Research Synopsis: Applied analytical chemistry, multi element trace analysis of environmental and geological samples, CBD and omega-3 dietary supplements forensics, counterfeit consumer products, advanced chromatographic separations, laser desoprtion ionization TOF-MS, E-cigarette components.
Members of Professor Gene Hall's research team will be trained with the latest state-of-the-art instrumentation in analytical chemistry. These instruments include a Finnigan Element High Resolution ICPMS, gradient HPLC systems coupled to a Q-TOF-MS, GC-TOF-MS, MALDI, and FT-IR, Raman and EDXRF. Your multidisciplinary education will include forensic, biological, statistical, environmental, and analytical chemistry. After receiving your training and degree under my supervision, you will have excellent marketable skills to obtain employment in the exciting field of analytical chemistry. Recent activity has been devoted to the study of the composition of the e-cigarette cartridges that involve detailed composition analysis to determine why so many users of e-cigarettes are getting sick. The focus of Professor Hall's research team is composed of five major themes.
Current event hot topic theme.
Untargeted mass spectrometric, FT-IR, and XRF analyses of the components of e-cigarette components to provide insight into why so many users of e-cigarettes are getting sick. The culprit maybe the polydimethylsiloxane (PDMS) silicone elastomer used in the pods as shown below. We found that the PDMS is leaching into the e-liquid and is becoming part of the vapor that is inhaled. I am curious as to the effects of PDMS on the lungs and since PDMS contains Si, could there be a possible silicosis type effect. Is there an immunological effect? We have fingerprinted the liquid in the pods and compared this fingerprint with the components of the silicone gasket shown below in black. Using high resolution with accurate mass Q-TOF and LDI-TOF-TOF we confirmed the assignments of the isotopologues between the two samples and they match. Stay tuned!!!
Figure A. Typical e-pod showing components. Recycle your pods as the metal contacts are gold (Au) coated.
Notice the black silicone elastomer is in contact with the vape oil that contains a mixture propylene glycol and vegetable glycerin with the flavor contents. Others have used GC-MS to show more than 40 compounds in the vape oil.
We used a non-targeted approach to determine other components of the vape oil. The figure below compares a partial high resolution fingerprint between the vape oil and the black silicone polydimethylsiloxane elastomer. Notice how the isotope pattern matches although the samples were analyzed using two different mass spectrometric techniques using different solvents.
Figure A. Compare partial mass spectra of e-vape liquid vs black silicone elastomer.
Figure B. Using our LDI-TOF-TOF (matrix free) for rapid sample analysis.
Figure C. Using our Waters' Synapt Q-TOF to perform untargeted analyses of e-liquids.
Theme One: Identification and quantification of Pb, Cu, and Zn binding proteins in human tissues
Theme one is bio-analytical chemistry and focuses on the identification and quantification of Pb, Cu, and Zn binding proteins in human biological tissues. In collaboration with the University of Medicine and Dentistry of New Jersey (UMDNJ) - Robert Wood Johnson University Hospital, we obtain samples of human blood, follicular fluid, placenta, and amniotic fluid. We use high performance liquid chromatography (HPLC) as the method to identify and separate metal binding proteins in these fluids. The output flow solution from the HPLC is fed to an inductively coupled plasma mass spectrometer (ICPMS) to quantify elements bound to the proteins. This research allows for the first time a unique analytical method (HPLC-ICPMS) to investigate the effects of Pb speciation on metal binding proteins and Pb toxicity during and after pregnancy. Major concern is the transfer of Pb to the fetus during pregnancy and the transfer of Pb to the new born after birth..
The HPLC-ICPMS is used to study pre-natal Pb poisoning with emphasis of the importance of avoiding Pb before, during, and after pregnancy.
Theme Two: Environmental analytical chemistry: identification of sources of Pb in drinking water
Our second theme of research is environmental analytical chemistry that focuses on identification of sources of Pb in drinking water, consumer products (watches, dinner plates, children's toys) and food and drink. Because Pb has four stable isotopes, we determine the differences in the amount of these isotopes in drinking water. This unique water-lead isotope "fingerprint" is compared to lead isotope "fingerprints" from plumbing materials that include solder, faucets, valves, and Cu pipe. The "fingerprints" are determined by high resolution ICP-MS and statistical methods are used to determine the source(s) of Pb contamination.
