NMR Spectroscopy is the premier tool utilized by chemists to obtain detailed chemical information on molecular structure and is used extensively in molecular structure verification, chemical purity analysis, and complex mixture analysis. We have developed a quantitative NMR analysis that yields a chemical fingerprint that brewers can utilize to follow detailed variations in the chemistry observed in the various stages of the brewing process (malting, mashing, boiling, fermentation, ageing, and blending). The analysis observes all molecules in the beer at the same time and each molecular component (acids, alcohols, amino acids, malt-oligosaccharides) yields a unique spectral fingerprint pattern that is related to the structure of the molecule. Though the spectrum consists of a large number of overlapped individual fingerprints it is possible to identify and quantify individual components because many components have signals that appear at unique and specific points in the spectrum. The quantitative analysis is performed by comparing the area under the individual molecule signals to that of an internal standard (Maleic Acid 99%). Molecular components are quantified on a weight/volume basis in mg/L (parts-per million). Ethanol is also quantified on a %volume/volume basis.
The technique is not only applicable to the brewing process but is also being utilized to gain detailed chemical understanding of cider-making process, as well as the production of wine, mead, sake, spirits, and kombucha. Our laboratory has been developing this method with the help of a number of breweries following changes in batches of standard beers (Kolsch, Stout, Scots Ale, Barley Wine) as the brewing process is tweaked and changed over the course of 2 years. We have looked not just at finished beers but have studied dextrin solubility and chemistry of wort made from different malts, the effect of temperature on sour mashing, the effect of wild yeast and bacteria on various aspects of beer chemistry, as well as troubleshooting of “out of sensory target range” beers. The analysis requires very little sample preparation, has a large (orders of magnitude) linear concentration range of applicability and observes a large number of components in a single test that does not require constant re-calibration with expensive standards.
A poster was presented on this topic at the World Brewing Congress held August 1-17 in Denver Colorado – the poster can be downloaded here.
The Flyer for the PNA Beer Analysis can be found at this DropBox Link.
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Over the past few years our analytical NMR service has been developing a detailed chemical fingerprint analysis of alcoholic beverages by quantitative 1H NMR (qHNMR). Beyond the typical analyses of beer, wine, port, hard cider, sake and spirits, we have been looking at other fermented beverages such as kombucha, kefir, kvass, mead, ginger beer and perry. As well as the final fermented beverages we have been actively investigating the various starting materials such as malt wort, apple juice, honey, grape juice, fruit juices, and tea. The NMR analysis can provide a rapid quantitative analysis without any sample preparation based on the molar ratio of integration value of unique molecular fingerprint peaks with the integrated signal of an internal standard. In our case we typically use maleic acid as an internal standard as it’s singlet signal peak appears in a non-overlapping are of the spectrum to the chemistry we are interested in following.
The information that can be derived from the NMR experiment covers a wide dynamic range of component molecule concentrations from 10-100,000 ppm. The analysis observes all fully dissolved chemical constituents and the spectral response is linear with regard to all chemical types. As a primary analytical method the chemist can utilize the well understood literature on the NMR chemical shifts and couplings that allow first principles analysis of each molecular fingerprint to identify and quantify the presence of targeted and non-targeted molecules in the complex mixture. The analysis provides quantitative information on the following chemical components: ethanol, higher (C3,C4,C5) alcohols, methanol, glycerol, organic acids (lactic, acetic, succinic, pyruvic, pyruvic hydrate, citric, malic, tartaric, quinic), free amino acids (alanine, isoleucine, valine, tyrosine, phenylalanine), carbohydrates (sucrose, glucose, fructose, sorbitol, xylose, galacuronic acid, maltose, 1,6- and 1,4-dextrin chemistry, maltotriose, lactose), polyphenols. It can also provide information on yeast metabolism products such as 2,3-butandiol (directly from Enterobacter or from the action of saccharomyces on diacetal which is a well-known beer flavor deviation), 1,3-propandiol (from yeast action on glycerol after carbohydrates have been entirely fermented from the beverage).
In recent years kombucha has been found to contain more than 0.5% v/v ethanol which would technically lead the product to be classified as alcoholic beverages and bring the product under scrutiny and taxation by the Alcohol and Tobacco Tax and Trade Bureau which federally regulates the alcoholic beverage industry. Kombucha is a sweetened black or green tea that has been inoculated with a symbiotic culture of bacteria and yeast (SCOBY) which ferments the sugars in the drink solution in bith the manufacturing process and in the sealed bottle shipped out to stores. The drink is sold under the premise that the SCOBY provides a probiotic culture to the consumer which means that in many Kombuch products the activity of the culture is not arrested by pasteurization or by addition of sorbate. Thus, the kombucha is bottled with active yeast and bacteria present in a high sugar containing tea drink. Fermentation is then thought to occur while the product sits on shelves and leads to >0.5% ABV when the drink is purchased or consumed. We have utilized 1H NMR to obtain quantitative ethanol concentrations on a number of kombucha beverages bought off the shelf at grocery stores. The samples we analyzed represent the entire dataset of kombuchas that we purchased and they represent the products of 5 different manufacturers. We also aged two of the products at room temperature for 7 months and analyzed them to observe the effect of long term aging on kombucha products.
Experimental: 1H NMR spectra were acquired on a Varian Mercury-300MVX spectrometer operating at a resonance frequency of 299.67 MHz and equipped with a Varian 5mm ATB PFG probe. The experiments are performed under quantitative conditions utilizing a 10 ms (p/3 tip angle) pulse with an 8 second acquisition time and a 7 second relaxation delay. 64 transients were acquired over a spectral window of 8 kHz at a controlled temperature of 27oC. Water suppression was achieved by pre-saturation and this can affect the quantitation of glucose in the samples under these conditions.
