polyacrylamide are used in very rare instances and special circumstances only. This polymer is used mostly in mining.

3. What are the uses of polyacrylamide?
Flocculate or coagulate solids in a liquid may be one of the largest uses for polyacrylamide. This process applies to wastewater treatment, and processes like paper making. Most polyacrylamide is supplied in a liquid form. The liquid is subcategorized as solution and emulsion polymer. Even though these products are often called 'polyacrylamide', many are actually copolymers of acrylamide and one or more other chemical species, such as an acrylic acid or a salt thereof. The main consequence of this is to give the 'modified' polymer a particular ionic character.
Another common use of polyacrylamide and its derivatives is in subsurface applications such as Enhanced Oil Recovery. High viscosity aqueous solutions can be generated with low concentrations of polyacrylamide polymers, and these can be injected to improve the economics of conventional waterflooding.
It has also been used for horticultural and agricultural use under trade names such as Broadleaf P4, Swell-Gel and so on. The anionic form of cross-linked polyacrylamide is frequently used as a soil conditioner on farm land and construction sites for erosion control, in order to protect the water quality of nearby rivers and streams.

4. How is polyacrylamide applied?
Three most common forms of polyacrylamide are dry granules, solid blocks (cubes), and emulsified liquids. The application method of polyacrylamide chosen depends on the form of polyacrylamide selected.
The use of dry granular polyacrylamide into irrigation water is facilitated by the use of an augured metering system and excellent mixing and thorough dissolving before the polyacrylamide reaches the irrigated furrows. In order for the polyacrylamide to dissolve properly in the irrigation ditch it must have proper agitation. Unlike sugar or salt which dissolve fairly quickly in water, granular polyacrylamide needs to be agitated thoroughly in order for it to dissolve.
Polyacrylamide blocks (or cubes) are usually placed in wire baskets that need to be secured to the edge of the ditch to avoid washing of the blocks down the ditch. In a concrete ditch, tins or boards will provide sufficient turbulence.
Liquid polyacrylamide can be metered directly from the container into the irrigation ditch, directly into the furrow, or through a pipe line or injector pump.
If you are interest in and want to know more about polyacrylamide, please come and click our website, you will find what you want.

5. Why would people want to use polyacrylamide?
PAM Polyacrylamide is highly effective in reducing soil erosion off of fields and can increase water infiltration into irrigated furrows. It has been shown to significantly that polyacrylamide reduce soil erosion by 90-95 percent when applied to irrigation water. Increases in water infiltration rates vary from 20-60 percent from trials and experiments listed below in the "links" section. The increased use and distribution of polyacrylamide products in the past few years has brought down product prices, making polyacrylamide a more economical BMP option. Polyacrylamide's many forms and application techniques make integration into the farmer's irrigation routine smooth and relatively easy once the initial set-up is complete. Relatively low cost, high reduction of irrigation-induced erosion and soil loss, ease of use and integration, make polyacrylamide a best management practice worth looking into by any agricultural operation.

6. Why would people prefer to use granular polyacrylamide to liquid polyacrylamide in irrigated furrows?
An experiment have done to test on two different application techniques of polyacrylamide (polyacrylamide liquid and polyacrylamide granular) showed both reduced sediment loss and increased water infiltration into the soil. The experiment was designed to determine if granular polyacrylamide could be as effective at reducing erosion in furrows when applied starting at the beginning of the head ditch (where it has not yet thoroughly dissolved) as when applied to the furrows further down the head ditch.
The two forms of polyacrylamide were supposed to be applied at similar rates, but liquid polyacrylamide ended up being applied at a rate of 0.9 lb/acre and the granular polyacrylamide at a rate of 1.8 lb/acre. The difference was caused by the changes in volume of water flowing in the head ditch during the experiment and by other changes in irrigation management on the commercial farm. For soil erosion the check furrows lost 322 lb/ac of sediment off of the field in the runoff water during a single irrigation. Furrows irrigated with granular polyacrylamide lost 7 lb/ac of sediment off of the field, while those irrigated with the liquid solution of polyacrylamide lost 104 lb/ac. Remember though, the granular polyacrylamide was applied at a rate double the liquid.
In increasing water infiltration, the check furrows lost 37.5 percent of the water as runoff and 62.5 percent was infiltrated. Out of the total water applied treated with granular polyacrylamide, 26.5 percent was lost as runoff and 73.3 percent of the water infiltrated into the soil. Out of the total water treated with liquid polyacrylamide, 29.1 percent was lost as runoff and 70.8 percent of the water infiltrated. Granular polyacrylamide used as a "patch" was effective to control the loss of sediment and increase water infiltration.