For example, Figure 1 shows a Pb pipe that connects the water company's line to the consumer's water line. Several cities are replacing the Pb service lines but this could lead to a false sense of security because if the house has a Pb service line, chances are that the house has Pb-based solder. In our analysis, we found the Pb-based solder to be a major contribution to Pb in the tap water. In addition, faucets contain brass that also has Pb. Figure 2 is a typical plumbing system in a house serviced with well water. Using our Pb isotope fingerprinting, we can apportion the amount of Pb coming from each of the plumbing components (Cu pipe, solder, valves, and faucets) in a glass of water. The method can also be used to determine the sources of Pb in human blood.
Figure 1. Two Pb pigs. Pb isotope fingerprint shows Missouri Pb.
Figure 2. Typical well water system. Each plumbing material has a unique Pb isotope signature.
We also use non-destructive macro and micro energy dispersive X-ray fluorescence (EDXRF) to determine the amount of Pb, Sb, As, and Ba in various consumer products. For example, Pb imaging in a cartoon character watch marketed for children ages three and above that this watch is in violation of consumer protection laws.
Theme Three: Dietary Supplements Forensics
Our third theme is forensic investigations of dietary supplements that are marketed as containing omega-3 and omega-7 fatty acids. According to recent statistics, fish oil is the sixth most consumed dietary supplement in the US. Unfortunately, a majority of these misbranded and/or adulterated dietary supplements do not contain "fish oil". Instead, they contain marine biodiesel that are the ethyl esters of fatty acids that are enzyme inhibitors. We are using cold electron ionization gas chromatography time of flight mass spectrometry (GC-TOF-MS) to perform impurity profiles of these misbranded dietary supplements. The analyses of more than 3,000 dietary supplements (See Figure 3) has shown that they contain omega-3 fatty acids EPA, DHA, EPA, and ALA in the less bioavailable form ethyl esters.
If you have a question about a particular omega-3 sourced dietary supplement, chances are that our research team has analyzed and molecularly fingerprinted your dietary supplement. For example, Figure 4 shows the total ion chromatogram of a fish oil dietary supplement marketed as "100% natural". The chromatogram shows more than 50 different impurities that are not disclosed on the supplement facts panel as required by law.
Figure 3. Some of the more than 3,000+ dietary supplements analyzed by Professor Hall's research team.
Figure 4. TIC Truenutrition "fish oil" dietary supplement. Notice all the undisclosed impurities.
We are using computational chemistry assisted GC-TOF-MS to identify each of the more than 50 chemical compounds in this example misbranded and adulterated "fish oil" dietary supplement.
We are using advanced chromatographic methods such a gas chromatography coupled to a mass spectrometer to perform impurity profile of misbranded and adulterated dietary supplements. This allows a molecular fingerprint to characterize all kinds of impurities in the supplements using computational chemistry structure fragmentation analysis. We are also using supercritical CO2 as the mobile phase (green chemistry) to separate isomers of triglycerides that are the main components in fish, algae, and krill oils. Our method developments using a mass spectrometer (Q-TOF) revealed numerous molecular species that have not been previously identified. Figure 5 shows typical direct infusion mass spectrum and MS/MS of the dietary supplement Quell that is a re-esterified omega-3 dietary supplement. Again, members of our group are developing electronic databases and molecular fingerprints of these dietary supplements to understand how they contribute to good health. The structure and location of the acyl groups on the glycerol backbone of these re-esterified omega-3 dietary supplements is important for determining the hydrolysis of these molecules and the fate of the acyl groups for transformation to other lipid classes such as phospholipids and cholesterol esters.
Figure 5. Mass spectrum and MS/MS of Quell re-esterified omega-3 dietary supplement using direct infusion.
As a community service to the elderly and as a consumer advocate, our laboratory provides analytical analyses of consumer suspect dietary supplements. We have found numerous dietary supplements that target the elderly. For example, a dietary supplement called Cardia7 (See Figure 6) was purchased on Amazon with the claim "Proven to help and potentially reverse issues that can lead to bypass and open heart surgery.". We strongly advise that consumers not follow these outrageous therapeutic claims. We analyzed this dietary
Figure 6. Adulterated and misbranded dietary supplement, contains 0 mg palmitoleic acid vs label claims 210 mg and 0 mg fatty acids.
supplement and it did not contain any palmitoleic acid as claimed. The total ion chromatogram (TIC) is shown below in Figure 7 for Cardia7
Figure 7. Cardia7 TIC showing absence of "palmitoleic acid" per label claim of "210 mg palmitoleic acid".
containing the new dietary ingredient ProvinalTM using a simple dilute and shoot approach. Instead, the dietary ingredient was Provinal that is a mixture of synthetic fatty acid ethyl esters (FAEE) with the ethyl ester of palmitoleic acid being the most concentrated at approximately 50% w/w. FDA requires that the dietary ingredients (ProvinalTM) in a proprietary blend are to be listed in order of predominance by weight. If these compounds are not listed, the dietary supplement is misbranded. This means that all those impurities in Figure 7 must be included in the supplement facts so that the consumer will know what compounds they are ingesting.