Sample preparation: Samples were purchased “off the shelf” at local grocery stores and were analyzed the same day that they were purchased. Samples were prepared by 1) degassing the samples by repeated vortex agitation, 2) samples are equilibrated at 27oC before pipetting to allow a mass to volume conversion to be utilized to calculate the %ABV utilizing an ethanol density value of 0.7816 kg/L, 3) pipetting 175ml of kombucha beverage into a 5mm NMR tube, 4) adding 100ml of a 100mg/ml solution of maleic acid (99.5% – Sigma Aldrich) in D2O (99.8%D), and 5) addition of 375ml of D2O (99.8%D – Cambridge Isotopes Laboratories). The final samples were thoroughly mixed using a vortex mixer.
Two of the kombucha samples were purchased in duplicate and not opened immediately but stored at room temperature for 7 months before being analyzed. These stored samples were compared with the same samples that were opened and analyzed immediately after purchase.
Calculations: Component concentrations were calculated on a mg/L basis based on a knowledge of the concentration of maleic acid internal standard present in the sample (10mg) using the following equation:
Component Concentration (C) in mg/L = 0.995 x 10 x ((IC/NC)/(IMA/NMA)) x (MC/MMA) x (1,000,000/175)
Where 10 mg is the mass of maleic acid used as the internal standard, IC = integral of the component peak, NC = number of protons represented in the component peak, IMA = integral of maleic acid internal standard, NMA = number of protons represented in the maleic acid integral (2), Mc = molecular weight of the component, MMA = molecular weight of maleic acid (116.1 amu). Other aspects of the equation are – 175ml of sample must be adjusted to 1 liter (1,000,000 ml), and the whole must be multiplied by 0.995 as the maleic acid can only be guaranteed to be 99.5% pure. The ethanol content is calculated based on a weight per volume basis (mg/L) and then a calculation is performed to convert this weight/volume concentration to a volume/volume basis using a density value of 0.7816 kg/L to convert the weight of ethanol to the volume of ethanol.
Results: Figures 1-7 show the 1H NMR spectra of the 7 kombucha samples purchased and analyzed immediately. All 7 samples were found to contain ethanol and only one of them was found to contain less than 0.5%. Figure 8 shows a stacked plot comparison of the chemistry observed in a kombucha that was aged for 7 months at room temperature compared to the sample when it was initially purchased. The alcohol content rose from 1.23 %ABV to 4.25 %ABV and it can be seen that all sugars in the original drink have been consumed by the SCOBY to produce this increased alcohol content. The acetic acid content of the aged drinks also increased but it is obvious that the conversion of ethanol to acetic acid by acetobacter present in the SCOBY does not offset the overall production of ethanol. The component concentrations of ethanol, sugars and organic acids in each of the kombucha beverages analyzed are provided in Table I.
Table I: Concentration of Chemical Components of Kombucha Beverages
|Component||#1||#1 Aged||#2||#2 Aged||#3||#4||#5||#6||#7|
|Lactic Acid (mg/L)||64||68||131||210||461||124||1809||24||248|
|Succinic Acid (mg/L)||74||97||116||277||142||134||110||64||131|
|Acetic Acid (mg/L)||3056||5637||2746||3333||387||2806||2051||3719||444|
|Malic Acid (mg/L)||175||190||175||190||185||515||0||0||99|
Figure 1: Kombucha #1 – 1H NMR spectrum – component peaks utilized in calculations indicated.
Figure 2: Kombucha #2 – 1H NMR spectrum – component peaks utilized in calculations indicated.
Figure 3: Kombucha #3 – 1H NMR spectrum – component peaks utilized in calculations indicated.
Figure 4: Kombucha #4 – 1H NMR spectrum – component peaks utilized in calculations indicated.
Figure 5: Kombucha #5 – 1H NMR spectrum – component peaks utilized in calculations indicated.
Figure 6: Kombucha #6 – 1H NMR spectrum – component peaks utilized in calculations indicated.
Figure 7: Kombucha #7 – 1H NMR spectrum – component peaks utilized in calculations indicated.
Figure 8: Kombucha #2 – Comparison of original analyzed “fresh kombucha” with same purchase date bottle aged at room temperature for 7 months – 1H NMR spectrum – sugar peaks are consumed by the yeast to produce higher alcohol in the aged sample.
Alcohol in Kombucha News Articles:
http://www.cnn.com/2015/12/09/health/kombucha-tea-alcohol-content/index.html – visited 12-13-15
Kombucha Product Information:
Kombucha Brewers International – http://kombuchabrewers.org/ – visited 12-13-15
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1H qNMR Applied to Alcoholic and Non-Alcoholic Beverages
“Quantitative determination of ethanol in cider by 1H NMR spectrometry”, A Zuriarrain , J Zuriarrain , M Villar ,I Berregi, Food Control 50 (2015) 758-762
“Quantitative determination of lactic and acetic acids in cider by 1H NMR spectrometry”, A Zuriarrain, J Zuriarrain, A I Puertas, M Dueñas, I Berregi
“Identiﬁcation and quantiﬁcation of the main organic components of vinegars by high resolution 1H NMR” spectroscopy, A Caligiani , D Acquotti , G Palla , V Bocchi, Analytica Chimica Acta 585 (2007) 110–119
“NMR-based metabolomics in wine science”, Y-S Hong, Magn. Reson. Chem. 2011,49, S13–S21
“1H NMR-based metabolomic characterization during green tea (Camellia sinensis) fermentation”, J-E Lee , B-J Lee , J-O Chung , H-J Shin , S-J Lee , C-H Lee ⁎, Y-S Hong, Food Research International 44 (2011) 597–604
“NMR methods for beer characterization and quality control”, J E Rodrigues, A M Gil, Magn. Reson. Chem. 2011, 49, S37–S45
“Metabolomic proﬁling of Cheonggukjang during fermentation by 1H NMR spectrometry and principal components analysis”, H-K Choi, J-H Yoon , Y-S Kim , D Y Kwon, Process Biochemistry 42 (2007) 263–266
“Quantiﬁcation of organic acids in beer by nuclear magnetic resonance (NMR)-based methods”, J E A Rodrigues , G L Erny , A S Barros , V I Esteves , T Brandão , A A Ferreira , E Cabrita , A M Gil, Analytica Chimica Acta 674 (2010) 166–175
“Monitoring a commercial fermentation with proton nuclear magnetic resonance spectroscopy with the aid of chemometrics”, S Clark , N W Barnett , M Adams , I B Cook , G A Dyson , G Johnston, Analytica Chimica Acta 563 (2006) 338–345
“Quality control of beer using high-resolution nuclear magnetic resonance spectroscopy and multivariate analysis”, D W Lachenmeier, W Frank, E Humpfer, H Schafer, S Keller, M Mortter, Manfred Spraul, Eur. Food Res. Technol. (2005) 220:215–221
“1H NMR spectroscopy for proﬁling complex carbohydrate mixtures in non-fractionated beer”, B O. Petersen , M Nilsson , M Bøjstrup , O Hindsgaul , S Meier, Food Chemistry 150 (2014) 65–72.