7. Is polyacrylamide stable?
In dilute aqueous solution, such as in the use of Enhanced Oil Recovery applications, polyacrylamide polymers are susceptible to chemical, thermal, and mechanical degradation. When the labile amine moiety hydrolyzes at elevated temperature or pH, chemical degradation occurs, It will resulting in the evolution of ammonia and a remaining carboxyl group. Thus, the degree of anionicity of the molecule increases. Thermal degradation of the vinyl backbone can occur through several possible radical mechanisms, including the autooxidation of small amounts of iron and reactions between oxygen and residual impurities from polymerization at elevated temperature. Mechanical degradation can also be an issue at the high shear rates experienced in the near-wellbore region. However, cross-linked variants of polyacrylamide have shown greater resistance to all of these methods of degradation, and have proved much more stable.

8. What is the potential health effects of polyacrylamide?
Eye: May cause eye irritation.
Skin: May cause skin irritation.
Ingestion: May cause irritation of the digestive tract.
Inhalation: May cause respiratory tract irritation.
Chronic: Chronic inhalation and ingestion may cause effects similar to those of acute inhalation and ingestion.

9. What is the safety and health issues of polyacrylamide?
Public perception is often that all chemicals are harmful. We have to tell you that they are not – chemicals are the building blocks of all living things. Acrylamide is a naturally occurring chemical found in a wide variety of foods, such as potatoes. There is a current controversy being debated in the media about “free acrylamide” causing cancer. It is being alleged that when the acrylamide component of a food product is heated to an extremely high temperature in the cooking process, its chemical structure is altered and in some instances acts as a cancer causing agent.
Actually, polyacrylamide, while using acrylamide as one of the raw materials in the chemical formula, is a man-made potassium-based chemical compound, it is found environmentally safe for use in drinking water treatment and agricultural production. It is not used in applications where extremely high heat is recommended. It’s like comparing apples and oranges - there is no correlation between the two.
Anionic polyacrylamide is a “non-toxic” chemical compound. Quality anionic polymers are:
1. Environmentally benign (safe)
2. Harmless to fish and aquatic organisms, wildlife, and plants
3. Non-combustible
4. Biodegradable

10. What is the first aid measures of polyacrylamide?
Eyes: Flush eyes with plenty of water for at least 15 minutes, occasionally lifting the upper and lower eyelids. Get medical aid.
Skin: Flush skin with plenty of water for at least 15 minutes while removing contaminated clothing and shoes.
Ingestion: If victim is conscious and alert, give 2-4 cupfuls of milk or water. Never give anything by mouth to an unconscious person. Get medical aid immediately.
Inhalation: Remove from exposure and move to fresh air immediately. If not breathing, give artificial respiration. If breathing is difficult, give oxygen. Get medical aid if cough or other symptoms appear.
Notes to Physician: Treat symptomatically.

11. What is the environmental effects of polyacrylamide?
It is known that polyacrylamide used in agriculture may contaminate food with the nerve toxin acrylamide. While polyacrylamide itself is relatively non-toxic, it is known that commercially available polyacrylamide contains minute residual amounts of acrylamide remaining from its production, usually less than 0.05% w/w. However, unpolymerized acrylamide, which is a neurotoxin, can be present in very small amounts in the polymerized acrylamide, therefore it is recommended to handle it with caution.
Additionally, there are concerns that polyacrylamide may de-polymerise to form acrylamide. In one (much debated) study conducted in 1997 at Kansas State University, the effect of environmental conditions on polyacrylamide were tested, and it was shown that degradation of polyacrylamide under certain conditions does in fact cause the release of acrylamide.

