Differences Between the Autosampler and the Manual Injector HPLCs
Development of commercially available high pressure liquid chromatography (HPLC) systems has opened up new horizons in research and development of cannabis/hemp. The application of biochemical and biomedical research conducted with high pressure liquid chromatography instrumentation is at the heart of transparency and compliance within the industry [1.] Start-up labs within the cannabis/hemp industry are consistently growing as regulations have shifted toward legalization and expansion of medical use across the United States and internationally. Regulation and testing are vital to ensure that products are consistent in quality for distribution to consumers.
Ionization Labs have become channel partners with Agilent Technologies and have developed an in-house potency testing software. The user is offered an Agilent 1220 HPLC instrument at a discounted rate alongside Ionization Lab’s proprietary testing software Cann-ID. Producers and other testing facilities can receive accurate and consistent results easily when using Cann-ID alongside an Agilent 1220 HPLC. The Cann-ID software operates on both a manual injector or an autosampler. Drawing a comparison between both an autosampler HPLC and a manual injector HPLC will highlight the benefits and drawbacks of both types of instrumentation. This will help illuminate where the future of this industry is heading in regards to cannabis testing instruments.
To begin, the most pronounced difference between an autosampler and a manual injector are the physical modules (pictured below in figure 1) that come with the Agilent 1220 HPLC. Manual injectors require more user involvement by utilizing an injector port, lever, and a waste vial. The autosampler is more complex with the addition of the vail tray for multiple samples and the robotic arm used to take samples to the needle/injection port. The autosampler employs a more elaborate method of injection compared to a manual injector which can impact results based on the consistency of the injection process.
Figure 1. A Manual Injector Module (left) and an Autosampler Module (right).
Another prominent difference between the autosampler and the manual injector is the accuracy and consistency of the injection. The consistency of the autosampler’s robotic arm injection is uniform and retains higher accuracy. This higher rate of accuracy reduces the percent error contributing to your sample data. The injection of the sample into the autosampler contributes to less than 1% of the probable error.
The manual injector’s accuracy is contingent on the user/technician’s accuracy and experience. The result’s validity is dependent on the technician, the injection port, and the syringe. Manual injectors have a higher percentage of probable error. These probable errors related to manual injection typically add up to approximately 4-10% of relative error or more. This can greatly alter the results, causing the data to be inconsistent even while testing the same sample.
Above all manual injectors face larger percent variance. The instrument relies on user proficiency, syringe performance, and injection technique to determine the repeatability of injections. These factors are not present when testing with an autosampler.
Additionally, a key difference between the two types of instruments is the time required to obtain results. The manual injector needs an operator to physically inject a vial every 15 minutes. This does not account for the extra time needed to clean the injection port , and sample loop between sample runs. Syringe injection can be relied upon for accuracy only if proper protocol is followed when handling the syringe. Often this method is time-consuming and even experienced lab analysts cannot always ensure result repeatability. This is due to variance in injected volumes. Syringes are fragile and users must be careful when handling throughout the injection. There is also an intricate cleaning procedure required prior to subsequent use  .
However the autosampler saves time in between vial injections compared to the manual injector allowing the operator time to prepare more samples. By reducing the time that the machine is idle, the autosampler uses less mobile phase, which is one of the more expensive consumables that is required to run the instrument. The autosampler does not require supervision while injecting and washing the needle unlike the manual injector. The autosampler minimizes the idling between vial injections; whereas the manual injector needs supervision before and after a vial is run . If the operator leaves during the run and gets busy with something else, the manual injector is idling, not running any samples, and wasting mobile phase. The autosampler does not require an operator present for each injection as it has an automated arm that picks up the vials, rinses the needle, and injects the samples. 
Consequently, the autosampler can run a higher volume of samples and can be programmed to run a shut-down sequence once finished. The manual injector can only run a limited number of samples in a given work day. During a regular eight hour work day, the manual injector is able to run about 32 vials (16 samples total), and that’s if the operator is running samples for eight hours straight with no breaks or letting the instrument idle in between injections.
On the contrary an autosampler can run more than 16 samples in an eight hour period; it can run samples all night, as long as there is enough mobile phase to accommodate the volume of samples. The shut-down method can be programmed at the end of a sequence, essentially putting the instrument to sleep. This sequence flushes out the system then turns off the oven, detector, and gradient pump when finished. With a manual injector, if the operator is restricted to an eight hour work day, they have to allocate the time needed for at least the start-up of the instrument, which increases overall wait time for results.
