Ask The Expert: How to Overcome Challenges with Chromatography
David Ji, laboratory director at Analytical Laboratories in Anaheim, Inc., talks about the different ways in which his service laboratory uses chromatography techniques to analyze complex sample mixtures for various clients.
David Ji, Laboratory Director at Analytical Laboratories in Anaheim, Inc., talks to Tanuja Koppal, Ph.D., contributing editor to Lab Manager, about the different ways in which his service laboratory uses chromatography techniques to analyze complex sample mixtures for various clients. They are often asked to analyze and quantify trace components in samples such as nutraceuticals, botanicals, cosmetics and others. Ji and his team of chemists develop new chromatography methods and modify existing ones to get the best possible results in the least amount of time.
Q: Do you use a lot of chromatography techniques in your work?
A: Yes, we routinely use liquid chromatography (LC), mainly high performance liquid chromatography (HPLC), ion chromatography (IC) and gas chromatography (GC). We analyze a lot of natural herbs, dietary supplements like vitamins and minerals, and we have to do a lot of separation work, since our sample matrix tends to be very complex. We have to separate and quantify individual components using different instruments and columns. Most of our work is done using LC. We use IC to separate and quantify all kinds of sugars, such as sucrose, fructose, glucose, ribose, inositol, etc. using a conducting electrochemical detector. We also use an IC/conductivity detector to test organic and inorganic ions and some organic acids. We use GC with a flame ionization detector (FID) to test all kinds of fatty acids, including phytosterols and ginkgoterpenoids.
Q: What are some of the challenges you face using techniques like HPLC?
A: For our analysis we have to separate individual components. Traditional HPLC can do the job, but it takes a long time for achieving good separation. We have to slow down the gradient and that consumes a lot of solvent. To overcome this challenge we have now shifted to using Ultra Performance Liquid Chromatography (UPLC), where the packing particles in the column have much smaller size and that increases column efficiency. The column efficiencies increase three to five times compared to traditional HPLC. With the larger particle size in a traditional HPLC we can’t increase the solvent velocity too much, as the back pressure in the column can increase too.
Q: Are there any disadvantages to using UPLC?
A: There are no disadvantages, but since particle size is small the back pressure is high, around 10,000 psi. The cost of the UPLC columns is not much different compared to the traditional HPLC. The lifetimes of the columns are also quite similar. We typically replace columns after about 5,000 injections, but that depends on the sample type. All HPLC instruments last at least ten years.
Q: Do you have to deal with sample preparation problems?
A: Our sample matrix is very complex. We can use solid phase extraction (SPE), but detecting trace amounts of ingredients is not possible with SPE and we would also need to do a lot of validation. The best way is for us to inject the sample directly into the HPLC after dissolving it in water or some other solvent. By doing this no sample preparation or pre-column clean-up is needed. However, we do need to increase the detection sensitivity in order to dilute the sample and increase the life of the column. We also need to wash the column regularly and perform routine maintenance.
Q: What do you look for in terms of features when investing in an HPLC?
A: First of all, because we work with very complex samples, I look for an instrument or column that would give us a high efficiency for separation. Next, I consider the time taken for separation and finally, I also consider the type of solvent used, particularly since some solvents tend to be very expensive these days. Everything else, including the software, tends to be fairly standard and often not an issue.
The images shown are chromatograms comparing the separation of components in a mixture using HPLC and UPLC. For a complex matrix, such as dietary supplements, UPLC is at least three times faster than HPLC and shows better separation. For a single ingredient UPLC can perform even faster. For method development UPLC significantly increases the efficiency by saving time and costs. It also helps save solvent and waste storage cost. In our lab, each UPLC saves ~300ml of acetonitrile every day. Every year we also save about $400 to $600 waste treatment fee. When compared with HPLC, UPLC doesn’t have additional cost for column and instrument maintenance. (Source: David Ji)
David Ji is the laboratory director at Analytical Laboratories in Anaheim, Inc., a FDA registered testing laboratory that specializes in the chemical analyses of vitamins, botanicals, nutritional supplements and cosmetics. Their services include testing of raw materials and finished products, stability testing, method development and validation and consultation for new product development. Ji and his team of chemists routinely employ various types of separation techniques to develop analytical procedures that are quality controlled, GMP-compliant and fully documented. As a part of their consultation services, they troubleshoot problems, improve upon existing protocols or recommend new ones. As the director, Ji is responsible for development and validation of analytical test methods, for laboratory troubleshooting and technical support, for customer service and technical consulting. Ji has a M.S. in chemistry from the University of North Carolina-Greensboro and has held similar managerial positions at other laboratories for more than a decade.