Effective Monitoring of Hydrogen Sulfide Gas in Wastewater for Odor Control; For On-site and Laboratory Analysis

Christopher Altamirano
Lab Manager, Arizona Instrument LLC

Hydrogen sulfide (H2S) is a colorless hazardous gas known for its ‘rotten egg’ odor that many of us can detect even at low levels.[1]  Every time we pass a landfill, a wastewater facility or are brave enough to pry open the long forgotten egg salad container in the back of the fridge we have encountered H2S.  Although a number of instruments are available that measure toxic gas in the parts per million (ppm) range, there is a need to detect and measure hydrogen sulfide in the much lower parts per billion (ppb) concentrations due to odor complaints, which are traced to ppb levels.  The human odor threshold is 8 ppb and the human annoyance threshold is 40 ppb.[2]  To take control of this problem many state and local regulations limit the ppb release of hydrogen sulfide into the environment.

Wastewater treatment facilities are therefore required by law to measure and control an acceptable level of H2S exposure for their employees and surrounding civilians.  Protocols established to control and limit the odor of the H2S generated from decaying organics often include adjusting the pH, adding nitrates, or disinfecting the wastewater with chemical additives.[3]  The cost in minimizing hydrogen sulfide odor may be substantial and may place a heavy financial burden on the facility. [4] Understanding why these protocols are used for odor control as well as accurately and precisely measuring the hydrogen sulfide gas is important for ensuring safety while minimizing the use of expensive additives.  This paper briefly outlines how hydrogen sulfide is created in wastewater, how it is controlled and how you can accurately detect its presence using the Jerome® 605 Hydrogen Sulfide Analyzer (J605).  Using the J605 as an analytical tool may aid in determining the appropriate concentration of additives required to minimize H2S odor on a laboratory scale.  These empirical concentrations could then be extrapolated to full scale proportions saving the facility time, money and resources.

Jerome605

What are anaerobic bacteria and how do they contribute to H2S production?

An anaerobic bacterium is a term for a species of bacteria that is capable of functioning in an oxygen free environment.  Many of these bacteria have a distant evolutionary predecessor that thrived 3.5 billion years ago when the earth was mostly barren of oxygen but rich in sulfates.[5]  Like all forms of life, anaerobic bacteria break down sugars, lipids and proteins for energy but require some form of a chemical electron acceptor to complete its biochemical/metabolic cycle.  These sulfates (SO4-2) had become that electron acceptor (reduced) and in the process had created hydrogen sulfide gas by anaerobic fermentation.  Today, sulfate reducing bacteria such as Desulfovibrio, still play an important role in reducing sulfates in anaerobic conditions (i.e. wastewater).[5] Scanning electron micrograph shown below.

Why are anaerobic bacteria important in wastewater treatment?

Most decomposition bacteria can be categorized as chemoheterotrophic, meaning its energy source is derived from organic materials made by other organisms.[5]  Wastewater and grey water are rich with undigested organics and are the proverbial ‘all you can eat buffet’ for bacteria.  This is important in reducing the amount of solid organic waste found in wastewater.  Early in the bacterial decomposition process, aerobic (oxygen loving) bacteria dominate the feeding frenzy.  However as more and more aerobic bacteria grow and utilize the dissolved oxygen in the water, an acidic anaerobic environment is created.  This acidic, oxygen-depleted environment is now suitable for anaerobic bacteria to proliferate and consume the partially digested organic material left behind.[3]   Having an anaerobic counterpart ensures that decomposition is always occurring in any environment.

Why do pH levels, nitrates, and disinfectants affect H2S concentrations?

All forms of life function at an optimal pH.  If the pH is too high or low, proteins and lipids may denature and the organism may cease to function properly.  In the case of minimizing H2S odor, pH plays two roles.  By increasing the pH (or making the wastewater more alkaline), the anaerobic bacteria stop growing and metabolizing since their optimal pH has been altered.  Raising the pH also inhibits the release of dissolved H2S from solution, so it remains in an aqueous versus gaseous state.[4]

Nitrates (NO3-1) can also serve as a final electron acceptor in place of sulfates.  Flooding the wastewater with nitrates, forces the bacteria to utilize nitrate instead of sulfates which minimizes the H2S fermentation byproduct.[6]  Commercially available nitrates are usually in the form of liquid or powdered calcium or sodium nitrates.   Like pH, the addition of nitrates has both a biological and chemical effect on the water.  Preventative or curative attempts to minimize H2S often result in the chemical alteration of hydrogen sulfide back to ionic sulfate in the equation shown below:

H2S Nitrate reaction

The downside of adding too much nitrate is that it may cause problems down the line in purifying the water which is now spiked with excessive amounts of nitrogen.[6]  Preventative nitrate measures also tend to be a major cost in water treatment and when utilized, should only be done sparingly.

Disinfectants such as chlorine, ozone, or peroxides essentially break down the cell wall of any microbes present in the water.  These reactive compounds also combine with sulfur compounds and other hydrocarbons.[4]  These additives are usually reserved for the final step in processing wastewater but are essential for killing any harmful bacteria.  Like any other additives, these chemicals come at a cost and the addition of too much or too little have detrimental effects later in processing. 

How can you accurately monitor H2S?

Monitoring your H2S in your facilities may be as easy as walking around your work site with a hand held H2S analyzer such as the Jerome® 605.  This level of detection is important for ensuring the air quality is safe for all who may be exposed.  The J605 uses gold film technology specific for hydrogen sulfide gas with a detection limit as low as 3 ppb.  The J605 offers a continuous ‘Autosample’ mode option to monitor air quality over a period of time and saves the data which can be exported onto a spreadsheet for further analysis.

Arizona Instrument LLC (AZI) has performed several studies in which the J605 has been used not only as a safety/odor monitoring tool but as an analytical instrument in the laboratory.  Due to the volatility of hydrogen sulfide (b.p. = -60°C), one effective method for testing the concentration of dissolved H2S is to test the headspace above a liquid sample.  If the temperature and the accumulation time are well controlled, then the concentration of H2S in the headspace will be proportional to the concentration of H2S dissolved in the sample.  AZI has done extensive studies with dissolved H2S in liquid samples such as beer and wine.  A similar method can be used to detect dissolved H2S in wastewater.  The experimental design is shown below.

H2S in Beer & Wine

To run a test, an Erlenmeyer vacuum flask is connected to a Jerome® J605 Hydrogen Sulfide Analyzer by Tygon® or other suitably sized inert tubing. A liquid sample of wine or beer is poured into an Erlenmeyer flask and allowed to stir for 5 minutes. The instrument is placed in ‘Autorange’ and ‘Autosample’, sampling the headspace above the liquid sample every 2 minutes.  The instrument is then allowed to sample for 30 minutes, and the results are then summed for a total concentration of H2S.[7]

Although this particular study relates to alcoholic beverages, anaerobic fermentation still plays a key role in both processes where hydrogen sulfide gas is generated as an unwanted byproduct.  By taking in-line samples of wastewater during various stages of processing it is theoretically possible to maintain an accurate measure of hydrogen sulfide concentration on a laboratory scale.  Moreover, scaling down water treatment (addition of base, nitrates, and disinfectants) may give you better insight on quantity, the rate of reaction, and the efficiency of each of these additives may be (or any combination).

Understanding the chemistry of hydrogen sulfide and the biology of its generation is important in treating wastewater odor control.  Moreover, having a reliable and accurate instrument to measure H2S is paramount for both safety monitoring and analytical measurement. The Jerome® 605 has been shown to be a useful and versatile instrument in determining hydrogen sulfide concentrations in both air and liquid samples.  This technique may allow for more efficient bulk usage and purchasing for these odor inhibiting additives while maintaining a safe and healthy work environment in accordance with the law.

References:

[1] “Hydrogen Sulfide – PubChem Public Chemical Database”. The PubChem Project. USA: National Center for Biotechnology Information.

[2] “Hydrogen sulfide (also known as H2S, sewer gas, swamp gas, stink damp, and sour damp) Hole”. OSHA. 12 February 2013. Retrieved 22 May 2014.

[3] Mitchell, Ralph ed,. Water Pollution Microbiology, Wiley-Interscience, 1972.

[4] Palmer, Tony et. Al, Hydrogen Sulfide Control in Wastewater Collection Systems, Water & Wastes Digest, December 28, 2000.