“Regulatory Control of Energy Drinks Using 1H NMR Spectroscopy”, Y B Monakhova, T Kuballa, H Reusch, K Wegert G Winkler, D W Lachenmeier, Lebensmittelchemie 66, 129–168 (2012)
“Qualitative and Quantitative Control of Honeys Using NMR Spectroscopy and Chemometrics”, M Ohmenhaeuser, Y B Monakhova, T Kuballa, D W Lachenmeier, ISRN Analytical Chemistry Volume 2013, Article ID 825318, “Qualitative and Quantitative Control of Carbonated Cola Beverages Using 1H NMR Spectroscopy” P Maes, Y B Monakhova, and D W Lachenmeier, T Kuballa, H Reusch, J. Agric. Food Chem. 2012, 60, 2778−2784
A PDF Version of this Article can be found here – Kombucha NMR.pdf
Process NMR Associates quantitatively analyzes the component chemistry of craft beverages for consumers or manufacturers – for more information contact John Edwards at +1 (203) 241-0143
In conjunction with the 3rd PANIC conference in La Jolla, California, a 1-day NMR validation workshop was held that attracted approximately 80 interested participants. The agenda of the meeting is provided at this link (http://www.nmrvalidation.org/index.php/events/event-review) and registered participants can now download the presentations presented at the meeting. At the meeting it was decided to proceed with the idea of founding an organization dedicated to the development of validate NMR methods for use throughout all industry sectors.
NMR spectroscopy provides a means to evaluate material with high compound and high material specificity. Information as to the chemical structure, stereochemistry, quantity, material composition, and material identity is encoded in the NMR spectrum. The high reproducibility of NMR spectroscopy from instrument to instrument and lab to lab makes NMR an excellent tool for material validation. Approaches to utilizing NMR as a material validation tool include using (1) targeted approaches, the identification and quantification of specific components, and (2) non-targeted approaches, the use of chemometric methods to evaluate the spectrum as a whole. Efforts to increase the number and the speed of validated NMR methods are underway. This promises to move NMR technology from R&D to a mainstream analytical tool for production leading to high quality product assessment.
Quantitative NMR spectroscopy (qNMR) provides the most universally applicable form of direct purity determination without the need for reference materials of analytes or the calculation of response factors, with the only requirement being the exhibition of suitable NMR spectral properties. Due to recent advances in the technical development of NMR instruments, such as acquisition electronics and probe design, detection limits of components in liquid mixtures have been improved into the lower ppm range (approx. 5–10 ppm amount of substance).
The development of validated procedures and qualified standards will give users the tools to routinely exploit qNMR and enable them to speed up analytical method development, with the added advantage of reducing the time and financial burden of multiple analytical testing.
Over the last few years a number of efforts have been made to include NMR in routine testing and analysis – especially in regulated fields such as those operating under GMP or GLP guidelines. Unfortunately it has been observed that approval authorities, standard method organizations, and auditors prefer to take analytical routes derived from classical chromatographic methods. Since NMR represents a direct comparison analysis method such decisions clearly fail to take advantage of the benefits that NMR can provide.
The PANIC validation group proposes to become a driving force in getting NMR methods validated, publicized, and supported by documentation and qualified standards. The organization will also provide a mechanism for repeatability/reproducibility assessment of NMR methods as well as the round-robin accreditation of NMR labs. We aim to proactively promote the technology and improve its acceptance by the analytical community across all industry sectors.
What we want:
It is expected that there will be an annual 1-day meeting in conjunction with future PANIC conferences. A website has been been created as an organizational repository. The website can be found here: http://www.nmrvalidation.org/index.php and details of future events and, eventually, contact information will be provided.
1H quantitative NMR (qNMR) has been utilized to assess the the small molecule and carbohydrate chemistry of a number of home-brewed and commercial alcoholic ciders. A quantitative chemistry distribution of the products of the various fermentations that occur in cider making. Malolactic fermentation as well as fermentation by saccharomyces and wild yeasts occur in the cider making process which traditionally occurred without the intentional addition of yeast by the manufacturer. The distribution of small molecules produced by the yeast and bacterial metabolomes at work in the process can yield information of the sensory perception of ciders produced in different ways. An investigation of the residual sugar chemistry of commercial ciders gives some indication of the process of sweetening commercial cider products with sugar additions after fermentation is complete. These typical commercial ciders are very different in chemistry distribution compared to very dry cider styles such as those found in the Basque region of Spain where fermentation is taken to the extreme resulting in complete conversion of sugars to alcohol but also glycerols to 1,3 propandiol. Finally it was decided to determine how much quantitative chemistry information could be obtained from benchtop NMR systems operating in the 60 MHz range. These benchtop NMR systems have a price and cost-of-ownership that would allow small laboratories of manufacturers to think about their use in QA and QC roles.