Although autosamplers are the future of HPLC technology and tend to be the preferred method within the industry; manual injectors can gain utility in some training or education settings. When training a lab analyst it may be beneficial to use a manual injector given it’s need for consistent attention from the operator. The manual injector gives first-hand experience that may aid learning about the testing process and how the instrument functions. Also if a user is able to duplicate manual injection results, that may indicate a level of precision and detail while cleaning and injecting the sample runs. Another reason manual injectors may be preferred is due to the cost being considerably less in comparison to autosamplers.
Specifically in laboratories where budgetary restraints are a factor, manual injectors may be more suitable. Commonly, this may be the case for a university laboratory or a start-up lab where the volume of samples is more manageable. Syringe injection is the most common introduction to HPLC instruments and can be more hands-on when practicing how to use the instrument. It is the more affordable option between the two which lowers the cost barrier of entry into the analytical lab economy. Manual injectors suit low volume laboratories and are great as an introduction into the system.
Thus autosamplers are more suitable for high volume testing and are more advisable for commercial laboratories. Sometimes the affordability of the manual injector may be an advantage, however the autosamplers are typically preferred based on their accuracy, efficiency , and capacity for a larger volume of samples. Consequently the overhead cost of paying analysts to work within the lab is reduced due to time saved , and lower margin of error.
To illustrate how injections are taken from vials by the robotic arm of the autosampler below is a screen capture from Cann-ID’s easy to use interface. In numbered positions users follow the software planned sequence and errors of selection from such a large number of vials is avoided. Moreover, the largest advantage offered by autosamplers is time saved. After the samples are prepared and loaded into the vial tray, the analysis sequence is set and the analyst is free to tend to other tasks (see figure 2 ). As stated previously autosamplers can operate overnight when no one is around which increases the laboratories potential sample output. These advantages have made autosampling the most desirable option of the two, however manual injections also have advantages for educational purposes, and the lower budget sector of hemp/cannabis lab operations.
Figure 2. Screen Capture of Cann-ID softwares vial tray interface in numbered sequence.
Therefore the best available option on the market currently is the Agilent 1220 Auto-injector with Cann-ID software and technical support. With the help of an autosampler it offers several advantages over a manual injector including but not limited to: regulating injection volume, automated needle wash, run time, vial selection, and shut down sequence. Cann-ID alongside the Agilent autosampler can load 95+ vials during a given work day into the numbered positions within the autosampler vial tray ( Figure 2). This makes testing 100+ samples a month much more manageable, and ensures lower potential of errors during sample testing.
In summary, using the Agilent 1220 Infinity II LC automated injector alongside Cann-ID software, is the most efficient, easy, and accurate tool to use when doing potency testing on a HPLC instrument. Cann-ID also accommodates more advanced instrumentation such as the 1260 Infinity II LC System and Ionization Labs is constantly working to evolve as HPLC systems upgrade and change. Throughout Ionization Labs has called attention to the pros and cons of manual and automated injection systems. We emphasize the difference between the two methods , and highlight their distinctive utilities.
For more information about Cann-ID and how to purchase an Agilent HPLC through Ionization Labs please visit our website at www.Cannid.com or www.Ionizationlabs.com . For answers to any questions regarding instrumentation and site requirements to start testing at your own facility today please email [email protected]. If you would like to call Ionization Labs to plan a demonstration of our Cann-ID software or inquiry about pricing please call us at (512) 470-7262.
 Brown, PR. High Pressure Liquid Chromatography; Biochemical and Biomedical Applications, Academic Press, Inc. ( London) LTD. , NY, Academic Press, New York, 1973, pp. 1–15.
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 Paul, Carsten, et al. “HPLC Autosamplers: Perspectives, Principles, and Practices.” Perspectives in Modern HPLC, LGC North America , Aug. 2019, https://www.researchgate.net/profile/Michael-Dong-3/publication/342901878_HPLC_Autosamplers_Perspectives_Principles_and_Practices/links/5f0ca3c092851c38a51a1540/HPLC-Autosamplers-Perspectives-Principles-and-Practices.pdf.
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