[5] Prescott LM, Harley JP, Klein DA (1996).  Microbiology (3rd ed.). Wm. C. Brown Publishers. Pp.130-131 ISBN 0-697-29390-4.

[6] V.L Mathiodakis et. Al, Addition of Nitrate for odor control in sewer networks: Laboratory and Field Experiments.  Global NEST Journal, Vol 8, No 1, pp 37-42, 2006 Copyright© 2006 Global NESTPrinted in Greece.

[7] Moore, James et. al.  H2S in Beer and Wine.  Arizona Instrument LLC Newsletter. http://www.azic.com/downloads/white_papers/Hydrogen%20Sulfide%20in%20Beer%20and%20Wine.pdf?BLOG=Newsletter+Blog+Post&TYPE=Hydrogen+Sulfide

 

Detecting and Measuring Microbial (Unicellular Yeast) Growth on Food Products using Loss-On-Drying Moisture Analysis

by Christopher Altamirano, James Moore

Introduction

In 1825 the human population was estimated to have exceeded the 1 billion mark; a true indication of how technology and innovation was allowing us to grow in number and with longevity.[1]  Farms were becoming more industrialized and methods for preserving food and preventing spoilage became paramount for transporting food over great distances.  By the early 20th century, methods such as canning, pasteurizing and vacuum sealing were being utilized to prolong food products until they reached their destination.[2]  However these preservation methods have proven to be imperfect with many consumers suffering from food-borne illness throughout the past century.[3]  Food-borne illnesses are caused by microbial contaminants that include a vast array of parasitic, fungal, bacterial and even viral agents.  When any one of these microbial agents is consumed, the agent will replicate quickly inside the body causing a number of medical ailments and in severe cases even death.[3]

Today, we have more stringent government oversight and continuous microbial testing to ensure food safety.  These tests include selective media culturing, various forms of microscopy (or spectroscopy), enzyme linked assays, and PCR (or genetic) screenings of food samples prior to being released to the general public.[4]  However, even with the multiple detection assays at our disposal, food-borne illnesses are still a public health risk.  Reports of bacterial contamination and food recalls of chicken meat, raw spinach and pet treats still resonate in our collective consciousness and have prompted food industries and government agencies to find better ways of detecting the presence of microbial contamination.

With the exception of viruses, most food-borne microbial agents have one common life process; they eat, they grow (and/or replicate), and they produce waste.  In fact, all living things do this using a process known as respiration. [5]  Although some microbes can do this in the absence of oxygen (known as anaerobic respiration or fermentation), aerobic respiration describes the process in which energy is released from complex organic molecules (i.e. sugars, fats or proteins) in the presence of oxygen.  When this occurs gaseous carbon dioxide and water vapor are released as by-products and the organism loses mass (or weight).  The loss of mass not only comes from water vapor being released but more so by the release of carbon from a solid state into a gaseous state in the form of CO2.[5]

Cellular Respiration Equation

To test this hypothesis, we employed the use of loss-on-drying moisture analyzer (Computrac® MAX® 4000XL) with an affixed humidity air pump to keep samples at a controlled humidity.  The Computrac® MAX® 4000XL was used due to its ability to maintain a stable temperature, give accurate weight loss readings over an extended period of time, allow for downloadable graphs and data, and its internal plumbing for gas flow (humidity vapor).  Although most types of food-borne pathogens consist of bacteria (e.g. Escherichia coli, Salmonella typhi-murium, Clostridium botulinum), we, at Arizona Instrument, chose to use a common Baker’s Yeast, Saccharomyces cerevisiae as our model organism.[3]  S. cerevisiae is a relatively fast growing (90 min. doubling time) unicellular yeast that is commonly used as a leavening agent in bread and pastries.[7]  It is also innocuous in nature and NOT a human pathogen, but shares a similar physiology to some pathogenic yeasts such as Candida albicans (an opportunistic human pathogen). [7]  In this experiment, we first used an in vitro approach to demonstrate proof-of-concept by measuring weight loss on yeast inoculated potato dextrose agar plates and later tested ‘real’ food (sliced bread) using similar methods.

Potato Dextrose Agar Plates

The outcome of this investigation will offer new insights on a novel loss-on-drying approach to bioactivity measurement for food industries who wish to expand their breadth of microbial testing.  This method does not intend to replace any single traditional detection assay, but hopes to add yet another checkpoint in the quality assurance of food safety.  Because Baker’s Yeast was specifically used in this study, this approach might be particularly useful in determining the viability of certain yeast cultures for bread or for brewing industries where traditional spectroscopy driven methods (optical density) may only tell of a yeast population density, not necessarily their carbon emission capacity.

Methods

Media Preparation and Yeast Culturing

Potato Dextrose Agar (PDA)

Potato dextrose agar (PDA) is a semi-solid media which is commonly used for fungal cultivation in the lab.  Agar is a seaweed extract that has consistency of a stiff gelatin mold, but is composed of cellulose in place of proteins (i.e. gelatin). [8]  Because protein is not used as scaffolding, it can be heat sterilized at 121° C and still congeal unlike gelatin which would denature at these temperatures.  It is important to note that the agar itself is not consumed when microbes are grown on it, it is simply scaffolding that retains the liquid broth that is mix in with it (in this case PDB).

Potato Dextrose Broth (PDB)

Potato dextrose broth (PDB) is created by stewing peeled potatoes and adding dextrose (which is a simple sugar) in the correct proportions to create a nutrient broth that can be consumed by fungi (including S. Cerevisiae).  Unicellular yeasts can grow in this liquid broth in a single cell suspension and is referred to as a liquid culture. PDA or PDB can be purchased through any major biomed distributor but we chose to make it ourselves using a recipe from ‘The mushroom cultivator’ (see reference page for more details)[9]

Pouring Agar Plates

For this experiment, aluminum deep-well plates (5 inch diameter) were used with the outer rim removed (traditional petri plates made from polystyrene would also work) and sterilized at 121° C for 30 minutes.  When the molten PDA mixture came out of the oven, it was allowed to cool inside a loosely capped 500 mL Erlenmeyer flask until it reached ~50° C.  Once at this cooler temperature, ~30 mL of molten PDA was directly poured into the sterile empty plates in a sterile hood where then each of the plates were covered with a heat sterilized aluminum ‘lid’ (AZI 4KXL waffle pans) to prevent contamination.  Once the PDA solidified, the lids and plates were sealed with strips of Parafilm® to prevent excessive moisture loss and stored in a refrigerator set at 4° C.

Liquid culture of S. Cerevisiae

A portion of the PDB was withheld to cultivate a growing stock from a 7 gram packet of freeze dried Fast Rising Instant Yeast S. Cerevisiae (Bakipan®).  100 mL of PDB and ~3.5 gram of freeze dried yeast was added to a heat sterilized 500 mL beaker and mixed with a sterile loop.  The culture was periodically agitated every 30 minutes for 2 hours and either used for the experiment or stored in a refrigerator set at 4° C for later cell culturing. [9]

Inoculating PDA plates with S. Cerevisiae

Once a robust culture of yeast had been created, a flamed loop was dipped into the liquid culture and spread evenly over all of the available surface area on each plate to create a ‘lawn’ of yeast.  3 ‘loop fulls’ were used on each plate in order to supply a sufficient amount of starting population of yeast.  All inoculated plates were then inverted to prevent condensation from the lid from falling onto the lawn and disrupting cellular growth (common practice). [9] See diagram below for a visual representation.

PDA Plates

Inoculating Bread slices with S. Cerevisiae

The bread slices used for this experiment were from a Nature’s Own Honey Wheat® loaf.  The sliced bread loaf was stored in its original packaging in a refrigerator set at 4° C, but was taken out one hour prior to the experiment to allow the whole loaf to warm up to room temperature.  For the control, a slice of bread was placed onto a waffle pan and 5 drops of sterile PDB was placed onto the bread slice (4 drops being placed on the 4 corners of the bread and 1 in the center using a sterile transfer pipette).  For the experimental, the same drop placement was performed on a slice of bread but with a homogenous mixture of liquid yeast culture suspended in PDB.  (See diagram below drop placement)

Bread Slices

 

Computrac® MAX® 4000XL Programing and Procedure

Programing

In order to track weight loss in real-time, twelve 2-hour tests had to be linked to provide an accurate and continuous depiction of weight loss over a 24 hour period.  Be sure to connect an Ethernet cable to the appropriate port in the back of the instrument and enable the ‘Web Server’.  Enabling the webserver allows the analyst to download the raw data onto a Microsoft® Excel® Spreadsheet at a later point in time for data analysis.  Once you enable the optional ‘Web Server’, feature, turn the instrument on and off again to allow the instrument to recognize the new connection.  To observe the newly established instrument IP address, select ‘Menu’ à ‘Set Up Menu’à ‘Ethernet Set Up’.  Record this IP address for later data analysis (see manual for further details).