John Edwards of Process NMR Associates will be presenting 4 papers at the 2015 ACS Northeast Regional Meeting that will be held in Ithaca, NY, June 10-13, 2015.
ABSTRACT ID: 2283171
ABSTRACT TITLE: 1H qNMR of Alcoholic Cider – Analysis of Small Molecule and Residual Sugar Chemistry (final paper number: 43)
SESSION: Food Chemistry
SESSION TIME: 5:00 PM – 9:00 PM
PRESENTATION FORMAT: Poster
DAY & TIME OF PRESENTATION: Wednesday, June, 10, 2015, 5:00 PM – 9:00 PM
ROOM & LOCATION: Emerson Suites – Campus Center
ABSTRACT ID: 2283063
ABSTRACT TITLE: Nutritional Supplement and Diesel Fuel Application Development for Benchtop NMR Systems Operating at 42, 60, and 80 MHz – Equivalency with Supercon NMR (final paper number: 336)
SESSION: Analytical Chemistry
SESSION TIME: 9:00 AM – 11:30 AM
PRESENTATION FORMAT: Oral
DAY & TIME OF PRESENTATION: Friday, June, 12, 2015 from 9:45 AM – 10:05 AM
ROOM & LOCATION: 222 – Williams Hall
ABSTRACT ID: 2283105
ABSTRACT TITLE: Survey of Low Field NMR Spectrometer Platforms for Successful Screening of Sexual Enhancement and Weight Loss Supplements for Adulteration with Drugs and Drug Analogs (final paper number: 386)
SESSION: Medicinal Chemistry
SESSION TIME: 1:00 PM – 3:20 PM
PRESENTATION FORMAT: Oral
DAY & TIME OF PRESENTATION: Friday, June, 12, 2015 from 2:20 PM – 2:40 PM
ROOM & LOCATION: 302 – Williams Hall
ABSTRACT ID: 2283153
ABSTRACT TITLE: From Mash to Bottle: Chemistry of the Beer Brewing Process and NMR-based Quality Control (final paper number: 284)
SESSION: Food Chemistry
SESSION TIME: 1:30 PM – 3:10 PM
PRESENTATION FORMAT: Oral
DAY & TIME OF PRESENTATION: Thursday, June, 11, 2015 from 1:35 PM – 1:55 PM
ROOM & LOCATION: 202 – Williams Hall
1H NMR shows excellent promise to be utilized in the quality control and authentication of essential oils. In order to ascertain if benchtop NMR systems reveal adequate “1H spectral fingerprints” for this purpose we have run several hundred essential oils at 300 MHz (Varian Mercury-300 MVX by 1H, 13C, COSY, HETCOR, DEPT) as well as at 82.3 MHz (Picospin 80), 60 MHz (Aspect-60), and 42.5 MHz (Magritek Spinsolve). The results plainly show that the spectrometers all yield similar proton line-widths with the difference in field strength leading to different levels of spectral dispersion and resolution. Though each spectrum is different it can plainly be seen that they all contain the same information with varying degrees of overlap. Chemometric and database comparative methods are being developed to allow identification of various essential oils as well as screening and quantifying different levels of adulteration. The figures below show examples from 6 different essential oils showing spectra obtained from all 4 spectrometers and plotted in the normalized chemical shift scale (ppm) as well as the absolute frequency scale (Hz).
1H NMR is an excellent tool for monitoring the residual olefin content of polymers after hydrogenation reactions. The fact that the olefin fall in a unique region of the spectrum means that it is a straightforward measurement to quantify the %H present as olefin or to correlate that olefin content with other analyses such as bromine number. Here is an example of a polyalphaolefin residual olefin analysis. The olefin proton content (%H) was plotted against bromine number values obtained on each of the samples. A linear correlation was obtain but two different correlations were observed that were dependent on the viscosity index of the polyalphaolefin being analyzed. Figure 1 shows the 1H NMR spectra obtained on neat samples on a Picospin-80 spectrometer operating at 82.3 MHz. The methyl and methlene protons of the polymer backbone are plainly seen and the olefin and alpha-olefin protons are observed.
Figure 2 shows the linear correlation between %H olefin and bromine number with the two correlations caused by different VI grade being indicated. The analysis shows that for the two viscosity grades the grade can be identified from the linear correlation that the data falls onto and the %H olefins content can directly yield the bromine number. This NMR method provides an alternative to the following ASTM standards: D1159 Test Method for Bromine Numbers of Petroleum Distillates and Commercial Aliphatic Olefins by Electrometric Titration D1491 Test Method for Test for Bromine Index of Aromatic Hydrocarbons by Potentiometric Titration D1492 Standard Test Method for Bromine Index of Aromatic Hydrocarbons by Coulometric Titration D2710 Test Method for Bromine Index of Petroleum Hydrocarbons by Electrometric Titration D5776 Test Method for Bromine Index of Aromatic Hydrocarbons by Electrometric Titration
Poster to be Presented at the 56th ENC, Asilomar CA, April 2015
|NMR Detection of Tomato Paste Spoilage in 1,000 L, Metal Lined Totes|
|Michele Martin1; Paul Giammatteo2; Michael McCarthy1; Matthew Augustine1|
|1University of California, Davis, Davis, California; 2Process NMR Associates, Danbury, CT|
Low field nuclear magnetic resonance (NMR) is used as a non-invasive method for detecting spoiled tomato paste. It is shown that the 1H T1 and T2 relaxation times change as tomato paste spoils due to changes in viscosity and/or changes in the concentration of paramagnetic compounds. With the goal of developing a spoilage detector that can be used in a tomato processing facility, a γBo = 19.5 MHz single-sided handheld NMR instrument is used. Due to the dominance of diffusion on relaxation measurements made with the single sided instrument, the slope of the amplitude of a spin echo for three different delay times is used to provide a viscosity dependent parameter that permits the differentiation between pristine and spoiled tomatoes.
