How does silver kill microbes?

[Gruff]

There's a new self-cleaning nanotechnology-based sock that cures smelly feet and athlete's foot (see www.agactive.co.uk). The socks contain nano-particles of silver. Does anyone know exactly how silver kills the bacteria/fungus/virus and why haven't they evolved against whatever the mechanism is?

 

Thanks!

[gcol]

Water filter manufacturers always talk up its effectiveness in conjunction with activated carbon, yet I have come across "independant" research that says its benefit is less clearcut, and some pathogens can become resistant.

 

Here is one quote "In its ionic colloid state, silver is recognized as a germicide, or in some cases as a bacteriostatic. It is believed that silver is able to disable the particular enzyme that pathogenic bacteria and fungi use for oxygen metabolism, thus suffocating them. Other pathogens are destroyed by the electric charge on the silver particles, causing their internal protoplast to collapse, and still others are rendered unable to reproduce. Parasites are also killed while in their egg stage. "

 

Silver contamination of potable water can be a health hazard, however.

 

The literature seems confusing, there appears to be no definitive answer.

 

Anyone else got an opinion?

[Martin]

There's a new self-cleaning nanotechnology-based sock that cures smelly feet and athlete's foot...!

 

That's funny!

 

maybe gcol has the answer, I can't add anything scientific to the discussion but I'll mention this:

 

I know for a fact that already 50 years ago some people thought that silver was self-sterilizing.

Wounds were sometimes sutured with silver wire (for thread) in hopes that they would not get infected! Before the widespread use of antibiotics it was a big problem that stitched-up wounds would get infected.

 

I also know of an apple-farmer near where we lived who used to make CIDER and he wouldn't PASTEURIZE his cider to keep it from spoiling. He would just filter the solid crud out of the stuff and then he would pump the cider thru a long SILVER COIL and he believed that this got rid of the germs or fungus that would otherwise make the cider spoil.

 

AFAIK it may be that silver is not effective, although people thought it was. It could be a "rural legend".

 

But I guess there could also be some truth in it----judging from what gcol said.

 

IMO silver is rare enough (in substantial concentrations) that germs would likely not evolve a tolerance of it. If they found it toxic or obnoxious they would just go live somewhere else where there wasnt any silver

[zyncod]

Well, I know that silver nitrate is an antimicrobial agent, because we used to use it to sterilize things that we couldn't heat in my old lab, where we worked on algae. Ethanol was another option - but this procedure was such a pain in the ass anyway that waiting around for the ethanol to sterilize the instruments would have made it horrible (basically, the procedure when you have a contaminated algal culture is to pick out single algae cells using a stretched-out pipette and drag them through the agar to knock off any bacteria stuck to them - and you can't heat the pipettes because even a match would destroy the stretched-out tip that it took you 30 minutes to make the right way). The silver ions likely covalently bond with various proteins/lipids/cell wall components on the bacteria. For whatever reason, it's less toxic for eukaryotes.

 

Anyway... I think in clinical situations, silver is mainly used as a salt with sulfa drugs. The sulfa drugs are the main active agents - the silver is just there as a little bonus. But, because the silver is still toxic to eukaryotic cells, silver/sulfa drugs are not used systemically.

[michael]

Some Alternative Medicine Practitioners use Colloidal Silver as an anti-bacterial agent.

See

http://www.silvermedicine.org/byustudy.html

or give it a google

[Mokele]

Some Alternative Medicine Practitioners use Colloidal Silver as an anti-bacterial agent.

 

The correct term is "quack". Want to see why? Google image search "argyria".

 

Yes, that's what happens if you use this colloidal silver BS: you turn BLUE!

 

This is why one should trust *real* doctors instead.

 

Mokele

[michael]

Thankyou.

I always thought it was to good to be true.

 

But don't throw out the baby with the bathwater.

Both 'alternative' (complimnetary?) and "real" doctors have strengths and weaknesses.

 

This is how real doctors use/used it

 

In the Middle Ages silver nitrate was used for the treatment of nervous system disorders such as epilepsy and tabes dorsalis.

After observing Dr. Halstead of Johns Hopkins University apply silver foil and gauze to wounds to prevent infection in 1897, Crede popularized the use of silver as an anti-infective measure [1].

In the pre-antibiotic era, silver was used in nose drops (Argyrol, a silver proteinate), in sinusitis and common-cold remedies, and for the treatment of syphilis (Neo-Silvol, a silver arsphenamine).

More recently silver arsphenamine has been used in topical astringent preparations.

Silver sulfadiazine (1 %) was formulated in 1967 by substituting a portion of the sulfonamide with an atom of silver; it is the most frequently used topical agent for burn treatment [2].

. . .

Inorganic silver compounds are germicidal and hence have been used extensively in the field of medicine.

These compounds denature proteins by binding to the reactive groups of proteins resulting in their precipitation.

They inactivate enzymes by reacting with the sulfhydryl groups to form hemisilver sulfides.

They also react with the amino-, carboxyl-, phosphate-, and imidazole-groups and diminish the activities of lactate dehydrogenase and glutathione peroxidase [5].

. . .

Argyria is a rare condition associated with chronic exposure to silver-containing products;

. . .

The official drug guidebooks (United States Pharmacopeia and National Formulary) have not listed colloidal silver products since 1975

http://dermatology.cdlib.org/111/case_reports/argyria/wadhera.html

[FixUrSelf]

I am Hep C positive. This was found out in 1995 when I went in for an ulcer operation. It was determined I most likely contracted the virus during a blood transfusion in 1985. I've been monitoring my viral load since 1995 and it has slowly increased until it reached 825,000 a couple months back. I was feeling terrible, tired, had trouble focusing for sustained periods, just a feeling of discomfort I find difficult to describe. This led to fear that I might end up a casualty of this disease. I couldn't afford any of the available treatments, nor was I a candidate of any assistance or drug sponsorship and so began looking for alternatives I might afford.

A month back I began a regime of three teaspoons a day of colloidal silver, one teaspoon every 8 hours mixed in 6oz of water. The colloidal silver was 30ppm. I chose a specific brand because of the amber glass bottle which I read inhibited degradation of the colloidal silver solution over plastic containers, and of course price, 39.95 for 32oz of 30ppm which I found the least expensive available. A few days back I went to the sliding scale clinic I've been taking advantage of since I lost my job (my illness had me so exhausted I couldn't stay awake at work). My current viral load is under 100 which I am told may be an indication I'm cured.

You can imagine how excited I am. My viral load will be monitored again next month. The doctor is amazed as am I. I shared with him what I did and although pleased with the results he said even if next months results showed the low viral level remains consistent, he would be apprehensive to suggest it to other patients because he could jeopardize his license. But he asked if I might consider coming in at various times to speak with patients about my results. Of course I will.