12. What is the handling and storage methods of polyacrylamide?
Handling: Wash thoroughly after handling. Use with adequate ventilation. Avoid contact with eyes, skin, and clothing. Avoid ingestion and inhalation. Store protected from light.
Storage: Store in a cool, dry, well-ventilated area away from incompatible substances.

13. What is the exposure controls and personal protection of polyacrylamide?
Exposure Controls
Engineering Controls: Use adequate ventilation to keep airborne concentrations low.
OSHA Vacated PELs: Acrylamide: 0.03 mg/m3 TWA Sodium Azide: No OSHA Vacated PELs are listed for this chemical. Water: No OSHA Vacated PELs are listed for this chemical.
Personal Protective Equipment
Eyes: Wear appropriate protective eyeglasses or chemical safety goggles as described by OSHA's eye and face protection regulations in 29 CFR 1910.133 or European Standard EN166.
Skin: Wear appropriate protective gloves to prevent skin exposure.
Clothing: Wear appropriate protective clothing to prevent skin exposure.

Respirators: Follow the OSHA respirator regulations found in 29 CFR 1910.134 or European Standard EN 149. Use a NIOSH/MSHA or European Standard EN 149 approved respirator if exposure limits are exceeded or if irritation or other symptoms are experienced.


Glycosaminoglycans

Fuming Zhang , . Robert J. Linhardt , in Handbook of Glycomics , 2010

Molecular Weight Analysis of GAGs

Polyacrylamide gel electrophoresis (PAGE) analysis can be conveniently applied to analyze the molecular weight of sulfated GAGs. Gels on which GAGs have been fractionated can be visualized with Alcian Blue with or without silver staining and the bands can be scanned and digitized. The average MW of a GAG is then calculated based on a mixture of HP-derived oligosaccharide standards prepared through the partial enzymatic depolymerization of HP. PAGE analysis of HS purified from human placenta is shown in Figure 3.2 [30] . The polydispersity of GAGs is observed as a broad smear in PAGE and a numerical value for the dispersity can be calculated.

Figure 3.2 . (a) Gradient polyacrylamide gel electrophoresis (PAGE) analysis with Alcian Blue staining of glycosaminoglycan samples before and after treatment with heparin lyases. Lane 1 is intact porcine intestinal heparan sulfate (HS) lane 2 is porcine intestinal HS after treatment with heparin lyase 1, 2, and 3 lane 3 is a hexasulfated tetrasaccharide standard derived from heparin (HP) indicated by the arrow lane 4 is a mixture of HP-derived oligosaccharide standards enzymatically prepared from bovine lung HP—the numbers indicate their degree of polymerization (i.e., 4 is a tetrasaccharide) lane 5 is intact porcine HP lane 6 is porcine intestinal HP after heparin lyase 1 treatment lane 7 is human liver HS lane 8 is human liver HS after heparin lyase 3 treatment. (b) A plot of log molecular weight of bovine lung HP-derived oligosaccharide standards as a function of migration distance of each oligosaccharide from which the average molecular weight of HP and HS can be calculated. (See color plate 6.)

Gel permeation chromatography (GPC), which separates molecule solely on the basis of differences in molecular size has been used for the MW analysis of GAGs. Dextrans, dextran sulfates, or GAGs of different MWs can be used as standards in a GPC column to calibrate the MW of GAGs. Refractive index detection is typically used in this method [52] .


SDS-PAGE protocol

Protein samples are denatured by heating them with a detergent SDS and mercaptoethanol. The former binds strongly to the proteins and gives them a high negative charge whilst the latter frees sulfhydryl groups, thus yielding polypeptide chains carrying an excess negative charge and similar charge to mass ratio. This helps the resolution of proteins strictly based on their size during gel electrophoresis.

The electrophoretic gel usually has several components including acrylamide, BIS, and a buffer. The mixture is degassed to prevent bubble formation during polymerization of the gel. Ammonium persulfate, a free radical source, and a stabilizer are added to start polymerization. BIS is also added to form cross-links between acrylamide molecules until a gel is ultimately formed.

As an electric current is applied proteins migrate through the gel to the positive electrode as they have a negative charge. Each molecule moves at a different rate based on its molecular weight – small molecules move more rapidly through the gel than larger ones. Migration is usually faster at higher voltages. After a few hours, the protein molecules are all separated by size.