Because S. Cerevisae is not a true human pathogen, 37° C (the temperature of human body) is too warm for optimal yeast growth; therefore the temperature setting was dropped to 30° C. [7]  The purpose of using a moisture analyzer was to use it as an incubator to grow the yeast at optimal temperature and measure the weight loss over a fixed amount of time.  Test parameters for each individual 2-hour test are listed below:

Sample Size: 25.0 +/- 15.0 g

Idle Temperature: 30° C

Testing Temperature: 30° C

Pan Tare: Standard

Sample Tare: Standard

Ending Criteria: Time, 120 Minutes (2 hour maximum limit per run)

Link Test: Yes

Because each individual test will automatically read as an individual ‘% Moisture’ result, a custom equation was created to track the total percent of weight Loss.  The first 2-hour link does not require this custom equation, but the following links will in order to give an accurate real-time reading of weight loss.

Custom Equation:

((WI1-WF)/WI1)x100

With WI1 being the initial weight of the first test in the series and WF being the final weight of the current test.

Computrac® Dry Air Generator (DAG) Humidity Flow

Most food products have a substantial amount of water and because of that have a unique evaporation profile dependent on sample size, distribution, surface area and the environment it is in (humid or dry).  To minimize the evaporation for both agar plates and bread slices, a humidifier was created out of a repurposed Computrac® Dry Air Generator (DAG) with ambient air being bubbled through distilled water held in a 500 mL spouted Erlenmeyer flask, and carried past the sample chamber of the Computrac® MAX® 4000XL.  Without the modified DAG apparatus, the internal chamber of the instrument read at relative humidity of 10.6%.  After a sufficient amount of time (~30 minutes) of humid air being pumped through the chamber, the relative humidity read between 26.5% – 30.1% with the instrument holding temperature at 30°C. 

Originally, the MAX® 4000XL was designed for a nitrogen purge function in which highly volatile and potentially explosive samples could be tested in an oxygen free chamber (preventing combustion).  However, this set up was repurposed to provide a semi humid environment for the sample and for the yeast growing on it.  For further information on the Nitrogen Purge setup, consult the manual.  A diagram below shows the humidified MAX 4000XL connected by Tygon® tubing.

Tygon Tubing on the 4000XL

MAX® 4000XL Testing

Each test was started when the first link in the 12 linked series was selected and allowed to reach 30° C.  Once the temperature was attained, the analyst pushed start, the instrument tared the pan and prompted analyst to add sample within the prescribed sample window (10-40 g).  For the PDA samples (both control and inoculated alike), pans were uncovered from their original ‘lids’ and placed upside down onto the waffle pan/pan support on the 4KXL.  The instrument recognized the change in sample weight and was within the acceptable weight limits and urged analyst to close lid.  Once the lid was closed, the test began and continued for 24 hours.   

Data Analysis

Data was stored on the MAX® 4000XL instrument as well as continuously being reported to the network.  The IP address, determined at an earlier stage, was entered into the address bar of an Internet Explorer 10 Interface and the twelve linked tests were highlighted and ‘downloaded with graphs’.  This gives you the raw data in the form of an Excel® spreadsheet.  The raw data had to be reoriented by cutting and pasting into a new spreadsheet since the series was reported in the opposite order.  The ‘total % weight loss’ equation allows you to observe in real-time on the display screen but is not captured from the download; so a similar custom equation must be used in Excel® to determine the continuous % weight loss for each data point.  The equation used was as follows:

Picture1

The MAX®4000XL reports the sample weight and the rate every ~ 30 seconds.  However when a new test begins, the instrument does not immediately recognize that it is a link test and the % weight loss and rate drops back to zero.  If the first 30 second data point was left in the series the graphs produced would have drastic dips every two hours upon the onset of each test and would affect the average and therefore was omitted. 

Graphing in Excel®

The graphs presented in the Results section of this this paper were created using standard a ‘Scatter Plot’ graph in Excel® taking the averaged ‘total % weight loss’ (calculated from the previous equation) from 3 individual runs and plotted against time (in hours).  The 4 dependent variables observed were:

1)      PDA (not inoculated) – Control

2)      PDA w/ yeast inoculation

3)      Bread slice with 5 drops sterile PDB – Control

4)      Bread slice with 5 drops liquid culture

The error bars represented in each graph corresponds to the standard deviation taken at each 2-hour interval among the 3 run of each variable.

Results

PDA Weight Loss

The first hurdle in our investigation was to support the theory that ‘bioactivity’ can be measured by weight loss.  It was understood from the beginning that whatever food product was placed in the chamber would lose weight from evaporation alone.  Therefore, we needed to get a baseline evaporation curve for the nutrient rich PDA plates specifically made as an optimal media for S. Cerevisiae.  The linear ‘blue’ line of the graph represented below corresponds to three separate PDA plates void of any S. Cerivisae.

Picture2

Once the baseline rate of evaporation (more accurately the change in total % weight loss) was determined, it was predicted that if a living organism (i.e. S. Cerevisae) began consuming the dextrose in the media, that carbon dioxide and water vapor would be produced and the agar plate would begin to lose weight at a faster rate that the control plate.  Although we cannot necessarily prove that carbon dioxide or water vapors are the culprits of this loss, we can still show that a greater percentage of weight loss occurred at ~8 hrs. into the incubation as compared to the control.  As time proceeds, a higher % weight loss is observed in the experimental group.  After a 24 incubation period, it was observed that ~ 6.88% weight loss was achieved by the control, whereas 11.21% weight loss was achieved by the inoculated experimental group; a difference of ~ 4.33%.  Both variables were each performed in triplicate and averaged. Error bars represent the standard deviation at every 2 hour interval.

Bread Weight Loss

After confirming that weight loss due to bioactivity can be measured under ideal conditions, we turned our focus to a simple yet practical food item; bread.  Because yeast is used in the process of bread making, bread inherently has nutrients that yeast can readily use.  As mentioned before, each food type has a different water loss profile, and must first be determined to set a baseline to compare against weight loss due to bioactivity (or respiration).  Because bread has high moisture content and has a greater surface area than an agar plate, the rate of evaporation proved to be much greater, but eventually plateaus.  With the PDA experiment, 3 ‘loop fulls’ of liquid yeast culture was used to inoculate the experimental group which does not add a substantial amount of moisture on to the PDA plate (certainly not 4.33%).  However for the bread slice experiment, a loop of liquid culture would not be a sufficient amount of starting culture considering the surface area of all of the pores of the bread.  In order to yield a higher number of yeast onto the bread, 5 drops of liquid culture suspended in PDB was added to the bread slice (this also gives the yeast ample moisture to grow in the start).  However, 5 drops of liquid culture certainly adds a great deal of moisture to the experimental group so to even the starting moisture levels between the control and experimental, 5 drops of sterile PDB was added to the control slice (no yeast).  Over a 24 hour incubation time, the control slice of bread (blue line) losses 26.76% from its initial starting weight while the Yeast inoculated bread slice (red line) losses 28.39%; a difference of 1.63%.  This is not as big of a difference in weight loss as compared to the PDA experiment, however considering how much moisture is lost by evaporation alone, we still see a statistically significant trend between the two groups by hour 14 in a 24 hour period. Both variables were each performed in triplicate and averaged. Error bars represent the standard deviation at every 2 hour interval.