We are in the process of determining the composition of all these undeclared impurities (>40) using computational chemistry with accurate mass molecular ion information. Once identified, we will use computational chemistry to determine how these compounds fit into the active site of the major hydrolyzing enzyme hCE2 (a human CES2 family isozyme) carboxylesterases (CESs, EC.220.127.116.11.) that release free fatty acids and ethanol into pancreatic acinar cells.
According to the FDA's 75-day pre-market review of this new dietary ingredient ProvinalTM, "For the reasons discussed above, the information in your submission does not provide an adequate basis to conclude that the dietary supplement containing "Provinal™", when used under the conditions recommended or suggested in the labeling of your product, will reasonably be expected to be safe. Therefore, your product may be adulterated under 21 U.S.C. 342(f)(1)(B) as a dietary supplement that contains a new dietary ingredient for which there is inadequate information to provide reasonable assurance that such ingredient does not present a significant or unreasonable risk of illness or injury. Introduction of such a product into interstate commerce is prohibited under 21 U.S.C. 331 (a) and (v) these are not dietary supplements. Most disturbing is that the NDI and GRAS/N state: "Toxicological studies on Provinal, the finished product, have not been conducted, and toxicological studies on pure palmitoleic acid were not found in the public domain. The absence of such studies is most likely due to the fact that this fatty acid, as well as the other fatty acids that constitute Provinal are considered “edible” and have an extensive history of consumption in foods." FDA's review of new dietary ingredients require at least "25 years of widespread use to be the minimum to establish a history of safe use." Since ProvinalTM was patented (US 8,703,818A) April 22, 2014, the 25 years of use was not obtained. In addition, both the GRAS/N and NDI claim a long history of use. If this was the case, then the patnet would be voided.
We have the certificate of analysis (CoA) from the South Korean company that manufactures ProvinalTM and the CoA contradicts the supplement facts of 210 mg palmitoleic acid. Under current good manufacturing practices (cGMP) the dietary supplement is adulterated because the CoA contradicts the identity of the dietary ingredient Provinal. Under 21 CFR 111.75(a)(1)(i) (cGMP), the manufacturer of a dietary supplement must conduct at least one appropriate test or examination to verify the identity of a component that is a dietary ingredient before using the ingredient in a dietary supplement. Our gas chromatographic method using a mass spectrometer verified that the dietary ingredient is not what was added to the supplement per supplement facts and therefore violation of cGMP and therefore adulterated. We are developing new analytical methods to assist in the confirmation and the identification of all different types of ingredients added to food and dietary ingredients. For example, we have developed a simple combination FT-IR GC-MS method to verify the identity and concentration (mass/mass, i.e. mass fraction) omega-3 dietary ingredients.
Note: Fatty acids have a statement of identity in 21 Code of Federal Regulations (CFR) . 101.9(c)(1) "Fat, total” or “Total fat”: A statement of the number of grams of total fat in a serving defined as total lipid fatty acids and expressed as triglycerides where fatty acids are aliphatic carboxylic acids consisting of a chain of alkyl groups and characterized by a terminal carboxyl group."
Therefore, consumers who purchased this and other Provinal based dietary supplements paid for what they thought was 210 mg palmitoleic acid but the dietary supplements contains 0 mg. Based on 21 U.S. Code 342(b) Adulterated food: "A food shall be deemed to be adulterated
(b) Absence, substitution, or addition of constituents
(1) If any valuable constituent has been in whole or in part omitted or abstracted therefrom; or
(2) if any substance has been substituted wholly or in part therefor; or
(3) if damage or inferiority has been concealed in any manner; or
(4) if any substance has been added thereto or mixed or packed therewith so as to increase its bulk
or weight, or reduce its quality or strength, or make it appear better or of greater value than it is."
Case law has confirmed this U.S. Code.
This is a nice example of using forensic dietary supplement investigation to determine the true identifity of a dietary supplement using paper trail and exploratory analytical chemistry analyses.
We were glad to assist this senior citizen to prevent them from perhaps further health damaging issues..
Figure 8. Total ion chromatogram (TIC) of >99% pure 9-z hexadecenoic acid (palmitoleic acid) and its fragmentation using cold electron ionization GC-TOF-MS.