My interest in this forum is trying to help share my experience with colloidal silver. My energy level is up and I feel great. I didn't pay exorbitantly for the product I used, and in fact found the least expensive one made by Garden of Eden to be effective. I can't believe eBay succumbed to the pressures of Big Pharma and the FDA and removed all domestic colloidal silver products from their site. I am disgusted to learn about how we the people are being kept in the dark about potential cures, and how people sharing knowledge are being targeted, and how there are sites demonizing cures simply because those cures can't be profited from by the current established medical industry.

It's frustrating to me, to see what I believe is the protectionism enforced by the FDA and other organizations supposedly looking out for the people, and that they are instead puppets of Big Pharma. But how much more frustrating is it for those who continue to suffer, and who are denied options because the health of the people is second to the economic health of an industry.

I'd welcome member feedback on this issue. I know there are many opinions and aspects on this subject which must be considered. If you've any to share I'd like to read it. After all, the more informed we are the better our chances for survival individually and as a society. It reminds me of an episode of Star Wars were Queen Amidala makes a statement, "It is clear to me now that the Republic no longer functions. I pray you will bring sanity and compassion back to the Senate."

In this case it is our medical system, FDA, AMA, etc., which no longer functions because they have been taken over by Big Pharma. It is only by our participation (not exclusion), through which we the people may be able to bring compassion back to medicine. It is sites like this which offer us the opportunity and a chance. Wishing peace and compassion to us all.

[John Cuthber]

"

Acute infection

Hepatitis C infection causes acute symptoms in 15% of cases.^[5] Symptoms are generally mild and vague, including a decreased appetite, fatigue, nausea, muscle or joint pains, and weight loss.^[6] Most cases of acute infection are not associated with jaundice.^[7] The infection resolves spontaneously in 10-50% of cases, which occurs more frequently in individuals who are young and female"

 

from

http://en.wikipedia.org/wiki/Hepatitis_C

 

Assuming that the disappearance of the infection is due to the silver is a logical fallacy.

http://en.wikipedia.org/wiki/Post_hoc_ergo_propter_hoc

 

However it is known that the colloidal silver is toxic.

 

The mechanisms by which silver is microbicidal are not well targeted towards viruses.

[FixUrSelf]

I'm neither young nor female (male 60 years old). So if somehow it's a coincidence that after 27 years with the virus it disappears the same month I use colloidal silver, then happy coincidence.

Back in 1991 I went to a doctor because of an ulcer. I mentioned I read an article in Time magazine about helicobacter pylori being found in 80% of ulcer patients and that when treated with antibiotics (I think I remember amoxicillin, flagil and salts of bismuth as the treatment regime), the ulcers were healed. My doctor was 'old school' and told me I should leave the practice of medicine up to him. He gave me a prescription for Ranitidine and sent me on my way. That same week I went to San Francisco General and spoke with a couple interns about the article. They seemed excited about the conversation put me on the antibiotics and my ulcer disappeared. About 6 months afterwards I read in the Chronicle that SF General had begun a study on the effects of an antibiotic regime in the treatment of ulcers.

I don't believe ignorance is bliss. But sometimes what we think we know can blind us just as much as what we don't know. Imagine how many experts told the Wright brothers they would never be able to fly, or told Columbus he would eventually sail off flat earth, or how many doctors on television commercials touted the benefits of smoking which reached millions of future cancer patients. There is so much information out there to choose from to substantiate which ever side of the fence you're on in reference to colloidal silver. The benefits of not being an expert is one doesn't need to understand something in order to accept the results.

By the way, I have personally spoken to a half dozen other Lyme and Hepatitis patients who describe similar coincidences with their use of colloidal silver. Let's hope the coincidences keep piling up. Not for your sake, but for those of us who are slowly dying because of the experts, or those protecting an industry which cannot exist without the sick.

Edited January 22, 2013 by FixUrSelf

[StringJunky]

The oral consumption of colloidal silver is a very bad idea. From the NIH National Centre For Complementary And Alternative Medicine on colloidal silver. Bolded emphasis was not added by me:

 

 

Key Points

• Colloidal silver is not safe or effective for treating any disease or condition.
• Colloidal silver can cause serious side effects. The most common is argyria, a bluish-gray discoloration of the skin, which is usually not treatable or reversible.
• The FDA and the Federal Trade Commission have taken action against a number of companies (including some companies that sell products over the Internet) for making drug-like claims about colloidal silver products.
• Tell all your health care providers about any complementary health practices you use. Give them a full picture of what you do to manage your health. This will help ensure coordinated and safe care.

Background

Silver is a metallic element. People are exposed to silver, usually in tiny amounts, through air, water, and food, and in certain activities such as jewelry-making, soldering, or photography.

In 1999, the FDA prohibited the sale of over-the-counter drugs containing colloidal silver or silver salts because they had not been shown to be safe and effective. However, colloidal silver products are still being sold as dietary supplements or homeopathic remedies. Consumers should be aware that unlike some homeopathic remedies, which are so diluted that none of the original substance is present, some colloidal silver products marketed as homeopathic may not be extremely diluted.

A few prescription drugs containing silver are still in use; for example, silver sulfadiazine is used to treat burns. However, of the few prescription drugs containing silver, all are for topical use; there are no FDA-approved prescription or over-the-counter drugs containing silver that are taken orally.

Scientific Evidence

Reviews of the scientific literature on colloidal silver have concluded that:

• Silver has no known function in the body.
• Silver is not a nutritionally essential mineral or a cure-all and should not be promoted as such.
• Claims that there can be a “deficiency” of silver in the body and that such a deficiency can lead to disease are unfounded.
• Claims made about the effectiveness of colloidal silver for numerous diseases are unsupported scientifically.
• Colloidal silver can have serious side effects.

Side Effects and Risks

Silver builds up in the tissues of the body. This buildup of silver can lead to a side effect called argyria, a grayish or bluish discoloration of the skin, conjunctiva (the clear membrane that covers the white part of the eye), nails, and gums. In 2009, the FDA issued a warning to consumers about the risk of argyria associated with the use of dietary supplements containing silver, including colloidal silver.

Argyria is usually permanent and may discolor large portions of the body, especially those exposed to the sun. Attempts to reverse the discoloration have usually been unsuccessful, except in instances where only small areas of skin needed to be treated. Cases of argyria have occurred in people who drank homemade colloidal silver liquids as well as in people who used commercial colloidal silver products.