Picture3

Conclusion

Incubation based assays for determining the presence of microbial contamination on food products is not a new technique in food safety screening.  In fact, selective media culturing and spectroscopy methods often rely on growing any number of potential forms of contamination in greater number so as to be visibly perceived or detected using light spectroscopy.  Using loss-of-weight as an indication for the microbial presence on food however is a novel approach but requires a baseline evaporation curve of each product in question.  This approach allows the analyst freedom from using expensive selective media or specialized vessels for optical  density (as in some spectroscopy methods).[4]  The food product in question can be directly placed into the chamber of the MAX® 4000XL without any processing or culturing techniques. Moreover, this approach allows the analyst to view the rate of growth (via loss of weight) in real time.  It is important to note that a 24 hour incubation period (held at 30° C) was selected due to the growth profile of S. Cerevisae (doubling rate of 90 minutes).  As mentioned before, S. Cerevisiae is not normally a human pathogen which tend to be bacterial in nature.  One prime example of a food-borne bacteria is Escherichia coli (E. coli).  Under ideal conditions (37° C), E. coli has a doubling rate of only 20 minutes and if bacterial contamination is suspected a shorter incubation time may be more useful for the analyst than a 24 hour period, however some preliminary work would be required by the analyst to determine this to be true. [6]

This loss-of-weight detection assay is not a complete assay for microbial detection, but offers another method in the process of microbial determination.  This approach assumes there is a high enough microbial load already present in the food to be measured under standard incubation conditions.   This loss-of-weight approach will only detect the presence of living organisms on the food in question, it will not differentiate between innocuous and pathogenic organisms.  However, if an organism is growing on a food product at 37° C it may be a good initial indication of the presence of a potential pathogen in which other detection assays can be utilized to elucidate.

Although the repurposed Computrac® Dry Air Generator was used as a humidifier, further investigation is required to determine if this apparatus is essential in applying moisture to a growing organism, but was used as a precautionary measure to resist excessive desiccation during testing.  This brings into light the possibility of other gasses being used to fill the growth chamber.  In this experiment humidified air was primarily utilized, but humidified nitrogen was also used in a separate experiment to observe a change in % weight loss under anaerobic conditions (void of oxygen).  However, after several tests, it was determined that the % weight loss of yeast on PDA under anaerobic conditions was not statistically different to the aerobic conditions (data not shown).  This may have been caused by the strain of yeast used in this experiment, since many commercial strains have been selected for their resiliency in anaerobic conditions (i.e. within a loaf of bread).  It is interesting to note that a mild ‘alcohol’ smell was detected by analysts after these anaerobic tests had been completed.  Although this is purely speculation, it is possible for the yeast to have undergone fermentation in which carbon dioxide and ethyl alcohol may have been released instead of carbon dioxide and water.

This data may be particularly useful for members within the baking and brewing industries.  Testing the viability of yeast using other methods besides cellular enumeration might be useful since the number or density of a culture is not necessarily an indication of a strong yeast culture.  Determining the rate of growth based on % weight loss of different yeast strains might help bakers or brew masters decide on the best strain to use for their products. 

Although there may be drawbacks to having a single plate incubator (MAX® 4000XL), the data collected from the growth of an organism offers great insight on how it is interacting with your food product.  Moreover, having a stable and controlled environment means that the lid never has to be opened once the test has begun, which prevents fluctuations in humidity and temperature.  If it is in the interest of analyst, it is even possible to link a final ‘sterilization link’ at the end of the 24-hour series (depending on the sample being tested).  The MAX® 4000XL can attain temperatures as high as 275° C, though a final link set to 121° C would be sufficient to sterilize most food products (NOTE: if an inverted agar plate is being tested, it would be important for the analyst to turn the plate right side up prior to running this test, in which case a linked option would not be recommended.) 

Overall, the Computrac® MAX® 4000XL proved to be useful in detecting the presence of microbial growth based on the loss-of-weight of respiration.  This method adds one more assay for detecting potential food-borne contaminants in food and gives us insight on how real-time growth occurs on solid and semi-solid food types.

References

  1. Kennelly, John J.  Feeding 9 billion People, Ninth Annual AIA Conference, Banff March 2013, University of Alberta link http://www.albertaagrologists.ca/files/documents/5662_John_Kennelly_Feeding_9_Billion_People.pdf
  2. Tauxe R.V. Emerging foodborne diseases: an evolving public heath challenge.  Emerg Infect Dis 1997: 3:425-34
  3. World Health Organization. “Chapter 2 Foodborne Hazards in Basic Food Safety for Health Workers <internet>” (PDF). Retrieved Dec 25 2013.
  4. American Public Health Association (1992) Compendium of Methods for the Microbiological Examination of Foods 3rd Edition APHA Inc. Washington DC.  Retrieved Dec 27, 2013
  5. Stryer, Lubert(1995).  Biochemistry (fourth ed.). New York – Basingstoke:W.H. Freeman and CompanyISBN 978-0716720096
  6. Fratamico PM and Bayles DO (editor). (2005). Foodborne Pathogens: Microbiology and Biology. Caister Academic Press ISBN 978-1-904455-00-4.
  7. T. Goekhout, V. Robert, ed. (2003).  Yeasts in Food: Beneficial and Detrimental aspects. Behr’s Verlag. P. 322 ISBN 978-3-86022-961-3.
  8. “Bacterial nutrition”. Microbiology Laboratories, University of Wisconsin. Retrieved Dec 27, 2013.
  9. Stamets, P. The Mushroom Cultivator: A Practical Guide to Growing Mushrooms at Home.  http://www.ehow.com/how_5872701_make-potato-dextrose-agar.html

Comparing Traditional Oven Methods to Rapid Loss-On-Drying Methods in Powders and Bulk Solids for Moisture Analysis

Christopher Altamirano, Lab Manager
Arizona Instrument LLC

Introduction

Powders and Bulk Solids is a broadly defined genre of materials from various industries that describe a material that can be milled or processed into a powder for bulk storage or use.  For the food industry this might include milled flours, flavors or additives while the plastic industry might include resin pellets or powdered polymers.  Before final processing, it is important for these materials to be within a certain moisture range to uphold internal and external quality standards.  The moisture level of these products may change while being stored in intermediate warehouses or from being shipped in bulk around the world.  Historically, traditional oven methods (standard or vacuum) have been used to monitor the moisture levels of these products with a high degree of accuracy.  However, using these methods have proven to be quite lengthy with test times ranging from 30 minutes to several hours or even days.  No matter what the industry, test times such as these may be too long for a practical business approach in such a fast-paced economy.

A rapid loss-on-drying (LOD) Instrument for bulk solid moisture analysis uses the same theory as standard oven methods:

  1. An empty sample pan is weighed and tared
  2. Sample is added and recorded as the initial starting weight
  3. Heat is applied to sample to evolve moisture (or other volatiles)
  4. Difference in sample weight is recorded and calculated as % Moisture using the equation

With a rapid LOD instrument, the balance and heat source are coupled together allowing the user to view real-time moisture curves and rate graphs as moisture is evolved.   In the integrated system, test times are substantially faster with the same level of accuracy one would expect from traditional oven methods.

Using the Computrac® MAX® 4000XL Moisture Analyzer, various powdered samples were run in tandem with traditional oven methods to determine moisture content.  Samples included flour, food flavorings and inorganic polymers ranging as high as 14% moisture to as low as 0.03% moisture.  The instrument is robust and designed for onsite analysis using small sample sizes (<40 g).  Along with being able to view real-time graphs during each test, data from each run can also be stored via Ethernet connection or USB memory stick for graphical analysis.

4K

 Method

Sample Prep – All samples tested were analyzed as they were received from the manufacturer.  Samples were stored in air tight Mason jars to prevent excessive desiccation during testing. 

Test Conditions – For flour samples AOAC 925.09 or 925.10 methods were used.  For all other samples a modified 925.09 or AOAC 925.45 were used.

Computrac® MAX® 4000XL – Flour Samples

Test Temperature:  150 °C – 170 °C
Results Display:  % Moisture (3 or 4 decimals)

Computrac® MAX® 4000XL – Food Powders

Test Temperature:  85 °C – 120 °C
Results Display:  % Moisture (3 or 4 decimals)

Computrac® MAX® 4000XL – Inorganic Polymers

Test Temperature:  75 °C – 160 °C
Results Display:  % Moisture (4 decimals)

For information regarding specific parameters for each sample, contact Arizona Instrument LLC for further assistance.

All samples were sifted evenly over waffle pan and run using parameters listed above.  Flour samples that were analyzed using these methods included All-Purpose, Pastry, Whole Wheat, Soy, Black Bean, and Pinto.  Food powders samples included Vanilla, Tomato, Blueberry, Parmesan, and 3 different cheese powders.  The last set of samples included inorganic powders named polymer 1, polymer 2 and powder coating.  All trade names and manufacturer names were withheld for this study.