This palmitoleic acid ethyl ester is cytotoxic and is the major cause of pancreas injury in alcoholics. Provinal is not GRAS for its intended use because the notification has numerous errors including wrong chemical structure, wrong ADME, and most importantly many publications are missing that contradict the notifier's claim of GRAS. Our research group are experts qualified by scientific training and experience to evaluate the safety of substances directly or indirectly added to food. This includes our graduate and undergraduate students.
Cannabidiol (CBD) is a hot new dietary ingredient used in dietary supplements. Unfortunately, it is not lawful to include CBD in dietary supplements because CBD is in the FDA approved drug product Epidiolex.
We are using non-destructive analytical methods such as ATR-FT-IR to show that the label contents are not correct and to support the FDA's conclusion that these products that claim they are dietary supplements are unlawful.
Figure 9. Unlawful CBD dietary supplements deemed adulterated near ATR 9 reflection diamond crystal for rapid FT-IR analyses.
Our research team is also experienced in food law and the marketing of dietary supplements. For example, we have used our extensive portfolio of state-of-the-art instrumentation to investigate GRAS notices in the context of "the same quality and quality of data needed to support a food additive petition". Therefore, a thesis could be centered around the composition of a GRAS substance evaluated under the regulations of a food additive petition. This includes chromatograms (FID), total ion chromatograms (GC-MS), IR spectra, absolute concentrations of the substance using mass balance, impurity profiling, toxicology, and bioavailability of the substance..
Fourth theme: Charactization of counterfeit consumer goods
Our fourth theme involves non-destructive characterization of counterfeit consumer products such as Rolex watches, Coach handbags, Barbie dolls, ink-jet cartridges and jewelry. We have an extensive collection of Coach handbags that were analyzed for trace elements using EDXRF, Raman, and ATR-FT-IR spectrometry. Using these non-destructive analytical methods, we can easily differentiate between the counterfeits and the genuines. Consumers who purchased these counterfeit products should be aware of high concentrations of Pb in some of the hangtags. Don't let your child suck or touch these toxic hangtags because the Pb can easily be transferred to your child via hand-to-mouth contact.
We are cautioning consumers on the purchase of fake Rolex watches because some of the crystals on the fake are arsenic oxide as shown in Figure 10.
Figure 10. Non-destructive micro EDXRF analyses of counterfeit Rolex watches.
Fifth theme: non-destructive characterization of postal stamps and banknotes.
Our fifth theme involves non-destructive characterization of postal stamps and banknotes. This is in collaboration with collectors as a means to characterize these precious cultural heritage objects. Figure 11 shows an old NJ banknotes that we used FTIR, Raman, and EDXRF to understand the trace element composition of the inks and papers.
We have collaborated with world-wide philatelists and numismatists to uncover counterfeit material and to study the history of the various paper documents. We welcome all collectors for joint non-destructive collaboration. One of our most challenging projects was to charaterize non-destructively the ARCHIMEDES PALIMPSEST and read the hidden text.
Figure 11. Early American NJ currency. Red ink is HgS and blue ink is Prussian blue.
We are using the non-destructive complimentary techniques, EDXRF, micro and macro FT-IR, and micro Raman spectroscopy to determine the materials used in counterfeiting banknotes. For example, we used these techniques to characterize a counterfeit Bank of England 50 Pounds banknote as shown below in Figures 12-15.
Figure 12. ATR-IR spectrum compare counterfeit paper vs genuine.
Figure 13. EDXRF spectra compare security stripes.
Figure 14. Micro Raman spectra compare security stripe counterfeit vs genuine.
Figure 15. Compare ATR-IR spectra of genuine vs counterfeit security stripe.
We also develop non-destructive analytic methods for characterization of art work using handheld-XRF instrument as shown below in Figure 16. A local resident wanted to know if the painting was "genuine" and who painted it.
Figure 16. Professor Hall analyzing a suspect painting by handheld EDXRF.
Figure 17. Adulterated and misbranded dietary supplements that contain 0 mg of "palmitoleic acid".
We used certified science-based analytical methods for our forensic molecular fingerprinting investigations of misbranded and adulterated fish oil dietary supplements. We found numerous errors in manufacturer’s marketing of fish oil dietary supplements. Based on our science-based approach, products with the claims of containing palmitoleic acid (9-Z-hexadecenoic acid, CAS # 373-49-9) were the most widespread misbranded and adulterated products being sold to unsuspected consumers. This amounts to what the FDA refers to as “health fraud”. One product that is advertised on Amazon called Cardia7 was sold to a senior citizen who asked our laboratory to investigate the contents of this dietary supplement. Below is the total ion chromatogram of Cardia7 that claims to be “100% natural” when in fact it is 100% synthetic based on the US patents and the GRAS/N and NDI notifications. Further inspection of the TIC and following the paper trail, Cardia7 with its Provinal is really marine biodiesel that has been treated with methanol or ethanol with urea to reduce the concentrations of palmitic acid ethyl ester.
