Highly concentrated heavy metal stocks have an acidic pH. For cadmium we have experienced pH=3.0. When pH rises they complex with ions (different metal species are present depending on pH and salt with different properties). So you have to find a tradeoff between pH toxicity and heavy metal concentration.

Precipitation of metals can be observed while using seawater.. when the concentration of the metal is more... as Dr. Xavuer said due to acidic pH.. some fraction of the salt transforms from dissolved to particulate phase and gets precipitated. I had this problem while using Copper sulphate so I used to adjust the pH. How much difference to u observe in ur dose and measure concentration?

Dear Pedram, precipitation will depend on chemical species present in the medium and highest metal concentrations tested. Could you detail these informations, since it's difficult to answer your question.

For the second question : geometric series in toxicity tests allow a constant difference between logarithms of concentrations, which results in optimization of concentration-effect relationship (linear relationship between effect and log(concentration)) and better estimation of the EC50. For an effluent, it is generally recommended to simply dilute by 2 (conc : 100-50-25-12.5% etc.), at least for a screening test, and for substances one can choose the following range : 18 - 10 - 5.6 - (unit : µg/L or mg/L).

Yes. It is generally recommended to simply dilute by 2 (conc : 100-50-25-12.5% etc.), at least for a screening test, and for substances one can choose the following range : 18 - 10 - 5.6 - (unit : µg/L or mg/L).

Yes. You have to find a trade off between pH toxicity and heavy metal concentration.

I think it can also depends on salt solubility and formation of different salt with "medium" ions. To try to avoid the problem you can choose another salt if you hare interested only on the metal ions.

It´s been said by previous comments, but I want to emphasize it. Your test water is too hard, and you are precipitating cadmiun carbonate. Either you decrease your Cd concentrations or lower the pH.

I would only like to add that there's no ecological relevance in performing an acute test beyond the solubility of the compound, so forcing the solubility just to obtain an LC50 may be too much, you might be better of calculating an LC20 or any other with your highest concentration at the highest solubility. Regarding using geometric series, the idea is to have, ideally, two concentration above your LC50, two below and one very close to his parameter so that when you apply either probit or logit transormation to your data the confidence limits are not too wide.

As far as my experience is concerned, metals have a good solubility at lower pH. based on your way of approaching the toxicity, it is highly difficult. because your sample has already precipitated and the bio-availability will also decrease simultaneously. what i recommend is that , first adjust the pH and then after proceed with the EPA standard for acute water toxicity testing.

When you use seawater or ( hardwater when using freshwater) precipitation of metals occur due to coprecipitation when the metals form metals with chlorides ( as in Nacl) in seawater) dissolved organic matter and other chemical compounds in the water. Cu and Pd are metals prone to precipitation. One way around problem so that dissolved metal concentrations ( of the metal toxicant) is maintained is to use a continuous flow system ( that puts in a constant flow of the test chemicals in water) . Or change the dilution water and add fresh metal toxicant every 48 hrs. Must monitor dissolved metal concentration in the test water by AAS

reduce the pHof the medium. Feeding should be stopped at least 24 hrs before bioassay and nofeeding be done during this test. reduce the pH of the medium n change the test medium at every 96 hrs. hope this may do.

Thanks for all useful answers and suggestions.

When we reduce the water pH in acute toxicity measurement, the experiment condition (water quality parameters) will be different with our future experiment, which will be prepared based on the obtained details from our LC50 test. In other word, we should reduce our pH in all future experiments due to be exactly close to LC50 test condition but it might be however dangerous for aquatic animals. Anyway, I used semi-static test (renewal every 24 h) previously to hinder metal precipitation but we did not find any mortalities because the precipitation occurred very soon (during first hours).

It is worthy of note that according to OECD protocol No 203, pH range should be 6.00 to 8.5 and hardness are preferred that not exceeding 250 mg CaCo3. We considered this condition to perform our study.

Thanks again!

You are most welcome!

Carbonate salts of lead are extremely insoluble.

Dear Peter,

We used lead acetate (Pb(CH3COO)2·3H2O ) and other metals as a chloride salt.

It is not completely clear what was in your water. But there is lots of carbonate in sea water and in many surface waters. Also substantial amounts in ground water. Thus the mention above of "hard" water. Formation of insoluble carbonate and IIRC sulfate salts is why environmental lead (say at gun ranges) tends to stay put and not get into ground water.

To reiterate: There is considerable carbonate in your water. Likely, this is precipitating out lead. Doesn't matter that you are adding lead as the soluble acetate. Once in solution, lead will from a salt with whatever anion is present.

Peter, how can I stop metal percipitation in water? This is a freshwater and I work with living organism. I cannot reduce the pH or carbonate because I'd like that water quality parameters be simislar to our future experiment, which will be prepared based on obtained results.

Now to answer the second part of your question.