Results

Figure 1 represents a typical graph of whole wheat flour that can be viewed during routine testing.  The blue line corresponds to the % Moisture while the red line corresponds to the Rate (% loss per minute).  The data was downloaded into Microsoft® Excel® and graphed using graphical analytical tools in Excel®.  Once a rate drops below a set ending rate the test will terminate. (time, prediction, temp then rate are also other ending options that can be used)

Picture1

Figure 2 shows a set of flour samples analyzed with their corresponding oven method.  All oven and MAX® 4000XL tests were performed in triplicate (x3).  Error bars represent the standard deviation (S.D.) of the individual data set.

Picture3

The array of flour samples demonstrate equivalency to their corresponding oven reference methods at various moisture levels.

 It is also important to test other material to demonstrate the versatility of the MAX® 4000XL.  Figure 3a &3b are a collection of food powder flavors that also demonstrate the MAX® 4000XL’s equivalency to the oven method as well as maintaining a tight standard deviation (error bars).  Figure 3b consists of three ‘cheese powder’ samples which highlight the resolution of the instrument measuring these slightly varying moisture levels from the same product.

Picture5

Table 3 represents the raw data from ‘Cheese 1’ to demonstrate the significant difference in test times.

Picture6

Figure 4 presents another variety of samples analyzed by both standard oven and rapid LOD methods.  This set of data corresponds to the inorganic powders used in the plastic/coating industries.  The data demonstrates that even at low moisture levels, accuracy and precision are not compromised on the MAX® 4000XL as seen by the error bars (± S.D.).

Picture7

Table 4 represents the raw data with statistics of figure 4.  The MAX® 4000XL (4K) has a smaller S.D. than the oven even at low moisture levels when run in triplicate.

Picture8

Conclusion

A rapid loss-on-drying (LOD) instrument such as the Computrac® MAX® 4000XL allows the technician to accurately and precisely measure % moisture at high and low levels in various powders.  Data presented in this paper demonstrate that test times are substantially shorter when compared to standard oven references, with virtually no prep work (no need to purge pans or allow for cool time).  Having a real-time graphing LOD unit, allows the technician to view the rate of loss as its occurring or digitally save it for later analysis.  This is helpful in determining the appropriate ending criteria and maximizing the efficiency of each test.  Using the MAX® 4000XL will save both time and money and is robust enough to withstand the stresses of a distribution or manufacturing floor.

Maintaining Your Instrument: Computrac® Vapor Pro® Series

By James Moore & Chris Altamirano

Arizona Instrument LLC

Adding a new piece of equipment or specialized instrumentation to your facility has the potential to add significant value to your product.  Whether it be in the quality assurance or manufacturing process, adding a new instrument may save the company time, money, and increase the overall quality of your product.  However, adding a new method of analysis of your product requires learning new testing procedures and developing new testing parameters.  This learning curve may be the primary focus of the operator, but other tasks are important to keep in mind while adjusting to your new instrument. Tasks such as general maintenance and calibration checks should be practiced regularly to ensure the accuracy of your instrument as well as preserving the longevity for years to come.

Routine Maintenance

For operators using the moisture specific Vapor Pro ® line of instruments, routine purging of the internal tubing is recommended.  When heating samples on a daily basis within the Vapor Pro®, there is a good chance that volatilized/re-condensed material will remain deposited on the sensor of the instrument.   Making sure the sensor and tubing is void of any residue ensures accurate results.  This is completed by filling a sample vial with isopropyl alcohol (IPA) and manually moving the bottle transport in to the oven chamber.  Since IPA has a relatively low vapor pressure, it is recommended that the temperature of the instrument be set below 80 °C.  Once the transport has moved into the oven, the IPA will be pushed through the flow path allowing the fluid to flush the sensor as it passes.  Be sure the instrument is fitted with brass outlet port fitting to allow for proper drainage.  It is helpful to place an empty vial under the outlet port fitting to catch the outflow during the purge.  Leave the clean IPA vial in the instrument for approximately 30 seconds then manually move the transport out.  Once this is complete the sensor will be saturated and it could take 15-20 minutes to return to proper testing conditions.  Arizona Instrument LLC recommends this be done a minimum of once a week or as needed (depending on the type of material).  For a sample that volatilizes more readily, this type of cleaning may be needed more frequently.

Instrument Verification

When utilizing the moisture specific Vapor Pro® line of instruments, the best check to verify the accuracy of your instrument is to run a ‘RH Sensor Cal’ test.  This requires NIST (National Institute of Standards and Technology) traceable 1000µg capillary and deionized water.  To run this test, select the preprogrammed ‘RH Sensor Cal’ test and allow the instrument to heat up to 215 °C.  Gently break the surface tension of the deionized water with the capillary and water will fill the tube by capillary action.  Gently wipe any excess water droplets with your finger (Avoid cloth or tissue) and place filled capillary into a sample vial.  Seal the vial and place horizontally into the transport and begin running the test.  The test should take no more than 4-6 minutes and should yield 1000 µg ± 50 µg.  If the test results are not within this range, run a second test.  If the results are still not acceptable, call our customer service department for troubleshooting details.

Following a maintenance and calibration schedule for your instruments may seem like an extra step in your busy day, but is well worth it in the long run.  This will optimize your instrument’s performance and give you peace of mind that your instrument is working properly and accurately.  This will reduce downtime, and ensure the quality of your product.

 

 

Maintaining Your Instrument: Computrac® MAX® Series

By James Moore & Chris Altamirano

Arizona Instrument LLC

Adding a new piece of equipment or specialized instrumentation to your facility has the potential to add significant value to your product. Whether it be in the quality assurance or manufacturing process, adding a new instrument may save the company time, money, and increase the overall quality of your product.  However, adding a new method of analysis of your product requires learning new testing procedures and developing new testing parameters.  This learning curve may be the primary focus of the operator, but other tasks are important to keep in mind while adjusting to your new instrument. Tasks such as general maintenance and calibration checks should be practiced regularly to ensure the accuracy of your instrument as well as preserving the longevity for years to come.

Routine Maintenance

During the course of testing, it’s inevitable that samples will be spilled and the instrument will get dirty.  If the operator is using a Loss-on-drying instrument such as the MAX® 4000XL or MAX® 5000XL then there is increased chance of spilling sample onto the instrument due to the open chamber available to the user.  Fortunately, these instruments are designed for this type of treatment.  However, continuous build-up of material can leave the instrument looking undesirable and if the material is hazardous, it may even become dangerous for the operator.  Personal health, instrument damage and accuracy of results could be negatively affected by this build up and is therefore recommended by Arizona Instrument LLC, that the operator uses a Shop-Vac® or other nozzle based vacuum cleaner between shifts.  If the material being tested is a liquid or a gel, then the instrument should be wiped down with a cleaning agent, such as Formula 409® between shifts.  If the material is hazardous then it should be cleaned immediately using the proper procedure described on the material MSDS.

When using the MAX® 5000XL as a loss on ignition application, it is quite common for soot to build up within the test chamber.  If a significant amount of soot is observed, be sure to utilize the ‘Self Oven Cleaning’ setting.  In order to use this setting, first select ‘Self Oven Cleaning’ setting under the Calibration menu. The message on the screen will prompt the operator to remove the pan and pan support prior to beginning the program.  As soon as the pan and pan support are carefully removed from the chamber, hit start and allow the instrument to run for 45 minutes (instrument begins countdown automatically).  The instrument maintains a temperature of 550° C during this time,  which allows any residue to bake off the chamber walls.  Self-oven cleaning is recommended as needed for samples that regularly produce a full bodied plume of smoke or thick residue within the chamber.

Instrument Verification

No matter what instrument being used or the material type being tested, it is a good idea to make sure the instrument is working properly.  For the MAX® 4000XL and MAX® 5000XL, this can be performed by using a preprogrammed test labeled as ‘Weight Test’.  For this verification, the operator uses a 5g and 3 g weight.  Once the test has started, the operator places both weights on the balance as prompted and closes the lid.  The instrument will calculate the initial weight (8 g) and then begin testing.  After the test has begun, open the lid and remove the 3g weight.  Removing 3 grams from a total of 8 grams (3g + 5g) leaves you with a simulated loss on drying event of 37.50% ± 0.02%.  This is a quick verification that your instrument is measuring weight loss accurately and should ideally be performed daily to ensure accurate results.