One-Sided NMR – Non-Invasive Analysis of Tomato Paste
Back in October we presented a talk at Gulf Coast Conference that concerned the prediction of the chemical and physical properties of heavy petroleum feeds being converted to higher value product in a residual catalytic cracker (RCC). Over the years we have analyzed these materials by 300 and 60 MHz NMR and obtained good PLS-regression models that can adequately predict properties for real-time process control and optimization in a petroleum refinery. With the advent of a large number of new benchtop NMR systems we have been convincing ourselves that these types of analyses can be performed by systems such as the Magritek Spinsolve 43 MHz. We ran a series of samples that had been sitting around our lab for 15 years by dissolving them at about 50 volume% in a 50/50 CDCl3/CS2 solvent system. For each sample we had laboratory test data for a number of chemical and physical properties of interest to process engineers. We regressed the lab data variability against the variability in the Magritek 43MHz 1H NMR spectra and obtained cross-validated PLS models. The presentation material is given here at this link – Gulf Conference Presentation – 43 MHz RCC Feedstream Regression Models
PNA has been conducting a number of studies into the adulteration of male enhancement herbal suplements with PDE5 inhibitors such as viagra, cialis and a wide number of analogs (Mw approximately 475 amu). Standard materials for the analogs are quite expensive so in th eprocess of developing ID and purity methods on our NMR systems we ran the 1H NMR on our 300, 60, and 43 MHz NMR systems at a concentration of 5mg/ml which equates to 0.01 Molar. This PDF of the NMR data demonstrates the sensitivity and resolution obtainable on these bench-top 5mm NMR systems. They can readily look at samples at these concentrations. They can readily be utilized as screening tools or as quantitation and identification analyzers.
The PDF file containing the Varian 300, Aspect-60, and Magritek 43 MHz data can be found here: NMR Data-0.01 Molar_PDE5 Analogs 300_60_43_MHz
Here is an example of spectral reproducibility. We are doing a lot of beer NMR at the moment on our 300 MHz NMR and for “giggles” we are running many samples through the various bench-top systems in our lab. We have been quantifying small organic acids (lactic, acetic, succinic , malic, citric, etc.) as they can give some idea of yeast activities and health during fermentation. We are also quantifying and studying the 1,4/1,6 linkage distribution of residual dextrins. The series of superimposed spectra below consists of 28 spectra of a freeze dried beer sample (a unique Belgian Dubbel. Each spectrum was 128 pulses and took approximately 30 minutes per spectrum. So the superimposed data represents a 14 hour continuous stability test.The data was automatically processed with 16K zero-fill and autophase.It looks pretty damn good.
We’ve been looking at a lot of sour beers – here is a home-brewed Flemish Red aged in an oak barrel – note the high lactic and acetic content.
We’ve also been analyzing a lot of hard ciders – commercial and home-brewed varitieties of various styles – very different from one sample to another in the small molecule and sugar chemistry.
1H qNMR at 300MHz or 60 MHz can be utilized to identify and quantify small molecule chemistry in fermentations. Below is an example of a quantitative chemistry report on a series of ciders.
Beer and Cider Analysis is offered with similar quantitative results is offered for $100 per sample in our analytical lab.
Survey of Low Field NMR Spectrometer Platforms for Successful Screening of Sexual Enhancement and Weight Loss Supplements for Adulteration with Drugs and Drug Analogs
John C Edwards1, Kristie M Adams2, and Anton Bzhelyansky2
1Process NMR Associates, Danbury, CT
2United States Pharmacopeial Convention, Rockville, MD
The adulteration of dietary supplements (or natural health products) with synthetic pharmaceuticals is an area of increasing concern, which presents substantial risk to public health. Widely available in retail and via the Internet, these products are often marketed as sexual enhancement, weight loss and/or bodybuilding supplements. Unlike prescription drugs, supplements do not require premarket approval by FDA before they are made available for public consumption. In fact, the agency can only take investigational action after the adulterated product has caused harm and the adverse event has been reported via MedWatch (FDA’s online portal for voluntary reporting of adverse events associated with drugs, medical devices and dietary supplements).
Development of analytical tools for screening and identification of adulterated products in the marketplace represents a significant step forward in the fight against adulterated dietary supplements. Several organizations, including AOAC and USP, have undertaken initiatives to evaluate and recommend analytical methodologies for screening supplements for adulteration. HPLC and mass spectrometry have so far dominated the screening and quantitation studies published in the literature, with NMR spectroscopy often relegated to the status of structure elucidation tool. In this work, we investigate the ability of several-low field NMR spectrometric platforms to successfully identify and quantify the presence of adulterating drug substances in sexual enhancement and weight loss supplements purchased online and in US retail. 1H qNMR of both types of samples was performed with 300 MHz NMR to confirm the presence of adulterants such as sildenafil, tadalafil, and their structural analogues (sexual enhancement supplements) and various synthetic stimulants (weight loss supplements). We have concluded that a simple sample preparation protocol combined with straightforward 1H NMR spectroscopic analysis yields a rapid, robust and reliable screening test for adulterated supplements, presenting an attractive alternative to more labor-intensive, expensive and expertise-demanding techniques du jour.
This was presented by John Edwards at SMASH in September 2014, and at the Carolina NMR Symposium in November 2014
presentation can found here: Benchtop NMR – Herbal Supplement Adulteration Screening
1H Benchtop NMR has great potential to increase the throughput of both routine and emergency fuel sample analysis in refinery laboratories. Currently fuel samples must be passed through multiple dedicated analyzers to obtain information such as density, H-Content, aromatics, olefins, saturates, benzene,
Octane numbers, cetane index, cetane number, distillation curves, vapor pressure, flash point, pour point, freeze point, cloud point, etc. Correlation of the 1H NMR spectra of these refinery fuel samples to these primary test results will allow all parameters to be predicted in about 40 seconds from the 4 pulse spectrum of the pure fuel. Here we have a few examples obtained on some diesel fuels that were submitted to our lab for ASTM D7171 – Hydrogen Content by TD-NMR. We had density, H-content, and aromatics wt% by GC. Below are three example correlation obtained on the Picospin 80 system (that requires 32 pulses per sample due to the capillary sample size). The results were very similar for the 300, 60, and 42 MHz data obtained on the three other NMR system in our laboratory. The comparative results are shown in Table II. The results are very similar independent of the field strength of the NMR system. The data from all 4 NMR systems is provided in this section.