The only reason to use a geometric series in environmental toxicology is as part of a range finding test. Basically you want to bracket the concentrations that have no toxicity and then partial or 100 percent mortality. Then you use concentrations that are selected to produce a concentration-response description of the toxicity to your specific endpoint. This is then the definitive test.

The toxicity ranges of these metals as ions are pretty well known. So did you make sure you used that information to set up your test concentrations.

If there are concentrations that are regularly found in the environment be sure to consider those in your definitive toxicity test.

Please use a regression approach to describing your concentration-response. NOECs and LOECs have severe limitations as descriptions of concentration-response (Landis WG, Chapman PM. 2011. Well past time to stop using NOELS and LOELs. Integr Environ Assess Manag 7:vi-viii. ). The recent editions of textbook by Mike Newman or Landis et al also provide a great deal of guidance on the design of toxicity texts.

If the material precipitates out of the water that is not a concentration that is likely to be useful in determining the concentration-response if your pH and other water quality parameters are within those ranges found in your typical receiving water.

The other reason to use geometric series is that most statistical methods (i. e. Logit, Probit, etc.) requiere the use of a geometric (or log) scale.

I absolutely agree with Wayne, however, dissolved organic matters (DOMs) should be measured in your water, because DOMs can affect the fate and effect of trace metals

Concur that lead at concentrations which precipitate out are unlikely to be of toxicological significane, as a practical matter.

I agree with Peter Proctor.

Do you mean (lethal dose) LD 50 or LC50

LC50!

I do not know how at the given conditions can be calculated LD 50.

What are the concentrations of the HMs you are using in your test?

From 2.5 to 160 mg/l as a geometric series for cadmium and similarly for other HMs.

What kind of salts are you using for preparing your stock/working metal solution? chloride/nitrate/sulfate/?

From 2.5 to 160 mg/l as a geometric series for cadmium and similarly for other HMs.

pH is a key element, I am not sure if you can reduce it or not. You can check the precipitation using geochemical model such as Minteq software.

Yes, pH is important!

Moreover... did you have EDTA or other chelating compound in the composition of your medium ? I work on D.magna and i use EDTA among other salts to grow algae as carbonium source. EDTA presence (also in very low doses) could reduce the apparent toxicity of metals

Dose- letal response dependence is often approximated as a log-linear one, and geometric concentrations (doses) series is a better (although not ideal) choice as compared with an arithmetic progression. As to sedimentation problem (due to formation of hydroxides od heavy metals), I believe that using buffered solutions with pH in neutral-t0-acid range is the simplesr answer

Is very common that metal salts to precipitate in contact with dissolved salts or with organic materials, as fish feces. Your water hardness can be the major responsible by the precipitation and a dilution using a ultrasonic device or more easily water soluble salts can resolve your problem.

Yes. Your water hardness can be the major responsible by the precipitation and a dilution using a ultrasonic device or more easily water soluble salts can resolve your problem.

But if you dilute the sample, or somicate it, then you have again the issue of the "reality" of your experiment, since your solutions will not reflect the real conditions you are trying to represent.

One variable i believe we're missing out is temperature? what is the temperature of the set-up? there is a strong relationship between temperature and the maximum amount of salt water can dissolve.

Second, confirm the purity of the salt being used and the hardness (Ca(HCO3-)2-) of the water being used is also a suspect as others have raised previously.

What is the water source bore-hole or municipal water supply? this may help trace the problem in a way.

This is true. This is a very important point.

Hi Pedram

1. Stabilize the test solution,

2. Control pH, DOC and hardness of the medium,

3. Measurements of metal concentration must be related to effect.

Best

Dola

Hello guys, 

I am trying to study the toxicity of lead acetate on PC12 since 5 months but my lead always participate and I have not seen any  toxicity so far. Any help in this issue will be highly appreciated.

Thanks 

Hi Sarah,

If you have studied above-mentioned comments, you will find the most issue is related to pH and hardness of water. Additionally. It's better to use lead nitrate instead of acetate salt because it is more soluble in water. I had the same problem with acetate and had to change it. Therefore, please check your pH first (acidic water provide more solubility) and if you could not overcome this issue, change your salt.

 Hi Pedram, 

Thank you so much for responding. I did changed the PH of the media and mad it acetic but no things happened. I reduced PH of water by using glutamic acid but also no toxicity. I changed my salt to lead nitrate and lead II acetate instead lead IV acetate and my Pc-12cells still acting so normal after 23, 48, 72 h.

please any body I need suggestion with that. I am working on that for 4 months.

Sarah,

Please ask a new question if you have still the same issue, this is an old question and I think people could hardly hear anything from you.

Please tag your question with more relevant topics like water chemistry or analytical chem.

Thanks,

I asked this question a couple months ago but I have not find good answer except change the cells and I did bought new cells but no toxicity.

To Sarah, You should use acidic medium to prepare lead acetate may be by adding few drops from acetic acid and prepare the mixture freshly every day.