Following a maintenance and calibration schedule for your instruments may seem like an extra step in your busy day, but is well worth it in the long run.  This will optimize your instrument’s performance and give you peace of mind that your instrument is working properly and accurately.  This will reduce downtime, and ensure the quality of your product.

Selecting the right sample size for Rapid Loss-On-Drying tests

When characterizing a material using rapid loss-on-drying technology, one of the vital parameters for accurate analysis is sample size. This is often very difficult to do because each material has unique characteristics that may influence how it should be tested. These are just some of the questions that need to be answered when determining the optimal sample size for a rapid loss-on-drying test:

Total Volatile Content – How much material is going to come off during a test? Is it 20%? Is it 2%? Is it 200ppm? The answer to this question will greatly influence the amount of material needed for an accurate test with an acceptable test time. For materials that expect losses greater than 20% only a few grams of material are needed. 5.0g ± 0.5g is a good initial testing sample size for materials with higher volatility. Once the initial analyses is finished the mass can be adjusted up or down to increase repeatability or reduce testing time, which is dependent on what condition is more desired by the user.
For materials that have a small amount of volatile content a larger sample size is needed. For materials with losses between 1 and 20% a good initial sample size is 10.0g ± 1.0g, and for materials with an expected loss less than 1% a samples size of 20.0g ± 2.0g is recommended. From these sizes the sample size can be adjusted for optimum instrument performance.

Sample Layering – Another factor to consider when selecting a sample size is sample layering. If the initial sample size selected is too large it could lead to sample material piling up. This causes the sample near the pan to be insulated from the heat source, which would cause a lower than expected result. Often times technicians attempt to compensate for this by increasing the temperature, which often leads to burning the top layer of sample material (similar to scorching the outer layer of food in an oven that is set too hot). If this is the case, try returning the testing conditions to a lower temperature and reducing the sample size while increasing surface area.

Flammability – When working with materials that can ignite, the amount of sample is a critical parameter of testing. Too much material could damage the instrument if it ignites, but too little material can hinder good repeatability from test to test. For samples that fall into this category it is best to start with 1.0g ± 0.2g of material, and increase the sample size in 0.5g increments until good testing conditions are achieved.

There are certainly other criteria that can influence the amount of sample needed, and each material will provide its own challenges for determining the optimal sample size.

Jerome J505 Portable Atomic Fluorescence Spectroscopy Mercury Vapor Analyzer

James Moore, Research Chemist
Arizona Instrument LLC

Introduction
Whether it is fluorescent lighting, dental fillings, antique switches, gold mining or thermometers, the element mercury (Hg) is present in the world we live.  Many of the mercury containing products give us comfort, are used to provide us with information, and even allow us to control our environment.  While these products are safe, they could potentially expose people to a plethora of toxic compounds if an accident should occur.  Symptoms of mercury exposure include seizures, memory loss, and in some cases, death.  Because of these risks, several guidelines and regulations have been developed that limit the amount of mercury people can be exposed to, and special methods are required for cleaning up mercury if an accident should occur.  Currently the time weighted average limit for mercury varies depending on regulating agency.  For OSHA, the limit is 0.1 mg/m3*; NIOSH sets the limit at 0.05 mg/m3*; and the ACGIH has a limit of 0.025 mg/m3*.  Since mercury vapor is not something people can see, how do they determine their amount of exposure?  Arizona Instrument LLC manufactures the Jerome® J505 Atomic Fluorescence Analyzer; a handheld atomic fluorescence spectrophotometer that measures the concentration of mercury in air.  The lower detection limit of this instrument is 50 ng/m3 (0.000050 mg/m3), and it can detect as high as 0.5mg/m3.  These detection limits exceed the current industrial exposure limits, as well as clean-up levels for public facilities.

Atomic Fluorescence Spectroscopy (AFS)
When an atom is excited by an input of energy, one of its electrons transitions from a stable ground state to an unstable excited state.  Once the source of energy is removed, the electron returns to its ground state and the absorbed energy is emitted as a photon (light).  This process is called fluorescence.  Often the amount of energy given off is not the same as the energy going in.  This is not the case for mercury, which makes it special.  When the energy required to excite an electron is the same energy as the photon it gives off when it returns to its ground state, it is called resonance fluorescence, and is easily detectable using Atomic Fluorescence Spectroscopy (AFS).  The Jerome® J505 instrument uses a mercury lamp to excite the mercury atoms at the 254nm wavelength, and then uses a detector to measure the emission of the photons, at the same wavelength, as the electrons return to their stable ground states.  Because AFS measures the emission of photons, this technique does not have interferences, such as hydrocarbons, hydrogen sulfide, and ammonia, which are often problematic for traditional detection methods.

The specifications for the J505 Atomic Fluorescence Analyzer are below.

Atomic Fluorescence Spectroscopy should not be confused with Atomic Absorption Spectroscopy (AAS).  In AAS, a light source of known wavelength and intensity is passed through a sample of interest. Some of the energy of the source light is absorbed by the sample as it energizes electrons in the material from the ground state to an excited state. A detector is placed at the end of the pathway to determine how much of the energy passed through. The difference between the energy of the source light and the energy of the light that arrives at the detector is directly proportional to the concentration of analyte in the sample.  One of the drawbacks of this technique is that there are a number of other common molecules that can absorb energy at the same wavelength as mercury.  To compensate for these unwanted absorptions, manufacturers use a variety of filtering techniques to limit background interference. While these filtration principles are sound, they come at the cost of a more complicated and bulkier instrument. Further, AAS can also have physical limitations that may limit low level sensitivity. At very low concentrations, the amount of absorbed light, when compared to the intensity of the incident light source, can become indistinguishable from electronic noise, making detection at these levels more challenging.

* These TWA averages are dependent on time.  For more information on exposure limits please visit each respective website.  

J505 Specifications

Test Mode, Standard, Quick

 

Jerome J505 Specifications

Search, Typical Test Time, Power Requirements, Operating Environment, Dimensions, Weight, Display, Unattended Autosample, Data Storage Capacity, USB, Certifications

Accuracy and Precision (Standard mode)

Gas Level, Accuracy, Precision (RSD)

 

ARIZONA INSTRUMENT LLC
3375 N Delaware Street | Chandler, AZ 85225
Tel (800) 528-7411 | Fax (602) 281-1745
www.azic.com | sales@azic.com

Testing for Hydrogen Sulfide at a Wastewater Treatment Facility Using the Jerome J605

James Moore, Research Chemist
Arizona Instrument LLC

Having never been to a wastewater treatment facility, I had some prejudices on what I would be exposed to while visiting the plant in Tolleson, AZ.

I was informed that the Operations Manager, David Tyler, was going to run some tests, and Garrett Rowe and I were welcome to join him. In accordance with their air quality testing protocol, when Maricopa County receives 3 odor complaints from nearby residents, the facility must test the air at the closest occupied location, which happens to be an elementary school on the corner of 91st Ave. and Lower Buckeye Rd. We checked in at the front office at 11:40 AM and discussed the local odors with some of the staff. From the discussion we learned that there
was a very pungent aroma in the area that smelled like burned materials. Since H2S has a distinct rotten egg smell we didn’t believe the odor they were describing was coming from the wastewater facility. We also learned that there is a meat packing plant and a dairy within 2 miles of the school, and these places may be
contributing to the unpleasant smells.

Once we left the office I took some deep breathes with my nose to try and characterize the odor of the air. There was a faint but distinct smell of
manure, but no rotten egg smell that would be expected with the presence of H2S. We sampled the air for 30 minutes in the parking lot using the J605 fitted with an ammonia scrubbing filter.

The instrument was set to survey mode and the location was named “school.” The J605 was in range 0 and did not detect any H2S during the testing period. We returned to the office of the elementary school and informed them that we
didn’t detect any dangerous gases.

In addition to the testing conducted at the school, the operations manager for the
wastewater facility also conducts a perimeter test, walking along the surrounding fence and measuring emissions in survey mode and the location set to “fence.”
During this testing the instrument read 0.00ppb for the vast majority of the fence‐line. The one exception was the area where the wastewater entered the facility.
There the instrument measured concentrations as high as 110ppb. As we left the high concentration area the readings returned to 0.00ppb. A few processing areas did have odors of ammonia and chlorine bleach, but no signal was produced.