Although argyria is the most common adverse effect of consuming colloidal silver, some cases have been reported where colloidal silver may have caused kidney, liver, or nervous system problems. Colloidal silver may interfere with the body’s absorption of some drugs, such as certain antibiotics, thyroxine (used to treat thyroid disorders), and penicillamine (used to treat conditions such as rheumatoid arthritis and metal poisoning).

http://nccam.nih.gov/health/silver

Edited January 22, 2013 by StringJunky

[FixUrSelf]

I must admit that I find nothing on the Internet, Medline, or anywhere else concerning the efficacy of colloidal silver ingestion. Nothing. No clinical trials or case reports or even animal studies.

Although I did find an in-vitro study conducted by Mark A. Farinha, Ph.D., Professor of Microbiology at the University of North Texas demonstrating effectiveness of a 15 PPM and a 30 PPM isolated colloidal silver product against a wide range of illness-causing bacteria.

And another study from Microbiologists at Brigham Young University documenting the effectiveness of silver as an antimicrobial substance concluding that a quality product may possibly serve as an antibiotic alternative.

So outside of toxicities as far as the medical industrial complex is concerned there is no data of any kind to support either position. Therefore, why is there such a demonization of it's use. As a layperson I can only go back on my own personal experience, and the people I've met as a Hospice volunteer who are no longer in Hospice, and others who I've met in the course of my service who have shared their experiences. And so this layperson must fall back on results.

Edited January 22, 2013 by FixUrSelf

[StringJunky]

I must admit that I find nothing on the Internet, Medline, or anywhere else concerning the efficacy of colloidal silver ingestion. Nothing. No clinical trials or case reports or even animal studies.

 

Silver in Heathcare It's Antimicrobial Efficacy and Safety In Use

 

http://books.google.co.uk/books?id=QxtLm7MgQhYC&printsec=frontcover&source=gbs_ge_summary_r&cad=0#v=onepage&q&f=false

 

Got from the reference section of Medical Uses of Silver in Wikipedia. http://en.wikipedia.org/wiki/Medical_uses_of_silver

 

There's plenty to be getting on with there.

[Lance]

I once had a pair of socks that had silver wire weaved into it. My feet weren't any less smelly after extended wear. The overclocking community also uses silver coils in water cooling systems because they believe it stops the growth of algae. They call it a "kill coil". I guess when you come up with a name that cool you don't really need any scientific evidence.

[dirtyamerica]

copper ions work too

[CharonY]

Generally the silver ions are what kill off bacteria, thus the form of silver is relevant. Nanoparticles are, due to their larger surface, more effective, for instance.

[alpha2cen]

Silve is a good chemical adsorber to the oxygen.

So silver surface is filled with much amount of oxygen.

Micro organism, speciallay bacteria has an anaerobic property.

At the rich oxygen state, the DNA in the bacteria plasma is damaged by the oxygen.

During the evolution process, the nuclear membrane is created by that reason.

To protect DNA from the oxgen attack, DNA should have been enveloped by the nuclear membrane.

Edited January 23, 2013 by alpha2cen

[CharonY]

OK, this pretty much all wrong. The major discussed actions of silver particles involve the formation of oxidative stress and inhibitaiton of enzymes due to interaction with thiol groups.

[alpha2cen]

The oxygen on the silver particle is more reversible than other metal.

[John Cuthber]

The oxygen on the silver particle is more reversible than other metal.

Not really.

[alpha2cen]

Not really.

This is a web site about the silver particle.

See the "Reactions" part which is in the next " Nanoscience".

http://www.biophysica.com/water_science/biophysical_properties_of_metals/silver.php

[vampares]

I think that the element may produce some free radicals. Either during the process of oxidation, or, as a catalytic property of the Ag metal or the ion (not unlike platinum catalyzes oxidation reactions).

 

It is also in the periodic column with copper. Silver may act as a nuisance element. Copper is an essential component the SOD (superoxide dismutase) enzymes. This being a critical step in aerobic metabolic processes. In the case of humans, iron would transfer oxygen to the copper bearing enzyme. I imagine treatments yield a high degree of lethargy.

 

Some stages of "anaerobic" respiration may also be able to utilize SOD.

 

It may act as a potent Aromatase inhibitor.

BUT, "How does silver kill microbes?" I don't know that it does. Bleach kill microbes. But this requires a certain concentration of bleach.

Edited February 1, 2013 by vampares

[vampares]

 

"Reactions"

 

 

 

 

http://en.wikipedia.org/wiki/Silver_perchlorate

 

Silver perchlorate is noteworthy for its solubility in aromatic solvents such as benzene (52.8 g/L) and toluene (1010 g/L).[1] In these solvents, the silver cation binds to the arene [aromatic hydrocarbon], as has been demonstrated by extensive crystallographic studies on crystals obtained from such solutions.[2][3] It is also amazingly soluble in water, up to 500 g per 100 ml of water.

Silver percholrate is a catalyst for Diels–Alder reaction. http://en.wikipedia.org/wiki/Diels–Alder_reaction

 

Following these reactions, it would not be unreasonable to suspect that Ag+ ions could create a variety of dioxins, furans, PCBs, ect, inadvertently in the environment.

 

 

Mechanism of silver nanoparticles action on insect pigmentation reveals intervention of copper homeostasis.

(note there is no suggestion this is a pesticide)

 

Biochemical analysis suggests that the activity of copper dependent enzymes, namely tyrosinase and Cu-Zn superoxide dismutase, are decreased significantly following the consumption of AgNPs, despite the constant level of copper present in the tissue.

[John Cuthber]

This is a web site about the silver particle.

See the "Reactions" part which is in the next " Nanoscience".

http://www.biophysica.com/water_science/biophysical_properties_of_metals/silver.php

You just cited some page that says "One is the reaction that produces ethylene oxide (the basic building block for flexible plastics)," and yet you still expect to be taken seriously?

[renshen1957]

These may be of some interest to this topic:

 

Study Title:

A review of the use of silver in wound care: facts and fallacies.

Study Abstract:

This review traces the use of silver in wound care, discussing its merits as an antibacterial agent and constituent of many new dressings, which are increasingly tailored to the treatment of wounds ranging from acute surgical lesions to chronic and diabetic leg ulcers. Misconceptions regarding the biological properties of silver, its possible physiological value in the human body and wound bed, absorption through the skin, and safety factors are addressed. The article aims to present silver and the new range of sustained silver-release dressings as important features in the management of skin wounds, providing effective control of wound infections while ensuring patient comfort and quality of life.

