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Analysis of Total Ergot Alkaloid Amount in Various Grain Samples by ELISA

By Dr. Frederik Wuppermann and Dr. Uwe Aulwurm

Introduction:

Ergot alkaloids are mycotoxins produced by fungi (Claviceps sp.) on various grain species with a highly toxic impact on the health of consumers (Figure 1). The toxins provoke symptoms summarized as ergotism.

ergot wheat

Figure 1: Sclerotia grown on crop

The toxins are characterized by two different isomers, which occur under different pH conditions, they are named –ines and –inines. The main toxins found are ergocristine, -inine, ergometrine, -inine, ergosine, -inine, ergocryptine, -inine, ergotamine, -inine, ergocornine, -inine. The assessment of toxin amount becomes necessary due to the toxic effects of ergot alkaloids. For this purpose an ELISA was developed to detect all ergot alkaloids in various crop material and the detected amounts of ergot alkaloids were correlated with LC-MS data.

 

 

Method

ergoread quick method
Figure 2.
ELISA procedure

The sample is extracted by 40/60 methanol / 0.25% phosphoric acid as described by Ware et al. 2000. After centrifugation the sample was diluted with sample dilution buffer and filtered again. The filtered sample was applied in the ELISA plate and compared with a standard solution (enclosed in the ErgoREAD kit). The enzyme immune reaction is performed as depicted in Figure 2.

 

 

Results:

Analysis of wheat samples:

analysis of wheat

 Analysis of Rye Samples:

analysis of rye

Comparative Analysis of Naturally Contaminated Samples:

The samples showed good correlation between 0 to 10 ppm, indicating that the ELISA is suitable for the screening of various crop material for the presence of ergot alkaloids. The partially over quantification in the comparative analysis could be due to the cross reactivity of the ELISA against various ergot alkaloids which were not monitored by the LC-MS or HPLC analysis. comparative analysis of naturally contaminated samples

Figure 3 cross reactivity of ErgoREADThe ELISA shows significant cross reactivity against various ergot alkaloids e.g. ergonovin (ergometrine or ergocristine) and can be used to determine the qualitative stuatus of a crop sample concernig the sum of ergot alkaloids.

 Conclusion:

The ErgoREAD ELISA is an appropriate tool for screening wheat, rye and triticale samples for presence of ergot alkaloids. The range of measurement is from 0 to 5 ppm. Due to the readout as the sum of all reactive ergot alkaloids, the cutoff for critical samples is 0.4 ppm. All shown ELISA samples are correlated with HPLC. The total amount of samples which need to undergo LC-MS analysis could be dramatically reduced.

More information about this kit and other Mycotoxin analysis products can be found on LCTech’s Website. The original poster can be downloaded here.

Pickering Laboratories is proud to bet the distributor for LCTech in North America. For more information about our long-time relationship with LCTech, check out our blog entry on Pickering’s Company & Product Offerings in 2013.

 

 

Pickering Laboratories in 2013

By Wendy Rasmussen

In recent years, we have released several new products and applications, and with still more on the horizon, it occurred to me that now would be a great time to summarize the Pickering of today – our mindset and our wide variety of products & applications.

Acai Berries
Acai Berries

No longer are we simply the “Post-Column Company”. We are the “Automated-Sample Antioxidants” company.  Think of us as the new “super fruit.” The Acai berry, or perhaps the new Chia Seeds (incidentally, we do have a post-column application for the identification & quantitation of  Antioxidants in a variety of matrices).

We are still very active, and we as a company plan to be here for many years to come. We are still the company founded on chemistry and a desire to to teach, to spread our technical expertise, to support our customers.

It’s been a few years now since Pickering began distributing and supporting our LCTech Product line.  The products have shown an ever increasing interest here in the US and in Canada (our official Sales Territory for this product line). We are very proud to offer these products and we hope we can develop it further in the future.

Historically, we have provided the back-end of an analysis (post-column derivatization). Nowadays, we can provide the front-end of analysis as well (the sample-cleanup).

In thinking about our product offerings, I realized that a simple list does not effectively show the scope of the products we have to offer in 2013 – primarily because we have a lot of overlap between products and product lines.  We’re not a vertical company in that regard.  I suppose one could say our product offerings are more circular in that many do not fall into a single distinct category.  I am a very visual person, and for me, a Venn diagram and our overall “product scale” really helped to understand and clarify our products:

Venn Diagram of Pickering Laboratories
Venn Diagram of Pickering Laboratories
General "Product Scale" for Pickering Offerings
General “Product Scale” for Pickering Offerings
For those of who like lists, you can find one Here, on Pickering’s website, and on LCTech’s Website

For any Questions, please feel free to contact us:

Pickering Laboratories, Inc.
Mountain View, California
Email: sales@pickeringlabs.com
Phone: (direct) 650-694-6700 or (toll-free) 800-654-3330

 

Image of Acai Berries:  http://acaiberryeducation.com/

Caltrans, the Blunder Lizard

By Michael Pickering

The largest known dinosaur was the Brontosaurus, literally the thunder lizard.  Its brain, estimated as the size of a fist, was too small to manage the whole beast. Apatozaur, Apatosaurus, Brontozaur, Brontosaurus, DinoAnimals.pl, Portal.DobreSciagi.pl Instead, it used a distributed intelligence in the form of neuro-bodies called ganglia.  Since they had only one neural system, the decision nodes were in constant, real-time communication.  This form of committee decision is the initial model for Caltrans management.  However, unlike the dinosaur, Caltrans adds an orthogonal system of decision node ganglia.  Not only is there a multiplicity of decision nodes within Caltrans, but other state agencies are nodes as well.  The dinosaur’s decisions were planar while Caltrans is a volume, both horizontal and vertical.