Following the perimeter walk the facility was walked from end to end through the middle of the compound, starting in the north and heading to the south, and again from the east and heading west. During the north/south testing the instrument did not register a reading above 0.00ppb. As we traversed from east to west we passed through the sludge drying beds where solid waste is air dried before being shipped or sold. The material beds were approximately 30% filled, and smelled like plant fertilizer. No rotten egg smell was noticed and the instrument
did not provide a reading above 0.00ppb in this area. Once we reached the area where raw sewage entered the facility the instrument did again respond.

The final areas we tested were the air scrubbers they use at the exhaust of the sealed sewage processing station. There were 2 cylindrical scrubbers rising approximately 20 feet off of the ground with diameter of approximately 6 feet. The scrubbers were filled with activated carbon and were in year 3 of 5 before needing to be replaced. Mr. Tyler informed me that no prior testing had been conducted and these were the initial tests to ensure the scrubbers are maintaining efficiency. At the top of the scrubber where the air enters the atmosphere the instrument measured close to 8ppm H2S for scrubber 2 and 3ppm for scrubber 1. The gases coming into the scrubbers were measured at 1ppm, but there was a pressure gradient that would influence the results.

During general conversation the topic of solid content in sludge was brought up. David stated that they have the capability of testing solids in sludge, but don’t typically do this because the sludge is air dried for weeks prior to distribution
off‐site. However, many larger facilities do not have the ability to air dry their sludge for weeks and a rapid loss‐on‐drying instrument would be vital to monitoring mechanical drying processes. We also discussed testing Hg in water, and he
stated they do test for Hg, but sends water samples to an independent lab for metal
analysis.

Overall the Tolleson wastewater facility did a lot
to dispel some of the prejudices I was expecting
to encounter. Very few unpleasant smells I
believed would be there were present, and the
sludge material did not appear grotesque. In
fact, it looked like topsoil available at a garden
center. From a testing standpoint we were able
to begin
understanding the process of cleaning the water,
and how our instruments will best be used to
ensure that wastewater facility’s H2S emissions
are below prescribed levels. For our first visit
we were able to point out that the cleaning
scrubber could be monitored to ensure it is
working properly. Additionally solid content
determination would best be marketed to
facilities that are only doing mechanical drying
processes, and shipping material shortly after
drying. This would help them ensure their
dryers are working efficiently, and they aren’t
spending money shipping water. Hg testing
would be worth pursuing, but difficult as a
stand‐alone method, since they are interested in
other metal contaminant levels as well.

 

Arizona Instrument LLC
3375 N. Delaware St. | Chandler, AZ 85225
800-528-7411 | (f) 602-281-1745
sales@azic.com | www.azic.com

 

Moisture Determination of Specialty Resins Using Relative Humidity (RH) Sensing Technology; a Solvent-free Alternative to Karl Fischer Titration

James Moore, Research Chemist
Arizona Instrument LLC

Introduction

The Health Care industry has increased its needs for specialized devices over the past decade, which has led to a new frontier of resin and polymer development designed to keep the quality of care high while minimizing cost.  With these goals in mind, the resins being used for medical devices are scrutinized more thoroughly than other resins that require less regulatory compliance [1].  Analyzing a product for outgassing, deformation, and reactivity, among other things, has become part of the daily routine for manufacturers, molders, and final inspection personnel before an item can be shipped or used [2].  This additional testing and control also includes the amount of water that is allowed in the resins, since this will greatly influence the final product’s rigidness, consistency, and lifetime, as well as the quality of care that will be provided to the customer [3].  Oftentimes the quality control of the materials is closely monitored using testing equipment defined in an IQ/OQ/PQ: installation qualification (IQ), operational qualification (OQ), and performance qualification (PQ) to ensure that the instruments are effective, and the quality of the product is consistent.

Moisture Determination

As an alternative to the Coulometric Karl Fisher titration, Relative Humidity (RH) sensor moisture detection was first used as a method for determination of water in materials in 1997, with the introduction of the Computrac® 3000 Moisture Analyzer by Arizona Instrument LLC.  This method uses a thermoset polymer capacitor that has a selective response when in the presence of water, the same way that many RH sensors work in traditional settings such as houses, laboratory controlled environments, and dry boxes.

Medical device resins are sealed in a sample vial, and then transported into an oven chamber with inert gas blown through it.  As the material gets hot, water molecules evolve off and are carried to the sensor via the carrier gas.  The sensor is exposed to the water molecules and a measurable change in the electronic activity takes place. This method requires no solvents, making it an environmentally friendly alternative to traditional chemical titration.  The instrument provides in-situ moisture measurements, which allows users to monitor performance in real time.  Additionally, it has detection Limit of 10ppm, and is more rugged than Karl Fisher titrators, making it a suitable instrument for moisture analysis in manufacturing facilities, as well as Quality Control and inspection labs.

This technology is now being adopted as the standard test method and is described by ASTM D7191, Standard Test Method for Determination of Moisture in Plastics by Relative Humidity Sensor.  This instrument also meets the high demands of performance given in an IQ/OQ/PQ.

With major advances in technology, the medical device community is also taking advantage of new RAPID loss-on-drying methods for moisture determination.  These instruments use the same principle as traditional loss-on-drying techniques, but address the shortcomings of the method.

Sample material is heated on a balance and real time measurements are providing immediate feedback and moisture concentration.  The Computrac® MAX® 4000XL instrument, manufactured by Arizona Instrument LLC, provides a parameter development expert program that allows users to optimize testing conditions, such as sample size, test ending criteria, testing temperature, idle temperature, temperature rate, etc.  The chassis of this instrument is made of steel, which prevents cracking in the case and cool air from entering the testing chamber, which would influence the results.

These new techniques are being adopted as standard testing methods and are described by ASTM D6980 -12, Standard Test Method for Determination of Moisture in Plastics by Loss in Weight.  Like the Vapor Pro® 3100L, the MAX® 4000XL meets the performance standards set forth in typical IQ/OQ/PQ testing.

Testing

Sample Prep – Medical grade thermoplastic polyurethane (TPU), polycarbonate (PC), and nylon 6/6 resins were selected for analysis.  The materials were stored wet in a 1 gallon plastic Ziploc bags prior to testing.  An initial analysis of TPU was conducted to determine the water content prior to drying.  The material was then dried in the Dri-Air HP4-X 25 plastics drying hopper for 6 hours prior to testing.  The material remained in the dryer during testing due to the hygroscopic properties of the material.  Once it was determined that 6 hours was sufficient for testing, the PC and nylon 6/6 were also dried for 6 hours.  These materials were stored in Mason jars, upside down, to prevent head-space moisture from influencing the resultsTest Conditions – Reference testing was conducted using the Mitsubishi CA-100 Coulometric Karl Fischer titrator.

The parameters for TPU testing were:
sample size – 0.5g +/- 0.1g,
temperature – 90°C,
purge/preheat/cooling – 1/2/2,
ending sensitivity – 0.1µg/sec.
PC: sample size – 0.3g +/- 0.1g,
temperature – 220°C,
purge/preheat/cooling – 1/2/2,
ending sensitivity – 0.1µg/sec.
Nylon6/6: sample size – 0.5g +/- 0.1g,
temperature – 220°C,
purge/preheat/cooling – 1/2/2,
ending sensitivity – 0.1µg/sec.

Corollary testing was conducted using the Computrac® Vapor Pro® 3100L.

The parameters for TPU testing were:
sample size – 2g +/- 0.2g,
temperature – 105°C,
purge – 50 seconds,
ending criteria – rate<0.1µg/sec.
PC: sample size – 1.0g +/- 0.1g,
temperature – 180°C,
purge – 40 seconds,
ending criteria – rate<0.30µg/sec.
Nylon 6/6: sample size – 1.0g +/- 0.1g,
temperature – 240°C, purge – 30 seconds,
ending criteria – rate<0.05µg/sec.

Results

Comparative results of TPU, Lexan 1 and Polyone testing

Comparative results of TPU, Lexan 1 and Polyone testing

 

Total moisture curve of pre-dried TPU

Total moisture curve of pre-dried TPU

 

Total moisture curve of TPU dried for 6 hours at 200°F

Total moisture curve of TPU dried for 6 hours at 200°F

 

Real time rate of loss for dried PC material

Real time rate of loss for dried PC material

From the tables, the results using the two different instruments with similar testing conditions correlate to each other for all three medical grade materials.  For two of the three materials the Vapor Pro® did show a statistical improvement in the relative standard deviation, but did require a slightly longer test time than the Karl Fischer.  Additionally, the Vapor Pro® provided real time data points that could be used to graph the total moisture and rate change curves.  This allows for better monitoring of the product, or diagnosing possible problems with the instrument, and allows users to adjust testing parameters.  Examining the rate curve for PC, it was noticed that the rate had a significant drop at 0.30µg∙sec-1.  This is what was used to determine the ending criteria for this material.  This feature was not available for Karl Fischer titrator.