4. Staining and visualization:

Once electrophoresis is complete, the gel can be stained using colored dyes such as Coomassie Brilliant Blue or ethidium bromide to make the separated proteins appear as distinct colored bands on the gel.

Unbound dye is washed out from the gel. The stained gels are then dried so the color intensity of the protein bands can be measured. Bands of radioactive proteins can be detected by autoradiography. The proteins can also be quantified as the protein content is directly proportional to the quantity of the bound dye.

Some gel systems introduce a tracking dye such as bromophenol blue along with the protein sample &ndash the visible distance travelled by the dye on the gel helps in deciding the required duration of electrophoresis. Bromophenol blue travels along with the sample molecules until it eventually reaches the bottom of the gel. Electrophoresis needs to stop at this point to ensure no protein molecules electrophorese out of the gel and into the buffer.


Similarities Between Agarose and Polyacrylamide

  • Agarose and polyacrylamide are the two main types of gels that are used for the separation of biomolecules, such as DNA, RNA, and proteins.
  • Therefore, they are important in molecular biology and biochemistry.
  • These biomolecules move between electrodes after applying the electric field, moving the charged molecules through the matrix.
  • On that account, both types of gels allow the separation of biomolecules based on their size and charge.

Use of Natural and Synthetic Oligosaccharide, Neoglycolipid and Glycolipid Libraries in Defining Lectins from Pathogens

Krista Weikkolainen , . Jari Natunen , in Lectins , 2007

3.3.5 Polyacrylamide conjugates, biotin glycans and glycan arrays

Polyacrylamide (PAA) conjugates developed by Bovin and colleagues [66b] are among the most useful reagents for various assays in glycobiology. The materials commercialized through Syntesome and later through Lectinity Holdings include a vast variety of natural glycans and their synthetic analogs. The reagents are available as monovalent biotinylated, as polyvalent PAA-conjugates and even as biotinylated polyvalent PAA-conjugates. The PAA conjugates have been used, e.g., for studies of influenza virus [67] .

A non-commercial US-based Consortium for Functional Glycomics ( http://glycomics.scripps.edu/CFGad.html ) has also created impressive library of oligosaccharides and glycoconjugates by chemo–enzymatic synthesis. These include great variety of oligosaccharides with azidoethyl spacer, O-glycan-Thr-conjugates, biotin conjugates and PAA conjugates ( www.functionalglycomics.org/static/consortium/resources/resourcecored.shtml ). The materials are also used as a printed glycan microarray with 285 glycan targets. The array concept was recently demonstrated with pandemic influenza viruses [17] . Another pioneering study demonstrated carbohydrate microarrays for studies of bacterial adhesion [20] .


Buffers, pH, Acids, and Bases

Figure 5. The pH scale measures the amount of hydrogen ions (H+) in a substance. (credit: modification of work by Edward Stevens)

The pH of a solution is a measure of its acidity or alkalinity. You have probably used litmus paper, paper that has been treated with a natural water-soluble dye so it can be used as a pH indicator, to test how much acid or base (alkalinity) exists in a solution. You might have even used some to make sure the water in an outdoor swimming pool is properly treated. In both cases, this pH test measures the amount of hydrogen ions that exists in a given solution. High concentrations of hydrogen ions yield a low pH, whereas low levels of hydrogen ions result in a high pH. The overall concentration of hydrogen ions is inversely related to its pH and can be measured on the pH scale (Figure 5). Therefore, the more hydrogen ions present, the lower the pH conversely, the fewer hydrogen ions, the higher the pH.

The pH scale ranges from 0 to 14. A change of one unit on the pH scale represents a change in the concentration of hydrogen ions by a factor of 10, a change in two units represents a change in the concentration of hydrogen ions by a factor of 100. Thus, small changes in pH represent large changes in the concentrations of hydrogen ions. Pure water is neutral. It is neither acidic nor basic, and has a pH of 7.0. Anything below 7.0 (ranging from 0.0 to 6.9) is acidic, and anything above 7.0 (from 7.1 to 14.0) is alkaline. The blood in your veins is slightly alkaline (pH = 7.4). The environment in your stomach is highly acidic (pH = 1 to 2). Orange juice is mildly acidic (pH = approximately 3.5), whereas baking soda is basic (pH = 9.0).