Selected quotes from study:

Discussion on the presumed mechanism(s) of silver and related compounds should acknowledge that:
Silver is a broad-spectrum antibiotic36.
Organisms (especially bacteria) show a low propensity to develop resistance to silver-based products.37,38

From the earliest reliable studies, the microbicidal action of silver products has been directly related to the amount and rate of silver released and its ability to inactivate target bacterial and fungal cells.39 The oligodynamic microbicidal action of silver compounds at low concentrations probably does not reflect any remarkable effect of a comparatively small number of ions on the cell, but rather the ability of bacteria, trypanosomes and yeasts to take up and concentrate silver from very dilute solutions.4,5,40

Therefore, bacteria killed by silver may contain 105–107 Ag+ per cell, the same order of magnitude as the estimated number of enzyme-protein molecules per cell.41 In culture media, uptake and toxicity of silver ions in Pseudomonas stutzeri is influenced by sodium chloride, which results in precipitation of relatively insoluble silver chloride.42 Kuschner found variations in the sensitivity of mutant strains of Salmonella typhimurium to the biocidal action of metals such as copper, cobalt, nickel and chromium, whereas both parent and mutant strains of the bacterium remained equally sensitive to silver (and mercury).43

Chemically, metallic silver is relatively inert but its interaction with moisture on the skin surface and with wound fluids leads to the release of silver ion and its biocidal properties. Silver ion is a highly reactive moiety and avidly binds to tissue proteins, causing structural changes in bacterial cell walls and intracellular and nuclear membranes.44 These lead to cellular distortion and loss of viability.45,46

Silver binds to and denatures bacterial DNA and RNA, thereby inhibiting replication.47,48 A recent study demonstrated the inhibitory action of silver on two strains of Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. It found that silver-nitrate exposure lead to the formation of electron-light regions in their cytoplasm and condensation of DNA molecules.49

Granules of silver were observed in the cytoplasm, but RNA and DNA damage and protein inactivation seemed to be the principal mechanisms for bacteriostasis. Intracellular protective mechanisms against silver differed in the Gram-positive and Gram-negative bacteria. The action of silver on bacterial infections in water supplies has also increased our understanding of its microbicidal action. Cell penetration of silver is considered the principal objective in the development of copper/silver ionisation techniques.50

Positively charged copper ions form electrostatic bonds at negatively charged sites on bacterial cell walls, and the resulting damage permits the uptake and release of silver ions. Silver-related degenerative changes in bacterial RNA and DNA, mitochondrial respiration and cytosolic protein lead to cell death. Silver filters and metal used in the control of Legionella suggest that silver and copper ion concentrations are 40:400g/l respectively.51

The action of silver ion on cell walls is illustrated by reference to the yeast Candida albicans. Silver has been shown to inhibit the enzyme phosphomannose isomerase (PIM) by binding cysteine residues.52 This enzyme plays an essential role in the synthesis of the yeast cell wall, and defects lead to the release of phosphate, glutamine and other vital nutrients.53 Silver ion did not inhibit PIM in Escherichia coli cultures.53

Recent literature suggests that the microbicidal action of silver products is partly related to the inhibitory action of silver ion on cellular respiration and cellular function, although the contribution made by ‘other’ silver radicals generated is also acknowledged.54 The exact nature of these silver radicals is not clear but Ovington44 noted that nanocrystalline silver products (Acticoat, Smith and Nephew) can release a cluster of highly reactive silver cations and radicals, which provide a high antibacterial potency on account of unpaired electrons in outer orbitals. Silver and silver radicals released from Acticoat also cause impaired electron transport, bacterial DNA inactivation, cell membrane damage, and binding and precipitation of insoluble complexes with cytosolic anions, proteins and sulphydryl moieties.5,50,55

Lansdown AB. A review of the use of silver in wound care: facts and fallacies. Br J Nurs. 2004 March 13(6 Suppl):S6-19.
Imperial College London, London, England, UK.

AND

Interaction of silver nanoparticles with HIV-1

Jose L Elechiguerra1, Justin L Burt1, Jose R Morones1, Alejandra Camacho-Bragado2,Xiaoxia Gao2, Humberto H Lara3 and Miguel J Yacaman1,2*

*Corresponding author: Miguel J Yacaman yacaman@che.utexas.edu

Author Affiliations

1Department of Chemical Engineering, The University of Texas at Austin, Austin, Texas 78712, USA

2Texas Materials Institute, The University of Texas at Austin, Austin, Texas 78712, USA

3Facultad de Ciencias Biológicas, Universidad Autónoma de Nuevo León, San Nicolás de los Garza, Nuevo León, Mexico

For all author emails, please log on.

Journal of Nanobiotechnology 2005, 3:6 doi:10.1186/1477-3155-3-6

The electronic version of this article is the complete one and can be found online at:http://www.jnanobiotechnology.com/content/3/1/6

Received: 28 March 2005

Accepted: 29 June 2005

Published: 29 June 2005

© 2005 Elechiguerra et al; licensee BioMed Central Ltd.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

The interaction of nanoparticles with biomolecules and microorganisms is an expanding field of research. Within this field, an area that has been largely unexplored is the interaction of metal nanoparticles with viruses. In this work, we demonstrate that silver nanoparticles undergo a size-dependent interaction with HIV-1, with nanoparticles exclusively in the range of 1–10 nm attached to the virus. The regular spatial arrangement of the attached nanoparticles, the center-to-center distance between nanoparticles, and the fact that the exposed sulfur-bearing residues of the glycoprotein knobs would be attractive sites for nanoparticle interaction suggest that silver nanoparticles interact with the HIV-1 virus via preferential binding to the gp120 glycoprotein knobs. Due to this interaction, silver nanoparticles inhibit the virus from binding to host cells, as demonstrated in vitro.

Background

Nanotechnology provides the ability to engineer the properties of materials by controlling their size, and this has driven research toward a multitude of potential uses for nanomaterials[1]. In the biological sciences, many applications for metal nanoparticles are being explored, including biosensors[2], labels for cells and biomolecules[3], and cancer therapeutics[4].

It has been demonstrated that, in the case of noble-metal nanocrystals, the electromagnetic, optical and catalytic properties are highly influenced by shape and size [5-7]. This has driven the development of synthesis routes that allow a better control of morphology and size [8-13]. Noble-metal nanomaterials have been synthesized using a variety of methods, including hard-template[14], bio-reduction[9] and solution phase syntheses[8,10-13].

Among noble-metal nanomaterials, silver nanoparticles have received considerable attention due to their attractive physicochemical properties. The surface plasmon resonance and large effective scattering cross section of individual silver nanoparticles make them ideal candidates for molecular labeling[15], where phenomena such as surface enhance Raman scattering (SERS) can be exploited. In addition, the strong toxicity that silver exhibits in various chemical forms to a wide range of microorganisms is very well known [16-18], and silver nanoparticles have recently been shown to be a promising antimicrobial material[19].

For these reasons, and based upon our previous work regarding interactions of noble metal nanoparticles with biomolecules[20], we decided to study the interaction of silver nanoparticles with viruses. Herein, we present the first findings of our investigation, the discovery that silver nanoparticles undergo size-dependent interaction with HIV-1.