Conclusion

The development of an alternative to Karl Fischer moisture analyzer has been achieved, and can be used for moisture specific analysis of medical device grade resins.  The Computrac® Vapor Pro® 3100L moisture analyzer successfully uses Relative Humidity sensor technology for selective and accurate moisture measurement.  The instrument reduces the use of hazardous organic solvents makes it an environmentally friendly alternative, when compared to current Karl Fischer technology.   The results between the two methods of detection of H2O content for three separate resins, TPU, PC, and nylon 6/6 strongly correlate, with the Vapor Pro® 3100L providing real-time data that can be used to provide a complete moisture profile of the materials.

ARIZONA INSTRUMENT LLC
3375 N. Delaware St. | Chandler, AZ 85225
800-528-7411 | (f) 602-281-1745
sales@azic.com | www.azic.com

References

[1] DUPONT, “Medical Device Materials Provide
Design Options and Performance,”
4 December 2013. [Online]. Available: http://www2.dupont.com/
Medical_Device_Material/en_US/
products_services/plastics.html. [Accessed 15 January 2014].
[2] FAI Materials Testing, “Plastics and Polymer
Testing,” 30 July 2013. [Online]. Available:

http://www.faimaterialstesting.com/

plastics-testing.html?gclid=
CK2rsJjEgLwCFQmDfgodlwkAHg.
[Accessed 15 January 2013].

[3] M. P. Sepe, “Resin Drying and Moisture
Measurement,” in UBM, 2011.
[4] Paulson Plastics, “A Few Plastic Drying Tips For Injection Molders, Extruders And Other Processors,” Paulson Training Programs, 17 04 2013. [Online]. Available: http://www.paulsontraining.com/a-few-
plastic-drying-tips/. [Accessed 20 11 2013].
[5] Eastman Spectar, “Tips on Extrusion of Plastic Sheet,” 27 03 2013. [Online]. Available: http://www.eastman.com/Literature_Center
/D/DDS8.pdf. [Accessed 20 11 2013].
[6] ASTM, “Standard Test Method for Coulometric and Volumetric Determination of Moisture in Plastics Using the Karl Fischer Reaction
(the Reaction of Iodine with Water),” ASTM ,
pp. D 6869 – 03; 1-4, 2003.
[7] ASTM, “Standard Test Method for Determination of Moisture in Plastics by Relative Humidity Sensor,” ASTM International, pp. D7191-05; 1-4, 2005.
[8] J. Bozzelli, “Plastics Technology,” January 2011. [Online]. Available: http://www.ptonline.com/columns/you-must-dry-hygroscopic-resins. [Accessed 20 11 2013].
[9] E. M. M. I. J. R. W. J. Harold F. Giles Jr, Extrusion: The Definitive Processing Guide and Handbook, William Andrew, 2007.

 

The Water Factor in Medical Device Resins

THE WATER FACTOR IN MEDICAL DEVICE RESINS

James A. Moore
Research Chemist, Arizona Instrument LLC

Introduction

The Health Care industry has increased its needs for specialized devices over the past decade, which has led to a new frontier of resin and polymer development designed to keep the quality of care high while minimizing cost.  With these goals in mind, the resins being used for medical devices are scrutinized more thoroughly than other resins that require less regulatory compliance.  Analyzing a product for outgassing, deformation, and reactivity, among other things, has become part of the daily routine for manufacturers, molders, and final inspection personnel before an item can be shipped or used.  This additional testing and control also includes the amount of water that is allowed in the resins, since this will greatly influence the final product’s rigidness, consistency, and lifetime, as well as the quality of care that will be provided to the customer.  Oftentimes the quality control of the materials is closely monitored using testing equipment defined in an IQ/OQ/PQ: installation qualification (IQ), operational qualification (OQ), and performance qualification (PQ) to ensure that the instruments are effective, and the quality of the product is consistent.

Moisture Determination

As an alternative to the Coulometric Karl Fisher titration, Relative Humidity (RH) sensor moisture detection was first used as a method for determination of water in materials in 1997, with the introduction of the Computrac® 3000 Moisture Analyzer by Arizona Instrument LLC.  This method uses a thermoset polymer capacitor that has a selective response when in the presence of water, the same way that many RH sensors work in traditional settings such as houses, laboratory controlled environments, and dry boxes.

Medical device resins are sealed in a sample vial, and then transported into an oven chamber with inert gas blown through it.  As the material gets hot, water molecules evolve off and are carried to the sensor via the carrier gas.  The sensor is exposed to the water molecules and a measurable change in the electronic activity takes place. This method requires no solvents, making it an environmentally friendly alternative to traditional chemical titration.  The instrument provides in-situ moisture measurements, which allows users to monitor performance in real time.  Additionally, it has a lower detection limit of 10ppm, and is more rugged than Karl Fisher titrators, making it a suitable instrument for moisture analysis in manufacturing facilities, as well as Quality Control and inspection labs.  This technology is now being adopted as the standard test method and is described by ASTM D7191, Standard Test Method for Determination of Moisture in Plastics by Relative Humidity Sensor.  This instrument also meets the high demands of performance given in an IQ/OQ/PQ.

With major advances in technology, the medical device community is also taking advantage of new RAPID loss-on-drying methods for moisture determination.  These instruments use the same principle as traditional loss-on-drying techniques, but address the shortcomings of the method.  Sample material is heated on a balance and real time measurements are providing immediate feedback and moisture concentration.  The Computrac® MAX® 4000XL instrument, manufactured by Arizona Instrument LLC, provides a parameter development expert program that allows users to optimize testing conditions, such as sample size, test ending criteria, testing temperature, idle temperature, temperature rate, etc.  The chassis of this instrument is made of steel, which prevents cracking in the case and cool air from entering the testing chamber, which would influence the results.  These new techniques are being adopted as standard testing methods and are described by ASTM D6980-12, Standard Test Method for Determination of Moisture in Plastics by Loss in Weight.  Like the Vapor Pro® 3100L, the MAX® 4000XL meets the performance standards set forth in typical IQ/OQ/PQ testing.

Testing

Sample Prep – A medical grade TPU was selected for analysis.  The material was stored wet in a 1 gallon plastic Ziploc bag prior to testing.  An initial analysis was conducted to determine the water content prior to drying.  The material was then dried in the Dri-Air HP4-X 25 plastics drying hopper for 6 hours prior to testing.  The material remained in the dryer during testing due to the hygroscopic properties of the material.

Test Conditions – Reference testing was conducted using the Mitsubishi CA-100 Coulometric Karl Fischer titrator.  The parameters were: sample size – 0.5g +/- 0.1g, temperature – 90°C, purge/preheat/cooling – 1/2/2, ending sensitivity – 0.1µg/sec.

Corollary testing was conducted using the Computrac® Vapor Pro® 3100L.  The parameters were: sample size – 2g +/- 0.2g, temperature – 105°C, purge – 50 sec., ending criteria – rate<0.1µg/sec.

Results Water-Factor-Graph-1Water-Factor-Graph-2

From the table, the results using the two different instruments with similar testing conditions correlate to each other.  The Vapor Pro® did show an improvement in the relative standard deviation, but did require a slightly longer test time than the Karl Fischer.  Additionally, the Vapor Pro® provided real time data points that could be used to graph the total moisture curve.  This allows for better monitoring of the product, or diagnosing possible problems with the instrument.  This feature was not available for Karl Fischer titrator.

Conclusion

The development of an alternative to Karl Fischer moisture analyzer has been achieved, and can be used for moisture specific analysis of medical device grade resins.  The Computrac® Vapor Pro® 3100L moisture analyzer successfully uses Relative Humidity sensor technology for selective and accurate moisture measurement.  The instrument reduces the use of hazardous organic solvents makes it an environmentally friendly alternative, when compared to current Karl Fischer technology.   The results between the two methods of detection of H2O content in TPU strongly correlate, with the Vapor Pro® 3100L providing real-time data that can be used to provide a complete profile of the TPU.