Acids are substances that provide hydrogen ions (H + ) and lower pH, whereas bases provide hydroxide ions (OH – ) and raise pH. The stronger the acid, the more readily it donates H + . For example, hydrochloric acid and lemon juice are very acidic and readily give up H + when added to water. Conversely, bases are those substances that readily donate OH – . The OH – ions combine with H + to produce water, which raises a substance’s pH. Sodium hydroxide and many household cleaners are very alkaline and give up OH – rapidly when placed in water, thereby raising the pH.

Most cells in our bodies operate within a very narrow window of the pH scale, typically ranging only from 7.2 to 7.6. If the pH of the body is outside of this range, the respiratory system malfunctions, as do other organs in the body. Cells no longer function properly, and proteins will break down. Deviation outside of the pH range can induce coma or even cause death.

So how is it that we can ingest or inhale acidic or basic substances and not die? Buffers are the key. Buffers readily absorb excess H + or OH – , keeping the pH of the body carefully maintained in the aforementioned narrow range. Carbon dioxide is part of a prominent buffer system in the human body it keeps the pH within the proper range. This buffer system involves carbonic acid (H2CO3) and bicarbonate (HCO3 – ) anion. If too much H + enters the body, bicarbonate will combine with the H + to create carbonic acid and limit the decrease in pH. Likewise, if too much OH – is introduced into the system, carbonic acid will combine with it to create bicarbonate and limit the increase in pH. While carbonic acid is an important product in this reaction, its presence is fleeting because the carbonic acid is released from the body as carbon dioxide gas each time we breathe. Without this buffer system, the pH in our bodies would fluctuate too much and we would fail to survive.


Introduction

Numerous studies have demonstrated that azo dyes are a series of synthetic organic compounds that are widely used and applied in different industrial sectors like textile dyeing, paper making, printing and cell biology (Osma et al. 2007 Gupta and Suhas 2009 Konicki et al. 2013). During dye production and textile manufacturing process, a large quantity of wastewater containing dye stuffs with intensive color and toxicity are introduced into the aquatic systems (Ibhadon et al. 2008). The discharge of industrial dye effluent into the water systems and environment has become a serious pollution problem and human health risk because of its high visibility and toxicity (Gupta and Suhas 2009 Konicki et al. 2013 Zhang et al. 2014). Due to increasingly drastic restrictions on the organic content of industrial effluents, it is essential to remove dyes from wastewater before it is discharged to the normal water bodies (Ahmad 2009). Many of these dyes are also toxic specifically, the crystal violet (CV) is an organic cationic dye and widely used as textile colorant and biological stain. Several studies indicated that accumulation of CV has been suspected to cause harmful effects such as cancer on human beings (Zhang et al. 2014). Therefore, considerable efforts have been devoted for the treatment of dye effluents using different techniques such as coagulation, flocculation, reverse-osmosis, photo-degradation processes and ion-exchange. Among the various techniques, adsorption process is the most preferred method to remove the dyes from aqueous solution due to its simplicity, efficiency, convenience, ease of operation and inexpensive nature (Martins et al. 2017).

Activated carbon as an adsorbent has been widely used for the removal of dyes, heavy metals and other organic pollutants from aqueous solution, but its high cost limits its commercial application. In recent years, extensive research has been undertaken to develop alternative and low-cost adsorbents (Ahmad 2009). Adsorbents utilized for dye removal should be characterized by an excellent adsorption capacity derived fundamentally from the combination of pore structure and surface chemistry (Pang et al. 2019). Recent literature survey revealed that various adsorbents have been employed in the treatment of dye-polluted waters includes clay minerals (Kausar et al. 2018), zeolites (Sivalingam and Sen 2019), polymer composite (Jayasantha Kumari et al. 2017), fly ash (Gao et al. 2015) and reduced the cost of adsorption process various biomasses have been suggested as dye adsorbents: rice husks (Franco et al. 2015), grape wastes (Vanni et al. 2017), ouricuri fibers (Meili et al. 2017) and para chestnut husk (Georgin et al. 2018).