Benchtop high-resolution NMR systems are available at a number of field strengths and probe configurations. However beyond the obvious academic instruction market for these instruments very few applications have been demonstrated across all available platforms and thus proving the general applicability of benchtop NMR technology to industrial quality control. We will present two chemometric-based applications that have been developed at 4 different field strengths utilizing Varian Mercury 300 MHz, Magritek Spinsolve 42 MHz, Aspect AI 60 MHz, and Thermo Picospin 80 MHz NMR systems. Partial-least-squares (PLS) regression correlations were obtained on all 4 platforms relating to:
1) Omega-3 fatty acid composition of samples taken from various points in a nutritional supplement manufacturing process. Excellent correlations were obtained on all 4 NMR instruments proving that NMR technology is applicable to in-lab, at-line. or on-line analysis of fish oil derived omega-3 fatty acid supplements. The 40 second NMR analysis effectively replaces a 60+ minute GC analysis.
2) Physical and chemical property determination of diesel fuels where excellent correlations were obtained between 1H NMR variability and parameters such as density, aromatic content by GC, hydrogen content by 1H TD-NMR (ASTM D7171 method), and sulfur content. Many more physical and chemical properties can be correlated to the 1H NMR spectrum allowing a single 40 second NMR experiment to predict 10-15 parameters that each require dedicated analyzers.
Finally, we will present the concept and initial results from an independent server-based NMR application software that can be utilized in conjunction with the NMR software of the current benchtop NMR systems, or alternatively as a stand-alone application platform. This software would effectively make chemometric and direct measurement NMR application ubiquitous across all NMR platforms.
A link to this presentation in PDF form is given here: PLS-Regression – 300_80_60_43 MHz NMR of Fish Oil Supplements and Diesel Fuel
Adam DiCaprio (ex PNA) gave an excellent Science Cafe Talk under the auspices of the ACS North Carolina Section at the Busy Bee Cafe in downtown Raleigh on December 2, 2014. CHanging gears from his previous talks he centered the discussion on malt and hop chemistry as well as an start-to-finish NMR analysis of production runs of a bottled commercial tavern ale. If you are interested in having Adam give a detailed chemistry seminar on beer at your section meetings please contact him directly at email@example.com.
A PDF version of his talk is available here …. ACS Science Cafe Talk – Dicaprio – Busy Bee Cafe – Raleigh NC – 12-2-14
Process NMR Associates is currently developing NMR applications based on direct measurement of chemometric modeling on NMR data obtained on numerous NMR platforms. Our intention is to develop solutions that can be executed on any NMR platform. With this in mind we are currently developing a fish oil analysis application that can provide the EPA and DHA omega-3 fatty acid content of fish oil supplements manufactured by an ethyl ester esterification process. We have obtained data at 42 MHz, 60 MHz, 82 MHz and 300 MHz. The chemometric modelling yielded PLS models for all 4 field strengths that yield effectively the same result – DHA can be measured to +/- ~1.1 wt% and EPA can be measured to +/- ~2.2 wt% by a 40 second 1H NMR measurement. THe correlation is derived from a regression of the 1H NMR variability with primary GC analysis values.
This analysis has been shown for the 300 and 60 MHz data in a previous post on this blog. The same analysis was also obtained, with similar results, on 42 and 82 MHz platforms proving that individual applications can be automated and provided at all relevant frequencies of NMR analysis whether on superconducting lab systems or permanent magnet benchtop systems.
At each field strength the relative lineshapes are pretty much the same (<1 Hz at half height). The field strength differences mean that the same spectrum is dispersed across frequency space proportionate to the magnetic field. Figure 1 below shows the frequency space spectra obtained at all 4 field strengths on the same sample.
Figure 2 shows the same spectra displayed on the usual normalized chemical shift scale (ppm). In these spectra the data is stretched in order to allow the chemical shift comparison of the data. IN effect the 42 MHz NMR is stretched by a factor of 7, the 60 MHz data by a factor of 5 and the 80.2 MHz data by a factor of 3.7. The effect of the relative size of J coupling compared to the frequency space occupied by 1 ppm is an interesting observation to see directly. In traditional NMR analysis the resolution of various peaks was always the driving force for increasing the magnetic field strength of NMR instruments. With todays powerful PC’s and advanced software information can be garnished readily from any of these spectra by way of global spectral deconvolution or multivariate statistics.It is no longer necessary to obtain baseline resolution in order to integrate a resonance and obtain quantitative information.
THough the data is more closely packed together in the low field spectra it can be acknowledged that the same information is present in all 4 spectra. Automation approaches can be developed that will allow accurate measurement of quality parameters, component quantification, or structural verification to be performed on data obtained from any of these NMR systems.
The development of readily deployable NMR benchtop systems at an affordable price point must surely lead to the development of NMR into a more widely utilized technique outside of the realms of scientific study and quality control that NMR has thus far been involved.
Poster presented by Adam Dicapro at the 3rd Carolina NMR Symposium, Kannapolis, NC, November 6, 2014
Agilent Technologies to Close Nuclear Magnetic Resonance Business
SANTA CLARA, Calif., Oct. 14, 2014
Agilent Technologies Inc. (NYSE: A) today announced it is exiting its Nuclear Magnetic Resonance business. Agilent entered the NMR business in 2010, with the acquisition of Varian. Since then, the business has not met growth and profitability objectives.