Findings

Characterization of the tested silver nanoparticle preparations

The physicochemical properties of nanoparticles are strongly dependent upon their interactions with capping agent molecules[21]. Indeed, the surface chemistry of the nanoparticles can modify their interactions with external systems. For this reason we tested silver nanoparticles with three markedly different surface chemistries: foamy carbon, poly (N-vinyl-2-pyrrolidone) (PVP), and bovine serum albumin (BSA).

Foamy carbon-coated nanoparticles were obtained from Nanotechnologies, Inc., and used without further treatment. These nanoparticles are embedded in a foamy carbon matrix which prevents coalescence during their synthesis. The as-received nanoparticle sample consists of a fine black powder. For the purposes of the present work, the as-received powder was dispersed in deionized water by ultra-sonication. TEM analysis shows that the nanoparticles tend to be agglomerated inside the foamy carbon matrix, although a significant fraction of the population is released from this matrix by the energy provided from the ultra-sonic bath (Figure 1a–1f). These released nanoparticles are mainly free-surface nanoparticles, and it was observed that only nanoparticles that have escaped from the foamy carbon matrix interact with the HIV-1 cells.

Figure 1. Transmission electron microscopy (TEM) of the foamy carbon-coated silver nanoparticles. a) TEM image of the sample prepared by dispersing the as-received powder in deionized water by ultra-sonication. The agglomeration of particles inside the foamy carbon matrix is observed. b) TEM image of nanoparticles outside of the carbon matrix. The broad distribution of shapes can be observed. c)-f) TEM images of nanoparticles with different morphologies. c) Icosahedral. d) Decahedral. e) Elongated. f) Octahedral. g) High Resolution TEM image of the foamy carbon matrix.

The interaction of the nanoparticles with the foamy carbon matrix is sufficiently weak that simply by condensing the TEM electron beam, even those nanoparticles that were not initially released by ultra-sonication are ejected from the foamy carbon agglomeration. In fact, after this experiment the complete size distribution of these nanoparticles is better observed, please refer to 1. High resolution transmission electron microscopy (TEM) revealed that the silver nanoparticles released from the foamy carbon matrix by ultrasonication have a size distribution of 16.19 ± 8.68 nm (Figure 2a–b). By releasing the remaining nanoparticles from the foamy carbon matrix with the action of the electron beam, the average size was ~21 ± 18 nm. Additionally, TEM examination demonstrated that the sample is composed of several morphologies including multi-twinned nanoparticles with five-fold symmetry, i.e. decahedra and icosahedra, truncated pyramids, octahedral and cuboctahedral nanoparticles, among others (Figure 1c–1f).

Figure 2. Silver nanoparticle preparations. a) TEM image of free surface silver nanoparticles released from the foamy carbon matrix by dispersing the as-received powder in deionized water by ultra-sonication. b) Size distribution of free surface nanoparticles measured by TEM analysis. c) UV-Visible spectrum of carbon-coated silver nanoparticles. d) HAADF image of PVP-coated silver nanoparticles. e) Size distribution of PVP-coated nanoparticles measured by TEM analysis. f) UV-Visible spectrum of PVP-coated silver nanoparticles. g) HAADF image of BSA-coated silver nanoparticles. h) Size distribution of BSA-coated nanoparticles measured by TEM analysis. i) UV-Visible spectrum of BSA-coated silver nanoparticles.

PVP-coated nanoparticles were synthesized by the polyol method using glycerine as both reducing agent and solvent. In this method, a metal precursor is dissolved in a liquid polyol in the presence of a capping agent such as PVP[22]. PVP is a linear polymer and stabilizes the nanoparticle surface via bonding with the pyrrolidone ring. Infrared (IR) and X-ray photoelectron spectroscopy (XPS) studies have revealed that both oxygen and nitrogen atoms of the pyrrolidone ring can promote the adsorption of PVP chains onto the surface of silver[23]. The sample size distribution was obtained from high angle annular dark field (HAADF) images. The nanoparticles exhibited an average size of 6.53 nm with a standard deviation of 2.41 nm. (Figure 2d–e)

Silver nanoparticles directly conjugated to BSA protein molecules were synthesized in aqueous solution. Serum albumin is a globular protein, and is the most-abundant protein in blood plasma. Bovine serum albumin (BSA) is a single polypeptide chain composed of 583 amino acid residues[24]. Several residues of BSA have sulfur-, oxygen-, and nitrogen-bearing groups that can stabilize the nanoparticle surface. The strongest interactions with silver likely involve the 35 thiol-bearing cysteine residues. By using sodium borohydride, a strong reducing agent, BSA stabilizes nanoparticles via direct bonding with these thiol-bearing cysteine residues, and provides steric protection due to the bulkiness of the protein. The sample size distribution was obtained from HAADF images. Nearly 75% of the BSA-conjugated silver nanoparticles were 2.08 ± 0.42 nm in diameter, but a substantial fraction of larger particles was also observed, bringing the total average to 3.12 ± 2.00 nm (Figure 2g–h).

UV-visible spectroscopy is a valuable tool for structural characterization of silver nanoparticles. It is well known that the optical absorption spectra of metal nanoparticles are dominated by surface plasmon resonances (SPR), which shift to longer wavelengths with increasing particle size [25]. Also, it is well recognized that the absorbance of silver nanoparticles depends mainly upon size and shape [26,27]. In general, the number of SPR peaks decrease as the symmetry of the nanoparticle increases [27]. Recently, Schultz and coworkers[28] correlated the absorption spectra of individual silver nanoparticles with their size (40–120 nm) and shape (spheres, decahedrons, triangular truncated pyramids and platelets) determined by TEM. They found that spherical and roughly spherical nanoparticles, decahedral or pentagonal nanoparticles, and triangular truncated pyramids and platelets absorb in the blue, green and red part of the spectrum, respectively. In all the cases the SPR peak wavelength increases with size, as expected.

The UV-Visible spectra for all the nanoparticle preparations are shown in Figure 2. All samples presented a minimum at ~320 nm that corresponds to the wavelength at which the real and imaginary parts of the dielectric function of silver almost vanish [27]. The sample with carbon-coated silver nanoparticles exhibits four peaks at ~400, ~490, ~560 and ~680 nm, as shown in Figure 2c. The optical signature of this sample can be better understood in terms of the distribution of sizes and shapes observed in the TEM. As we previously mentioned, the distribution of shapes in the sample is broad, and a significant amount of nanoparticles are not spherical such as multi-twinned with five-fold symmetries. The presence of nanoparticles with pentagonal and triangular cross-sections could be responsible for the absorption at longer wavelengths. Thus, it is clear that the characteristic absorption of these nanoparticles arises from the contribution of different shapes and sizes, which agrees with the TEM observations.