Recently, intense research is going on to search for new conventional and low-cost green adsorbent for waste water treatment. Moreover, to further increase mechanical strength and sorption capacity of the conventional adsorbent, the lignocellulosic part was modified by various organic and inorganic compounds, acid and bases and also blended with several synthetic polymers reported elsewhere (Wang et al. 2015), biopolymer (Zhang et al. 2013), metal oxide nanoparticles (Gopalakannan and Viswanathan 2015), nano-clay materials (Hassani et al. 2015) and poly (methyl metha acrylate)-grafted alginate @Cys-bentonite copolymer hybrid nanocomposite (Hasan and Ahmad 2019). Moreover, phosphate-treated sawdust (lignocellulosic material) shows a remarkable increase in sorption capacity of Cr(VI) as compared to untreated sawdust (Ajmal et al. 1996). Therefore, to further enhance the capacity of the adsorbent we introduced a suitable procedure for surface modification which involved the grafting of polyacrylamide onto Actinidia deliciosa peels powder using N,N’-methylenebisacrylamide as a cross-linking agent and subsequent functionalization of the polymer network with desired reagent for treating dyes wastewater (Unnithan and Anirudhan 2001). After modification, it was found that the adsorption capacity and stability of the adsorbent materials have increased, which is an important aspect of commercial development of biosorbent materials. In this quest, author has attempted to explore a novel low-cost and abundantly available material Actinidia deliciosa peels powder (ADP) to remove hazardous crystal violet dye (CV) from aqueous solution and waste water.

Herein, we report the preparation of a novel and eco-friendly adsorbent, polyacrylamide-grafted Actinidia deliciosa peels powder (PGADP) and its application for the sequestration of crystal violet (CV) dye from aqueous solution. The experiments were carried out to see the effects of various parameters such as pH, contact time, initial dye concentration and temperature on the adsorption of dye onto (PGADP). The obtained experimental adsorption data were applied to validate the Langmuir, Freundlich, Temkin and Dubinin–Radushkevich (D-R) isotherm models. The equilibrium data were further tested by using pseudo-first, pseudo-second order, intra-particle diffusion and elovich kinetic models. In order to make the process more economical and feasible, the exhausted material was further desorbed and regenerated. To see the morphology of the prepared adsorbent, PGADP was characterized by FTIR, XRD, SEM, EDX, TEM and TGA techniques.

The main novelty of present material PGADP is its high monolayer adsorption capacity of (75.19 mgg −1 ) as compared to other adsorbents in the literature (Table 5). The PGADP also exhibits very good regenerative capability, and it can be used up to fourth cycle successfully without much loss in efficiency for the removal of CV dye. Therefore, PGADP can be used very effectively and economically for the removal of CV dye from the developing countries.


Types of charged lipids

Fatty acids

The simplest of the charged lipids, fatty acids are a large group of amphipathic molecules consisting of short, medium or long-chain hydrocarbon &ldquotails&rdquo (C4 to C36) and a polar carboxylic acid &ldquohead&rdquo. The aliphatic chains can be fully saturated or unsaturated to some extent, and provide the hydrophobic character of the fatty acid. Regardless of the length of the aliphatic tail group the hydrophilic carboxylic acid group will have a pKa around 4.0, which means that under physiological conditions, the majority of fatty acids will have a negative charge of -1. The tails can also contain carbon ring structures, hydroxyl groups, and additional methyl group branches (Figure (PageIndex<1>)).

Figure (PageIndex<1>): An example of fatty acids with differing chain length, saturation, and linearity. Image used with permission (CC BY-SA 3.0 Lojban via Wikipedia).

The physical properties of fatty acids are largely dependent on the length and degree of unsaturation of the hydrocarbon tail group. The major factor influencing properties such as melting point and water solubility is the ordering of water molecules around the hydrophobic tails. Fatty acids will cluster together as result of the hydrophobic effect. The clustering of the aliphatic tail groups forms a crystalline lattice, minimizing the exposed hydrophobic surface area and the formation of a hydration shells around individual fatty acid tails. With increasing degrees of unsaturation in the tails (increased number of double bonds), the effect is reversed. Unsaturation introduces kinks into the tails so that they no longer pack as uniformly and tightly, which allows water molecules to become ordered around the individual tails. As such, the crystalline lattice is weakened, increasing the solubility of the fatty acid and decreasing its melting point.