In the last century, during my early teens, perhaps middle or high school, I heard a Q/A joke:

Q:  What’s orange and sleeps four?

A:  A Caltrans van.

Caltrans was founded in 1895 as the Bureau of Highway.  Today it owns and operates ~15,000 miles of the California State Highway System.  Its annual budget is in excess of six billion dollars and it has more than 20,000 employees.

Because of the amount of public moneys involved and the scale of their projects, Caltrans regularly makes front-page news.  Unfortunately, it is always bad ink: delivery deadlines missed by years, budget overruns by many zeros.

In 1993, in order to address these common failures, Caltrans Director Van Loben Sels issued a charter to hold a peer review of the project management implementation plan.  The study group included Bechtel Corp., the U.S. Corps of Engineers, and the US Department of the Navy.  Some of their findings include the lack of:

  • realistic goals and objectives linked to civil service constraints;
  • communication, with specific roles and responsibilities not uniformly understood;
  • consistent management support with different district agendas; and
  • authority, with micromanagement by headquarters.

Similarly, in 1994 SRI International evaluated project management in response to Senate Concurrent Resolution No. 72.

bay bridge new span 2013
Eastern Span of Bay Bridge in 2013
Photo: Michael Macor, The SF Chronicle

The study found that Caltrans remains “rule driven” rather than “product driven” due to its longstanding bureaucratic culture.  SRI concluded that the Caltrans culture, not the organizational structure, was the culprit.  At the time, news analysts across the state described the audit as “scathing.”

So here we are in June of 2013.  Caltrans is front-page news because the replacement Bay Bridge span is years overdue and seriously over budget.  (Does anybody even remember that the reason for building the new span is that the old span is damaged and unsafe?).  The headlines are “Who Picked the Bad Bolts?”  The federal government is investigating California, the California legislature is investigating Caltrans, Caltrans is investigating vendors and other State agencies, and vendors are professing that all products were produced to the ordered specifications.  Although the bolts were made to spec (we hope; the jury is still out on this point), using them in this particular bridge design was inappropriate.

Eastern Span of Bay Bridge 2005
Photo: Wendy Rasmussen

The news reports of the multiplicity of investigations will drift off of the front page.  No little brain will be found, no ganglia identified.  Nothing will be revealed in real time.  The volume of decision nodes will hide all culpability.  The last report will be silence.

 

 

Editor’s Note:

The dinosaur which many of us know as Brontosaurus never really existed. The paleontologist who assembled the beast mistakenly placed the head of camarasaurus on the body of an apatosaurus. NPR has a fun story on this topic: http://www.npr.org/2012/12/09/166665795/forget-extinct-the-brontosaurus-never-even-existed

So does the UnMuseum  http://www.unmuseum.org/dinobront.htm

 

Images:

http://dinoanimals.pl/wp-content/uploads/2013/02/Apatozaur-Apatosaurus-Brontozaur-Brontosaurus-1.jpg

http://commons.wikimedia.org/wiki/File:Pasta-Brontosaurus.jpg

http://www.sfgate.com/bayarea/article/Broken-bolts-may-delay-Bay-Bridge-opening-4390299.php#photo-4385726

Chromatography Quiz No. 13

Chromatography Quiz #12 Results

We would like to congratulate our grand prize winners of our last newsletter’s Amino Acid Analysis Chromatography Quiz: Luiz Paulo Mousinho from Chemetric in Brazil, Helene Lachance from Shur-Gain Nutreco in Canada, and Narjes Ghafoori from the LA County Environmental Toxicology Laboratory!!!

They have each won and will be receiving: a $100 gift card from Amazon.com!  We would like to thank all of you for your submissions.

The correct answer for the modified Amino Acids chromatogram: the back pressure regulator on the outlet of the detector stopped functioning properly.  This was indeed another hardware troubleshooting quiz!  The pattern of noise seen in the customer’s baseline is characteristic to this problem, and a new back pressure regulator eliminated the noise.

Thank you!
Pickering Labs

Chromatography Quiz #13:
Identify the error made when running the Carbamate chromatogram below and win a prize!  Simply email your answer as well as your full contact information to Rebecca at rlsmith@pickeringlabs.com by August 30, 2013 in order to win.  You will receive email confirmation that your submission has been received.  The troubleshooting answer and winner congratulations will be published in the next issue (to be anonymous, please notify Rebecca in submission).