Come join our exciting research team to participate in cutting edge NMR, FT-IR, Raman, XRF, chromatographic, mass spectrometry, forensic, computational chemistry, and chemometric characterization of dietary supplements. This will make a difference in marketing dietary supplements so that consumers especially the elderly are not scammed out of their fixed-income dollars. In addition, health-fraud dietary supplements are a federal crime.
Our consumer advocacy has already resulted in the removal from interstate commerce numerous “bogus” and “snake oil” dietary supplements both marketed as “fish oil” , “krill oil”, and “ultra-pure palmitoleic acid”.
We also welcome corporate collaborators.
What is in those popular infused bottled water samples? What the manufacturers don't know what they don't know. We are taking an untargeted approach to the analysis using high resolution mass spectrometry either direct infusion or using special LC columns to fractionate the different unknown compounds in the infused bottled water. So far, we have discovered several compounds that are not listed on the labels. Pesticides are one of our focus.
Figure 18. Untargeted molecular fingerprinting of bottled waters.
With the passage of the 2018 Farm Bill, states can now grow hemp products after state approval. However, the plant can't contain more than 0.2% THC. We are developing rapid methods for in-field applications to measure the composition of cannabinoids especially CBD and THC in the flowers as shown below in Figure 18.
MALDI imaging of embryo cells and plants.
New Waters' SYANPT G2 Si ion mobility mass spectrometer, we will be performing characterization of phospholipids in brain tissue extract to determine the acyl positions of DHA and ARA on the glycerol backbone.
Pharmacokinetics of omega-3 and omega-7 sourced dietary supplements in different lipid fractions of human blood. This will be performed without performing any derivatization of the fatty acyl chains. This will allow for the analysis of the native lipids as they appear naturally in the different blood fractions.
Isotope dilution Ag Au nanoislands laser desorption time of flight mass spectrometry to determine the concentrations of free fatty acids in human blood, omega-3 and omega-7 sourced dietary supplements, and edible oils to determine rancidity.
All the information presented on this web page is copyrighted by G Hall. Information is for educational purposes and the research is supported by and corroborated by using science-based approaches using well established scientific protocols. This science-based approach contradicts the marketing fiction found in the marketing of dietary supplements.
Here's How Scientists Detect Fake Fish Oil in Dietary Supplements: http://www.selectscience.net/editorial-articles/heres-how-scientists-detect-fake-fish-oil-in-dietary-supplements?artID=45760
Highly Efficient Luminescent Metal−Organic Framework for the Simultaneous Detection and Removal of Heavy Metals from Water” Nathan D. Rudd, Hao Wang, Erika M. A. Fuentes-Fernandez, Simon J. Teat, Feng Chen,∥ Gene Hall, Yves J. Chabal, and Jing Li., ACS Appl. Mater. Interfaces 2016, 8, 30294−30303
Some Insights on Retention and Selectivity for Hydrophilic Interaction Chromatography.” Yang, Mei (Monica); Thompson, Richard; and Hall, Gene. Journal of Liquid Chromatography & Related Technologies (2009) 32: 1–19.
Determination of total toxic arsenic species in human urine using hydride generation inductively coupled plasma mass spectrometry.” Xie, Ruimin; Johnson, Willie; Spayd, Steve; Hall, Gene S.; Buckley, Brian. Journal of Analytical Atomic Spectrometry (2007), 22(5), 553-560.
Determination of Ti, Zn, and Pb in Lead-Based House Paints by EDXRF”. Hall, G.S.; and Tinklenberg, J. Journal of Analytical Atomic Spectroscopy, 2003, 18, 775 - 778.
Lead In Breast Milk and Maternal Bone Turnover”. Sowers, M.F.; Scholl, T.O; Hall, G.; Jannausch, M.L.; Kemp, F.W.; Li, X.; and Bogden, J.D. American Journal of Obstetrics Gynecology, 2002, 187, 770 - 776.
Determination of Lead Sources in Water Samples Using Isotope Ratios”. Hall, G.S.; and Murphy, E. Bulletin of Environmental Contamination and Toxicology, 2000, 65, 314- 321.