Figure (PageIndex<2>): Image depicting the various structures formed by charged lipids. Image source: Wikipedia

Fatty acids have a variety of important roles. The hydrocarbon tails are a major source of energy when each successive two-carbon unit is converted into acetyl-CoA during beta-oxidation, the process of which generates NADH and FADH2 which are subsequently used in the formation of ATP in oxidative phosphorylation. Fatty acids and their derivatives also function as soaps and detergents. At high enough concentrations, fatty acids will aggregate to form spherical structures called micelles, which are energetically favorable structures that increase the entropy of water by minimizing water clathrate formation around the hydrophobic regions. They accomplish this by clustering the hydrophobic tails into the center of the sphere, while the hydrophilic heads form a shell that is water soluble. The spherical structure is favored by fatty acids because the cross-sectional area of the head group is larger than that of the tail, forming a conical shape. When soaps (the salts of fatty acids) are mixed with water, the micelles that are formed sequester oils and hydrophobic &ldquodirt&rdquo into the micelle center, while the hydrophilic outer shell helps to wash the dirt-carrying micelles away. Lastly, fatty acids form the foundation of di- and triacylglycerols, a family of molecules with two or three fatty acid chains bound by glycerol. When the third hydroxyl group of the glycerol is bound to a charged molecule such as phosphate instead of a hydrocarbon chain, the diacylglycerol is known as a phospholipid.

Phospholipids

As with any molecule, the formation of a charge requires the inclusion of an ionizable element, most often covalently bound to the parent compound. In the case of diacylglycerols, non-ionizable hydrocarbons require the covalent addition of an ionic group to the glycerol to impart a charge. By far the most common of these is the negatively charged phosphate group ((PO_4^<3->)) which, when covalently bound to the glycerol moiety of a two-chain fatty acid, forms the main group of charged fatty acids known as phospholipids. Phospholipids are amphipathic molecules composed of three sections: a diglycerol, a hydrophilic &ldquohead&rdquo consisting of the charged phosphate moiety, and a small organic molecule covalently bound to the phosphate (Figure (PageIndex<3>)).

Figure (PageIndex<3>): Structure of a phospholipid, containing the fatty acid tails, glycerol, phosphate moiety, and an additional head-group structure. Image source: Wikipedia

Phospholipids comprise the major components of lipid bilayers in cellular membranes. Similar to fatty acids, phospholipids will favor the clustering of the hydrophobic tails to exclude water as much as possible. Unlike fatty acids, however, the presence of two tails favors the formation of a more planar membrane because the cross-sectional area of the head and tail groups is similar, forming a cylindrical shape which is most efficiently packed as a bilayer. In the bilayer, the hydrophobic tails interact with one another, whereas the hydrophilic head groups interact with the aqueous environment on either side of the bilayer. While the bilayer is most practical as a two dimensional planar structure, it can fold in on itself to form a closed, vesicle called a liposome. Unlike the micelle, the liposome has a bilayer wall enclosing a hollow, aqueous cavity, and is used organisms as a transport mechanism for any number of substances.

The structural complexity of the membrane bilayer goes beyond a single type of phospholipid molecule. There exist a variety of phospholipids with varying head groups and hydrocarbon tails. Additionally, membrane lipids are not symmetrically distributed throughout the membrane bilayer, whereby certain phospholipids are more likely to be found in the inner monolayer with the others more likely to be found in the outer leaflet. On the other hand, the physical properties of the different head groups dictate the interactions of each phospholipid with the aqueous environment, and resulting in the movement of the lipids across the bilayer in both the vertical and horizontal directions, continuously changing its shape and phospholipid organization. To give a better idea of the different phospholipid head groups, the following sections will describe the structural features some of the more common ones.