Carbamate Analysis for US EPA Method 531.1

Pickering Standard: 1700-0063 Carbamate Test Mixture, 2.5 µg/mL, 10 µL injection

Pickering Column: 1846250 Carbamate Column, C18, 4.6 x 250 mm

Normal Operating Conditions: (for reference only, condition changes may be reflected in chromatogram)

Column Temperature: 42 °C

Flow rate: 1 mL/min

Eluent Gradient:

TIME

WATER

MeOH %

0

85

15

1

85

15

44

25

75

44.1

0

100

49

0

100

49.1

85

15

57

85

15

 

Post-column conditions:

Reagent 1: Hydrolysis reagent CB130

Reagent 2: 100 mg of OPA, 2 g Thiofluor™ in 950 mL of CB910

Reactor 1: 100 °C, 0.5 mL

Reactor 2: ambient. 0.1 mL

Reagent flow rates: 0.3 mL/min

Detection: Fluorometer ex 330 nm, em 465 nm

Troubleshooting Chromatogram:

Troubleshooting chromatogram_CarbamateReference Chromatogram:

Reference chromatogram_Carbamate

Pickering Exhibits at Pittcon 2013

This year’s Pittcon was held in Philadelphia, Pennsylvania from March 17-21, 2013.

view of philly from the museum On display were Pinnacle PCX, EluVAC, and a video of the FREESTYLE Automated Sample Preparation Instrument.  The video was a new technique for us, and it was well received.

The video in our booth was a combination of the separate videos for each module of the FREESTYLE Automated Sample Preparation Instrument. You can view the originals here (they are very informative, and we highly recommend them): FREESTYLE VIDEOS

Wendy, Mike and David enjoyed visiting with existing and future customers, as well as our distributors from around the world, including our Canadian distributor, Chromatographic Specialties.

Before the show opened, we had the opportunity to explore the city.  Philadelphia is one of our oldest cities and is full of history and landmarks. Some of this editor’s favorites include Independence Hall and the surrounding neighborhoods, and the Art Museum (see more below).  On this trip, we explored Reading Terminal Market, and sampled that famous cuisine, the Philly Cheese steak Sandwich. I’m told it is not truly a Philly Steak sandwich unless you have it with Cheez Whiz. Any type of real cheese doesn’t count. I made that mistake on my first trip to Philadelphia, and the man behind the counter let me know it!  I wasn’t about to make that mistake again:

philly cheesesteak

The Philadelphia Museum of Art is known for its amazing collection of artwork (viewed on a previous visit), but is also made famous by the scene of Sylvester Stallone running up the stairs in the movie Rocky.  Since the author of this entry happened to be training for a race, she decided that a little cold wasn’t going to stop her. So she put on her iPod (Rocky Theme playing, of course), and ran from the hotel to the top of the steps and back (total distance: about 2.8miles).

view from the bottom

wendy philly top of steps

wendy rockyOn the return to the hotel, I came across a version of the LOVE sculpture, by American Artist Robert Indiana. I always thought it was was much bigger…

love sculpture

 

For those of you interested in learning more about the sculpture, click HERE

If you’re ever in Philadelphia, the Philadelphia Museum of Art is definitely worth a visit.

But if you can’t make the trip to Philadelphia, you could also stay home and watch Rocky…

Pickering Attends 4th Annual Latin American Pesticide Residue Workshop

By David Mazawa

The Fourth Latin American Pesticide Residue Workshop (LAPRW) was held from May 26 to 29, 2013 at the Compensar Convention Center in Bogotá, Colombia. About 400 people from universities, private companies, and government agencies responsible for monitoring and regulating pesticide residues in agricultural and environmental samples attended the valuable presentations, discussions, and workshops.Photo May 26, 17 53 47 About 19 different exhibitors, including Pickering Laboratories, proudly sponsored the 4th LAPRW. We would like to thank the Universidad Nacional de Colombia, Sociedad Colombiana de Ciencias Químicas and the Scientific Committee of the Latin American Pesticide Residue Workshop for putting together a great show. We look forward to the advancements of the Fifth Latin American Pesticide Photo May 25, 15 54 43Residue Workshop in Santiago, Chile.

http://laprw2013.com/ Photo May 25, 16 03 43 Photo May 25, 16 54 34 Photo May 25, 17 02 48

David and Laszlo
David.mazawa@pickeringlabs.com
Laszlo@pickeringlabs.com

Chromatographic Grade Water Recommended for Carbamate Analysis

By Maria Ofitserova & Mike Gottschalk

REh-body-image-2Purity of the mobile phase is an essential consideration in HPLC analysis. Solvents used to prepare mobile phase determine background noise level, presence of impurity peaks, as well as baseline drift during gradient formation. Use of highest purity solutions reduces undesirable baseline phenomena and improves overall sensitivity and selectivity of analysis.

EPA methods 531.1 and 531.2 require detection of carbamate pesticides in water on sub-ppb levels. To achieve such low detection limits it is crucial to have very low background noise and stable baseline without interfering peaks or drifts. Our customers long relied on Pickering Laboratories for highest quality reagents and chemicals. Now laboratories running carbamates analysis can purchase Pickering Laboratories Chromatographic GradeTM water tested using post-column analysis to be guaranteed free of fluorescence contaminations.

Guaranteed Chemistry