“Today’s announcement represents a difficult decision necessary to drive improved profitability,” said Mike McMullen, president and chief operating officer, and CEO-elect. “The NMR team has been extremely dedicated and has made many excellent contributions. However, this action is a step in ensuring that our investments are placed on higher-value life sciences, applied markets and diagnostics solutions that will continue to drive growth across the company.”
Agilent will stop taking new NMR system orders immediately, but the company will continue to meet customer commitments for orders in progress and for ongoing support contracts. Agilent will continue to provide service on all installed NMR systems.
The company expects that this decision will eliminate about 300 jobs, mostly within the next 12 months. The majority of the affected positions are located in Yarnton, U.K., and Santa Clara, California.
Today’s announcement is part of Agilent’s strategy to address the business shortfalls of its Research Products Division. In early 2013 Agilent announced its exit of the OEM and Specialty Magnet business and later the MRI business to focus resources on the core NMR portfolio. Despite these adjustments, the NMR business has continued to fall short of growth and profitability objectives.
To cover the cost of exiting this business, Agilent will take an approximate $72 million restructuring charge in the fourth quarter. It expects a $20 million to $30 million decline in revenues in fiscal year 2015 due to the NMR business closure, but a positive impact of about $10 million in operating profit in FY15.
For the fourth quarter of 2014, Agilent anticipates non-GAAP earnings per share of $0.87 to $0.91, and projects revenues to be negatively affected by currency at about $13 million, and lower NMR-related revenues by about $12 million.
Keysight has posted its investor roadshow slide deck on its website at www.investor.keysight.com. “New Agilent” will post its investor roadshow slide deck on Friday, Oct. 17, after the market closes at www.investor.agilent.com.
Nuclear magnetic resonance (NMR) spectroscopy is an analytical chemistry technique used in quality control and research for determining the content and purity of a sample as well as its molecular structure. It is used primarily in academia and government, the pharmaceutical, biotech and chemical industries.
About Agilent Technologies
Agilent Technologies Inc. (NYSE: A) is the world’s premier measurement company and a technology leader in chemical analysis, life sciences, diagnostics, electronics and communications. The company’s 20,600 employees serve customers in more than 100 countries. Agilent had revenues of $6.8 billion in fiscal 2013. Information about Agilent is available at www.agilent.com.
On Sept. 19, 2013, Agilent announced plans to separate into two publicly traded companies through a tax-free spinoff of its electronic measurement business. The new company is named Keysight Technologies, Inc. The separation is expected to be completed in early November 2014.
This news release contains forward-looking statements as defined in the Securities Exchange Act of 1934 and is subject to the safe harbors created therein. The forward-looking statements contained herein include, but are not limited to, information regarding the separation of Agilent’s electronic measurement business; future revenues, earnings and profitability; the future demand for the company’s products and services; and customer expectations. These forward-looking statements involve risks and uncertainties that could cause Agilent’s results to differ materially from management’s current expectations. Such risks and uncertainties include, but are not limited to, unforeseen changes in the strength of our customers’ businesses; unforeseen changes in the demand for current and new products, technologies, and services; customer purchasing decisions and timing, and the risk that we are not able to realize the savings expected from integration and restructuring activities.
In addition, other risks that Agilent faces include those detailed in Agilent’s filings with the Securities and Exchange Commission, including our latest Form 10-K and Form 10-Q. Forward-looking statements are based on the beliefs and assumptions of Agilent’s management and on currently available information. Agilent undertakes no responsibility to publicly update or revise any forward-looking statement.
# # #
+1 408 345 8396
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+44 186 529 1472
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“Beer Manufacturing and Analysis by NMR”, John C. Edwards and Adam Dicaprio, Presented at the Mestrelab MNova Users, Meeting – SMASH, Atlanta, GA, September 7, 2014
Get PDF Here
“Survey of Low Field NMR Spectrometer Platforms for Successful Screening of Sexual Enhancement and Weight Loss Supplements for Adulteration with Drugs and Drug Analogs”, John C. Edwards, Kristie Adams, Anton Bzhelyansky , Presented at SMASH Conference, Atlanta, GA, September 7-10, 2014.
Get PDF Here
“Liquid and Solid-State 27Al qNMR of an Elemin Senonian Trace Minerals Supplement for Identification, Chemical Structure, Quantitation of Active Ingredient in the Product, and Product Stability”, Boris Nemzer, John C. Edwards, presented at XII International Conference on the Applications of Magnetic Resonance in Food Science: Defining Food by Magnetic Resonance, Cesena, Italy, May 20-23, 2014.
Get PDF Here
“Liquid and Solid-State Multinuclear 13C and 11B qNMR FruitexB Fructoborate Complex Nutritional Supplement. Identification, Chemical Structure, Quantitation of Active Ingredient in Product, and Product Stability”, Boris Nemzer, John C. Edwards, presented at XII International Conference on the Applications of Magnetic Resonance in Food Science: Defining Food by Magnetic Resonance, Cesena, Italy, May 20-23, 2014.
Get PDF Here
“1H qNMR of EPA and DHA Omega-3 Fatty Acid Esters – PLS Regression Models Obtained by 60 and 300 MHz NMR – At-Line and On-Line Monitoring of a Fish Oil Nutritional Supplement Manufacturing Process”, John C. Edwards, Paul J. Giammatteo, invited oral presentation at XII International Conference on the Applications of Magnetic Resonance in Food Science: Defining Food by Magnetic Resonance, Cesena, Italy, May 20-23, 2014.
“Application of High Field and Cryogen-Free Bench-Top NMR Platforms to the Monitoring and Quantitation of PDE5 Inhibitor Adulteration of Male Sexual Enhancement Supplements”, John C. Edwards, Paul J. Giammatteo, Kristie Adams, Anton Bzhelyansky , presented at XII International Conference on the Applications of Magnetic Resonance in Food Science: Defining Food by Magnetic Resonance, Cesena, Italy, May 20-23, 2014.