On the other hand, the PVP-coated and BSA-coated silver nanoparticles present only one peak at ~450 and ~390 nm, respectively. These results indicate that both preparations are mainly composed by small spherical silver nanoparticles. It is also well know that for small particles a broadening of the plasmon absorption bands is expected, since there is a linear dependence of the full-width at half maximum (FWHM) with the reciprocal of the particle diameter[29]. The results for BSA-coated nanoparticles agree with the last statement, presenting just one broad symmetric peak at ~390 nm. On the other hand, the spectrum for the PVP-coated silver nanoparticles is not symmetric around the maximum absorption wavelength. In fact, this spectrum can be deconvoluted in two different curves, one centered at ~430 and another one at ~520 nm. The peak at ~430 nm could be assigned to the out-of-plane dipole resonance of the silver nanoparticles indicating the presence of spherical particles with small diameters. In addition, the synthesis of silver nanoparticles by the polyol method in presence of PVP promotes also the formation of multi-twinned nanoparticles (MTPs), being decahedra nanoparticles the most thermodynamically stable MTPs [23]. Therefore, the observed read shift is a consequence of both nanoparticles of larger size and the presence of decahedral nanoparticles with pentagonal cross sections.

Interaction with HIV-1

High angle annular dark field (HAADF) scanning transmission electron microscopy was employed to study the interaction of silver nanoparticles with HIV-1. In our previous works, HAADF has proven to be a powerful technique for analysis of biological samples, such as proteins[20] and bacteria[30], interfaced with inorganic nanoparticles. HAADF images are primarily formed by electrons that have undergone Rutherford backscattering. As a result, image contrast is related to differences in atomic number [31] with intensity varying as ~Z2. Therefore, image contrast is strongly related to composition. As a good approximation, lighter elements appear dark and heavier elements appear bright. Due to a large difference in atomic number, silver nanoparticles are easily distinguished from the organic matter that composes the virus.

In Figure 3, we present HAADF images of the HIV-1 virus with (3a) and without (3b) silver nanoparticles. For complete experimental details, refer to Methods Section. The presence of silver was independently confirmed by Energy Dispersive X-ray Spectroscopy (EDS), shown in Figure 3c. Interestingly, the sizes of nanoparticles bound to the virus (Figure 3d) were exclusively within the range of 1–10 nm. In the case of the silver nanoparticles released from the carbon matrix, the fact that no nanoparticles greater than 10 nm in diameter were observed to interact with the virus is significant, since the size of ~40% of the overall population is beyond this range. This provides strong evidence for the size-dependence of interaction.

Figure 3. HAADF images of the HIV-1 virus. a) HAADF image of an HIV-1 virus exposed to BSA-conjugated silver nanoparticles. Inset shows the regular spatial arrangement between groups of three nanoparticles. b) HAADF image of HIV-1 viruses without silver nanoparticle treatment. Inset highlight the regular spatial arrangement observed on the surface of the untreated HIV-1 virus. c) EDS analysis of image a) confirming the presence of Ag. The C signal comes from both the TEM grid and the virus, O, and P are from the virus, and Na, Cl, and K are present in the culture medium. Ni and Si come from the TEM grid, while Cu is attributed to the sample holder. d) Composite size distribution of silver nanoparticles bound to the HIV-1 virus, derived from all tested preparations.

Additionally, the nanoparticles seen in Figure 3a are not randomly attached to the virus, as regular spatial relationships are observed among groups of three particles. Both the spatial arrangement of nanoparticles and the size dependence of interaction can be explained in terms of the HIV-1 viral envelope, and can provide insight into the mode of interaction between the virus and nanoparticles.

The exterior of the HIV-1 virus is comprised of a lipid membrane interspersed with protruding glycoprotein knobs, formed by trimers consisting of two subunits: the gp120 surface glycoprotein subunit is exposed to the exterior, and the gp41 transmembrane glycoprotein subunit spans the viral membrane and connects the exterior gp120 glycoprotein with the interior p17 matrix protein[32]. The main function of these protruding gp120 glycoprotein knobs is to bind with CD4 receptor sites on host cells. Numerous cellular proteins are also embedded within the viral envelope[33]. However, the protruding gp120 glycoprotein knobs are more exposed to the exterior, and should be more accessible for potential nanoparticle interactions.

Leonard and coworkers[34] reported that the gp120 subunit has nine disulfide bonds, three of which are located in the vicinity of the CD4 binding domain. These exposed disulfide bonds would be the most attractive sites for nanoparticles to interact with the virus. As mentioned previously, the nanoparticles in Figure 1a appear to be located at specific positions, with regular spatial relationships observed among groups of three particles. The observed spatial arrangements correlate with the positions of the gp120 glycoprotein knobs in the structural model for HIV-1 proposed by Nermut and coworkers[32].

Regular spatial relationships are also found on the surface of the untreated virus, as seen in the inset of Figure 1b. The observed darker contrast at these sites could indicate the locations of the glycoprotein knobs. As mentioned previously, contrast in HAADF images is strongly dependent on differences in atomic number. However, this is not the only factor in determining the image contrast. If the material is composed of elements of similar atomic numbers, as is the case for the organic constituents of the pure virus, local variations in sample density will provide noticeable contrast. The majority of the viral envelope consists of a densely-packed lipid membrane. However, for the glycoprotein knobs, we would expect a localized region of lower density due to the presence of membrane-spanning gp41 glycoproteins rather than the densely-packed lipids. Hence, these areas should appear darker than the rest of the viral envelope.

It has previously been determined that the center-to-center spacing between glycoprotein knobs is ~22 nm[35]. In the inset of Figure 3a, the average measured center-to-center spacing between silver nanoparticles is ~28 nm, which correlates with the expected spacing between glycoprotein knobs. The average center-to-center spacing between the small darker regions on the untreated virus is ~22 nm, which again suggests these sites are the gp120 glycoprotein knobs. Thus, the observed spatial arrangement of nanoparticles, the center-to-center distance between nanoparticles, and the fact that the exposed sulfur-bearing residues of the glycoprotein knobs would be attractive sites for nanoparticle interaction strongly suggest that silver nanoparticles interact with the HIV-1 virus via preferential binding to the gp120 glycoprotein knobs.