Definition of Simple Staining

Simple staining is defined as one of the ordinaries yet the popular method used to elucidate the bacterial size, shape and arrangement to differentiate the various bacteria groups. It stains the bacterial cell uniformly and thus increases the visibility of an organism. A term simple staining sometimes interchangeable with the terms like direct, positive or monochrome staining. Now let us understand why simple staining is called by such alternative names.

  • Direct staining: Because it is a direct method that directly stains the bacterial cell with a colourless background.
  • Positive staining: Because it uses positively charged basic dyes that bind with the negatively charged bacterial cell.
  • Monochrome staining: Because it adds contrast to the specimen by using a single stain only.

Simple Stains

Simple stains can be defined as the basic dyes, which are the alcoholic or aqueous solutions (diluted up to 1-2%). These can easily release OH – and accepts H + ion, due to which the simple stains are positively charged. As the simple stains are positively charged, they usually termed as positive or cationic dyes.

It is commonly used to colour most of the bacteria. As the simple stain carry a positive charge, it firmly adheres to a negative bacterial cell and makes the organism coloured by leaving a background colourless. Examples of simple stain include safranin, methylene blue, crystal violet etc.

The basic stains have different exposure time to penetrate and stain the bacterial cell.

Basic stainsExposure time to stain the bacteria
Methylene blue1-2 minutes
Crystal violet20-60 seconds
Carbol fuschin15-30 seconds
Safranin30-60 seconds

Principle

Its principle is based on producing a marked contrast between the organism and its surroundings by using basic stain. A basic dye consists of a positive chromophore, which strongly attracts to the negative cell components and charged molecules like nucleic acids and proteins. Thus, a simple staining technique results in a coloured bacterial cell against a colourless background.

Procedure of Simple Staining

It involves the following three steps:


Smear Preparation

Bacterial smear appears as a thin film of bacterial culture. For the smear preparation, we need to perform the following steps:

  1. Take a clean, grease-free glass slide.
  2. Add a drop of distilled water at the centre of the glass slide.
  3. Then, add inoculum from the bacterial culture with sterilized inoculating loop on the glass slide.
  4. After that, mix the inoculum with a drop of distilled water to make a thin film by uniformly rotating the inoculating loop until the formation of a thin bacterial film.


Heat Fixing

After smear preparation, move the prepared slides over Bunsen burner’s flame for at least three times. Then, allow the slide to air dry. There are many reasons to perform heat fixing, and it can not be skipped because:

  • Heat fixing helps in the fixation of a specimen to the glass slide.
  • Heat fixing helps the stain to penetrate the smear.


Staining of Bacteria

It is the last and the most crucial step, in which one can identify the morphological characteristics of the bacteria through microscopic examination, once the cells get stained. This stage involves the following steps, which are as follows:

  1. Add stain to the heat fixed smear.
  2. Allow the stain to stand for at least 1 minute so that it can penetrate between the cells.
  3. Wash off the glass slide carefully.
  4. Blot dry the slide with absorbent paper (do not wipe the slide).
  5. Examine the glass slide under the microscope from low to high power to get a magnified view of the specimen. One can also add a drop of oil immersion over the glass slide’s stained area to observe it under 100X objective.


Video

Advantages

  • Simple staining is a very simple method to perform, which stains the organism by using a single reagent.
  • It is a rapid method which reduces the performance time by taking only 3-5 minutes.
  • Simple staining helps to examine or elucidate the bacterial shape, size and arrangement.
  • It also helps us to differentiate the bacterial cells from the non-living structures.
  • Simple staining can be useful in the preliminary study of the bacteria’s morphological characteristics.

Disadvantages

  • It does not give much information about the cell apart from the bacteria’s morphological characteristics.
  • Through simple staining, we cannot classify a particular type of organism.

Conclusion

Therefore, we can conclude that a simple staining method is the easiest way to colour the microscopic object as it uses a single basic stain. The results of simple staining are based on the type of basic stain that has been used.

The colour of a stain will decide the colour of a specimen that has to be identified. For example, when the bacteria retain the safranin colour, they appear pink-red, and the same goes with the other stains.

There is an attraction between the positive stain and the negative bacterial cell in simple staining, which results in the observation of coloured bacteria with a bright background.



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