Get PDF Here
“NMR Based Authentication of Nutraceuticals, Herbal Supplements, and Food Additives: Economic- and Efficacy-Driven Adulteration of Aloe Vera, Herbal Erectile Dysfunction Supplements, and Acacia Gum”, John C. Edwards, invited presentation at the 2nd PANIC, Charlotte, NC, February 3-5, 2014.
Get PDF Here
“Development of an Automated Complex Mixture Analysis qNMR Method within Mestrelab MNova – Application to Aloe Vera and the Beer Brewing Process”, John C. Edwards, Adam J. Dicaprio, Michael A. Bernstein, presented at the 2nd PANIC, Charlotte, NC, February 3-5, 2014.
Get PDF Here
“Small Molecule Chemistry of Spontaneously Fermented Coolship Ales”, Adam J. Dicaprio, John C. Edwards, presented at the 2nd PANIC, Charlotte, NC, February 3-5, 2014.
Get PDF Here
“Liquid and Solid-State 1H, 13C, and 11B qNMR Analysis of Fruitex-B®– A Calcium Fructoborate Comple: Chemical Structure and identification, quantitative analysis and stability study”, Boris Nemzer, John C. Edwards, presented at the 2nd PANIC, Charlotte, NC, February 3-5, 2014.
Get PDF Here
I have been invited to be a guest editor for Wiley publishing to pull together 2 special issues of Magnetic Resonance in Chemistry. I am putting out a general “call for papers” but I will harass people personally. We are looking for 10-20 papers for each issue.
The deadline for submission of the papers is June 30 so no-one has an excuse that there isn’t enough time. I do ask that you email me at the contact below to let me know that you are thinking of submitting and a title would be nice also (though I won’t hold you to it). We hope to have the review and revide papers (if necessary) by late October and then publish by December 2014.
The first will be on high resolution benchtop NMR in which I would like to include all permanent magnet systems capable of obtaining a spectrum. This is an open invitation to all the vendors and their customers of (in no particular order) Anasazi, Nanalysis, One Resonance Sensors, Magritek/ACT, Aspect, Qualion, Picospin, Resonance Systems, Oxford Instruments, Bruker, home built devices, I would also make the exception that HTS systems be included as they are also cryogen free.
Any papers on spectrometers, magnets, educational, industrial, academic applications, chemometrics, automated approaches, reaction monitoring, online/at-line utilization, 2D NMR, combined spectral/relaxation applications.
The second will be on low resolution benchtop NMR but I would like to exclude applications that have been around for decades (H content, SFC, oil content, spin finish). I would like to encourage new applications to be submitted and they should include hardware, magnets, spectrometers, probes, 1D/2D Laplace inversion (I’d love a review/overview of that software aspect), applications. Again – all vendors and all users of commercial and home grown benchtop TD-NMR systems please submit.
We’re looking for articles on FPGA spectrometers, software approaches (1D/2D Laplace, chemometrics), magnet design and construction, dedicated rheology analysis instruments, field cycling NMR, unilateral NMR, core analyzers, applications of all whether educational, academic, industrial.
The types of articles can be Reviews, Mini-Reviews, Tutorial, Historical, Spotlight, Perspective, Communication, Article, Application Note, Case Report, Spectral Assignments, Correspondence.
Here is the official word on the use of color (note you can have all the color figures you wish in the online version but the print version is restricted): two colour illustrations per submission are allowed free of charge, however further colour illustrations are allowed at the Editors discretion and where they are justified. Authors can have as much colour as they like in the online version as the restrictions are just for the print version.
Please contact me directly if you would like to make a submission of your work to either of the two issues: firstname.lastname@example.org
I have posted the authors instructions for special Issues that I was given on my website at:
The MRC author guidelines can be found at:
A herbal supplement marketed to alleviate erectile dysfunction was recently submitted for testing in our laboratory because it was surprisingly effective considering it should only contain the traditional herbals utilized for this problem such as Oyster, 2-Deoxy-D Glucose, Barberry, Snow Lotus, Bombyx Mori L., Ginger Root, Salfron Crocus.
Above: 1H NMR of ED Herbal Supplement – this is an extract of the powder from the capsules into 80:20 CD3CN:D2O – Observed suspicious specific small molecule peaks – herbals are usually broad featureless complex mixture spectra. This is something in different – a specific drug material appears to have been added.Comparison with the spectrum of Viagra (Sildenafil Citrate) below shows a strong similarity between the 2 active ingredients.
We also obtained the COSY, 13C and DEPT NMR Spectra on the ED herbal supplement. These are shown below.
The analysis of the NMR data led us to find that the extractable components of the sample contained predominantly Sulfoaildenafil (thioaildenafil) which is a strutural analog of Sildenafil (Viagra) and is not approved by any health regulation agency for use in erectile dysfunction medicines or supplements. Apparently these structural analogs to Viagra are found widely in herbal supplements sold for this malady. I guess we have one more for the list that the FDA keeps on such dangerously adulterated products.
We have an excellent solid-state Doty Scientific Supersonic 7mm CP-MAS probe for our Varian UP-200 spectrometer. This is a nice field strength for solid-state 13C analyses as it allows for spectra at higher sensitivity than a 100 MHz but still has the advantage that 7 kHz is enough to push the aromatic MAS sidebands beyond 0 ppm so that more accurate aromaticity values can be calculated. The probe is still working after 20 years and we are using the original rotors and have never had to replace any of the capacitors. We have run approaching 20,000 experiments with this probe. Here are some of the nice spectra we have been getting recently on drug API and tablets.
Above: Solid-State 13C NMR of Rapamycin API
Above: Solid-State 13C NMR of Ranitidine HCl – Zantac Tablet