Presuming that the most attractive sites for interaction are the sulfur-bearing residues of the gp120 glycoprotein knobs, there are only a limited number of bonds that a nanoparticle can form. This limited number of stabilizing sites can explain why larger nanoparticles are not observed to attach to the virus. Assuming that each nanoparticle interacts with a single glycoprotein knob, and that each nearest-neighbor knob is occupied by another nanoparticle, from geometric considerations the theoretical upper limit for the diameter for these nanoparticles would be ~20 nm. However, if a nanoparticle larger than the diameter of one knob (~14 nm[35]) were to be attached, only a small fraction of the total nanoparticle surface would be anchored, resulting in a less stable interaction. Thus, if the nanoparticles are interacting with HIV-1 via preferential binding at gp120 glycoprotein knobs, we would expect to find mostly nanoparticles less than 14 nm in diameter, as particles in this size range would have the most stable surface interactions. This corresponds closely with our experimental observation that particles greater than 10 nm were not attached to the viral envelope.

Although the mechanism by which HIV infects host cells is not yet fully understood, there are two steps that are broadly agreed to be critical. The first step involves binding of gp120 to the CD4 receptor site on the host cell. Then, upon binding to CD4, a conformational change is induced in gp120, resulting in exposure of new binding sites for a chemokine receptor, i.e. CCR5 or CXCR4[36-38]. An agent that preferentially interacts with the gp120 glycoprotein would block the virus from binding with host cells. Therefore, we measured the inhibitory effects of silver nanoparticles against HIV-1 in vitro.

The capacity of silver nanoparticles to inhibit HIV-1 infectivity was determined by testing against CD4+ MT-2 cells and cMAGI HIV-1 reporter cells. For complete experimental details, refer to Methods Section. The cytopathic effects of CD4+ MT-2 infection were analysed by optical microscopy assessment of syncytium formation as described elsewhere[39,40], as well as by the HIV-1 infection of cMAGI cells using the Blue Cell Assay[41,42]. The cytotoxicity of all the nanoparticle preparations against MT-2 cells was determined using the Trypan Blue exclusion assay [43]. For all three nanoparticle preparations, at silver concentrations above 25 μg/mL, viral infectivity was reduced to an extent that it could not be detected by syncytium formation, as shown graphically in Figure 4. For each nanoparticle preparation, we found a dose-dependant inhibition of HIV-1 infectivity, with an IC50 where only moderate cell toxicity was observed, as seen in Figure4.

Figure 4. Inhibition of HIV-1 and toxicity data. a) Assessment of HIV-1 mediated syncytium formation in MT-2 cells. b) Percentage of HIV-1 transmission in cMAGI cells. The toxicity of the nanoparticle preparations against MT-2 cells was determined using the Trypan Blue exclusion assay. The samples were incubated at 37°C, and the cells were evaluated via optical microscopy after c) 3 h and d) 24 h of exposure to silver nanoparticles.

Although the findings regarding interaction with HIV-1 were congruent among nanoparticles with markedly different surface chemistry, the toxicity and inhibition results were not the same. The differences in the observed trends in HIV-1 inhibition can be explained in terms of the capping agents employed for each nanoparticle preparation. BSA- and PVP-protected nanoparticles exhibit slightly lower inhibition because the nanoparticle surface is directly bound to and encapsulated by the capping agent. In contrast, the silver nanoparticles released from the carbon matrix have a greater inhibitory effect due to their essentially free surface area. The fact that the carbon-coated nanoparticles present higher cytotoxicity can also be explained in terms of surface chemistry. Since a significant amount of these silver nanoparticles possess nearly free surfaces, they are able to interact stronger with the host cells, thus increasing their toxicity. Clearly, selection of capping agents will be crucial for future research on the interaction of nanoparticles with viruses, microorganisms, and more complex biosystems, and many more variables require further testing, including the long-term effects of the presence of nanoparticles, and the impact of traces of precursor molecules and reaction by-products.

In conclusion, we have found that silver nanoparticles undergo size-dependent interaction with HIV-1, and that the bound particles exhibit regular spatial relationships. These observations lead us to suggest that the nanoparticles undergo preferential binding with the gp120 subunit of the viral envelope glycoprotein. Silver nanoparticles inhibit the HIV-1 virus infectivity in vitro, which also supports our proposal regarding preferential interaction with gp120. These findings only provide indirect evidence for our proposed mode of interaction, and we are currently undertaking testing to determine conclusively if direct conjugation between gp120 and silver nanoparticles exist.

The interactions of inorganic nanoparticles with biosystems are just beginning to be understood, and potential applications are being discovered at an increasing rate. However, in order to realize the future promise of nanoscience, it is imperative that the toxicity and long-term health effects of exposure to nanomaterials be fully explored. The flexibility of nanoparticle preparation methods, the multitude of functionalization techniques, and facile incorporation of nanoparticles into a variety of media provide the incentive for further research on the interaction of metal nanoparticles with viruses.

Methods

a) HIV-1 strains and cell lines

HIV-1IIIB laboratory strain of HIV-1 an X4 wild type (wt) virus was obtained through the AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH. CD4+ MT-2 cell line was obtained from the American Type Culture Collection. The cMAGI HIV-1 reporter cells were a generous gift from Dr. Phalguni Gupta from the University of Pittsburgh. All other reagents used were of the highest quality available.

cMAGI cells were cultured in DMEM Dulbecco's Modified Eagle Medium (DMEM) (1X) liquid without sodium phosphate and sodium pyruvate. The medium contained 4,500 mg/L D-glucose and L-glutamine (Invitrogen, Paisley, UK), with 10% fetal calf serum (FCS), 0.2 mg/mL geneticin (G418), and 0.1 μg/mL puromycin. MT-2 cells were cultured in RPMI 1640, containing 10% fetal calf serum (FCS) and antibiotics.

HIV-1IIIB primary clinical isolates were propagated by subculture in MT-2 and cMAGI cells. HIV-1IIIB was reproduced according to the DAIDS Virology Manual for HIV Laboratories, version 1997, compiled by Division of AIDS of the National Institute of Allergies and Infectious Diseases and the National Institute of Health, and Collaborators. Aliquots of cell-free culture viral supernatants were used as viral inocula.

All the work related to HIV-1 cells, except for TEM analysis, was done in a Biosafety Level 3 (BSL-3) Laboratory.

b) Synthesis of the three different silver nanoparticle preparations

Carbon coated silver nanoparticles tested in this study were obtained from Nanotechnologies, Inc. and used without further treatment. For more information about the synthesis of these nanoparticles, please visit http://www.nanoscale.com webcite

PVP-coated silver nanoparticles were synthesized by the polyol method using glycerine as both reducing agent and solvent. Silver sulfate (Ag2SO4, reagent grade) and poly (N-vinyl-2-pyrrolidone) (PVP-K30, MW = 40,000) were purchased from Sigma Aldrich and 1,2,3-Propanetriol (Glycerin, >99%) was purchased from Fischer Chemicals, all the materials were used without any further treatment. Briefly, we added 0.2 g of PVP to a round bottom flask following by the addition of 30 mL of glycerin. Once PVP was dissolved, we increased the temperature to 140°C. After 30 minutes we added 2 mL of 0.015 M Ag2SO4 and left to react for 1 h.

Silver nanoparticles directly conjugated to bovine serum albumin (BSA) protein molecules were produced as following described. Silver nitrate (AgNO3, 0.945 M), sodium borohydride (NaBH4, 99%) and 200 proof spectrophotometric-grade ethanol were purchased from Aldrich. Bovine serum albumin (BSA) was purchased from Fisher and was used without further treatment. Briefly, sodium borohydride was added to an aqueous solution of silver nitrate and BSA under vigorous stirring. The molar ratio of Ag+:BSA was 28:1, and the molar ratio of Ag+:BH4- was 1:1. The reaction volume was 40 mL, and contained 13.50 μmol BSA. The reaction was allowed to proceed for 1 h, and the product was purified by precipitation at -5°C, followed by cold ethanol filtration.

c) Characterization of the different silver nanoparticle preparations

Transmission electron microscope was conducted in a HRTEM JEOL 2010F microscope equipped with Schottky-type field emission gun, ultra-high resolution pole piece (Cs = 0.5 mm), and a scanning transmission electron microscope (STEM) unit with high angle annular dark field (HAADF) detector operating at 200 kV. Briefly, a droplet of each different solution of silver nanoparticles was placed on a Cu grid with lacey carbon film (Ted Pella), and allowed to evaporate. Size distributions for each nanoparticle preparation were obtained from TEM analysis based on the measurement of 400 particles, and 600 particles in the case of BSA-coated nanoparticles.

UV-visible spectra were obtained at room temperature using a 10 mm path length quartz cuvette in a Cary 5000 spectrometer. All the solutions were diluted 30 × in deionized water before acquiring the spectra.

d) Electron microscopy of HIV-1 and silver nanoparticles

Samples were prepared for electron microscopy as follows: 105 TCID50 samples of HIV-1IIIB cell free virus were treated with solutions of the different silver nanoparticles at a concentration of 100 μg/mL. After 30 seconds, a 10 μL droplet was deposit on a carbon coated nickel TEM grid and exposed to a 2.5% solution of PBS/glutaraldehyde vapors for 30 minutes. Microscopy was done using a JEOL 2010-F TEM equipped with an Oxford EDS unit, at an accelerating voltage of 200 kV and operated in scanning mode using an HAADF detector.

e) Inhibition of HIV-1 with silver nanoparticles

RPMI medium only or containing varying concentrations of silver nanoparticles were mixed with samples 105 TCID50 of HIV-1IIIB cell free virus. The highest concentration of silver nanoparticles used was 100 μg/mL. After 30 seconds, sequential 2-fold dilutions of the solutions were added to cultures of target cells (2 × 105 MT-2 and 2 × 105 cMAGI HIV-1 reporter cells with 0.2–0.5 multiplicity of infection (m.o.i) of HIV-1IIIB virus) prepared as previously mentioned. Each dilution was exposed to four replicate wells. After that, the cells were incubated in a 5% CO2 humidified incubator at 37°C for 3–5 days. Assessment of HIV-1 mediated syncytium formation was performed for the MT-2 cells, while for cMAGI cells, the percentage of transmission was estimated as follows: the number of blue-stained cells obtained from the supernatant of each of the tested wells was divided by the number of blue-stained cells obtained from the culture supernatant in the well of the positive control.

f) Cytotoxicity of silver nanoparticles against MT-2 cells

The cytotoxicity of the nanoparticle preparations against MT-2 cells was determined using the Trypan Blue exclusion assay. In all cases, the initial concentration of silver nanoparticles was 50 μg/mL and sequential two-fold dilutions were made and mixed with 2 × 105 MT-2 cells. The samples were incubated at 37°C, and the cells were evaluated via optical microscopy after 3 h and 24 h of exposure to silver nanoparticles. Briefly, an aliquot of the cell suspension was diluted 1:1 (v/v) with 0.4% Trypan Blue and the cells were counted using a haemocytometer. Viability was expressed as the percentage of number of unstained treated cells to that of the total number of cells.

Additional File 1. Supporting information. The file is a word document that contains the complete size distribution of the carbon-coated silver nanoparticles evaluated by TEM

Format: DOC Size: 177KB

This file can be viewed with:

Acknowledgements

The authors want to thank Nanotechnologies, Inc. for providing their silver nanoparticles. J. L. Elechiguerra, J. R. Morones, and A. Camacho-Bragado acknowledge the support received from CONACYT- México. J.L.Burt thanks the University of Texas at Austin College of Engineering and Mr. Robert L. Mitchell for their financial support through the Thrust 2000 Robert L. and Jane G. Mitchell Endowed Graduate Fellowship in Engineering. This material is based upon work supported under a National Science Foundation Graduate Research Fellowship (to JLB).

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"

Acute infection

Hepatitis C infection causes acute symptoms in 15% of cases.^[5] Symptoms are generally mild and vague, including a decreased appetite, fatigue, nausea, muscle or joint pains, and weight loss.^[6] Most cases of acute infection are not associated with jaundice.^[7] The infection resolves spontaneously in 10-50% of cases, which occurs more frequently in individuals who are young and female"

 

from

http://en.wikipedia.org/wiki/Hepatitis_C

 

Assuming that the disappearance of the infection is due to the silver is a logical fallacy.

http://en.wikipedia.org/wiki/Post_hoc_ergo_propter_hoc

 

However it is known that the colloidal silver is toxic.

 

The mechanisms by which silver is microbicidal are not well targeted towards viruses.

I wonder if that is the reason that Pure Bioccience Axen 30 (and other products) received EPA approval as a hard surface cleaner? http://www.purebio.com/products/sdc_hard_surface:

 

The Link is to the graphic (prohibited from posting on this forum) with viral pathogens killed:

 

HIV I Type 1

Herpes Simplex I

Rotavirus

Respiratory Syncytial Virus

Human Coronavirus

Norovirus

Murine Norovirus

Avian Influenza A

Influenza A

Influenza A (H1N1)

Swine Influenza A (H1N1)

Adenovirus

Rhinovirus

Polio Type 2

For those of you who speak or read Russian:

 

“Activity of colloidal silver preparations towards smallpox virus,

Pharmaceutical Chemistry Journal,” N. E. Bogdanchikova, A. V. Kurbatov,

V. V. Tretyakov, P. P. Rodionov. 26, 9-10, 778 (1992)

The above was cited by the EPA as part of the reason for silver to be regulated under FIFRA.

Best regards

Steve