Artificial Intelligence Applications in Chemistry - American Chemical

23 An Expert System for Optimizing Ultracentrifugation Runs Downloaded by NORTH CAROLINA STATE UNIV on December 29, 2017 | http://pubs.acs.org Publication Date: April 30, 1986 | doi: 10.1021/bk-1986-0306.ch023

Philip R. Martz, Matt Heffron, and Owen Mitch Griffith Beckman Instruments, Inc., Fullerton, C A 92634

The SpinPro U l t r a c e n t r i f u g a t i o n Expert System i s a computer program that designs optimal u l t r a c e n t r i fugation procedures to s a t i s f y the investigator's research requirements. SpinPro runs on the IBM PC/XT. Ultracentrifugation is a common method in the separat i o n of b i o l o g i c a l materials. I t s c a p a b i l i t i e s , however, are too often under-utilized. SpinPro addresses t h i s problem by employing Artificial Intelligence (AI) techniques to design e f f i c i e n t and accurate u l t r a c e n t r i f u g a t i o n procedures. To use SpinPro, the investigator describes the centrifugation problem i n a question and answer dialogue. SpinPro then offers detailed advice on optimal and a l t e r n a t i v e procedures for performing the run. This advice results i n cleaner and faster separations and improves the e f f i c i e n c y of the u l t r a c e n t r i f u g a t i o n laboratory.

U l t r a c e n t r i f u g a t i o n i s a common and powerful method i n the separa t i o n of b i o l o g i c a l materials. Despite i t s widespread use, however, few investigators f u l l y e x p l o i t i t s c a p a b i l i t i e s . As a r e s u l t , run times are unnecessarily long and separations are i n d i s t i n c t . In the long run, the e f f i c i e n c y and performance of the laboratory s u f f e r . The fundamental cause of t h i s s i t u a t i o n i s the increasing complexity of the u l t r a c e n t r i f u g a t i o n environment; the investigator must select the run parameters from a growing l i s t of rotors, gradient materials, and l i t e r a t u r e references. Knowing which rotor to use and at what run speed and run time i s a d i f f i c u l t matter. Furthermore, the s e l e c t i o n of one parameter complexly l i m i t s the a v a i l a b l e choices f o r the remaining parameters. Reliance on procedures reported i n the l i t e r a t u r e has compounded the problem. Often these procedures, perhaps i n i t i a t e d by investigators with a l i m i t e d set of rotors, are i n e f f i c i e n t by today's standards: the rotor i s inappropriate, the run speed i s too slow, or the run time i s too long. A new investigator applying t h i s procedure does not take f u l l advantage of the p o t e n t i a l of u l t r a centrifugation. 0097-6156/ 86/ 0306-0297S06.00/ 0 © 1986 American Chemical Society

Pierce and Hohne; Artificial Intelligence Applications in Chemistry ACS Symposium Series; American Chemical Society: Washington, DC, 1986.

ARTIFICIAL INTELLIGENCE APPLICATIONS IN CHEMISTRY

Downloaded by NORTH CAROLINA STATE UNIV on December 29, 2017 | http://pubs.acs.org Publication Date: April 30, 1986 | doi: 10.1021/bk-1986-0306.ch023

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One s o l u t i o n to the problem i s to provide the i n v e s t i g a t o r with technical advice. Good advice should y i e l d several immediate benefits: 1) Reliance on inappropriate or outdated techniques can be eliminated. 2) Better use can be made of the a v a i l a b l e equipment; shorter run times and improved separations w i l l r e s u l t . 3) The advice can be s p e c i f i c to the research requirements of the investigator. 4) The time usually wasted i n performing standardization runs, designing an u l t r a c e n t r i f u g e procedure, or researching u l t r a c e n t r i f u g a t i o n techniques can be minimized. In general, good advice w i l l improve the procedures, and thereby, improve the e f f i c i e n c y of most laboratories. Designing e f f i c i e n t u l t r a c e n t r i f u g a t i o n procedures and prov i d i n g good advice, however, i s a complex task; the knowledge and experience of an u l t r a c e n t r i f u g a t i o n expert are often required. In t h i s paper we describe a computer program, the SpinPro U l t r a c e n t r i fugation Expert System, that designs u l t r a c e n t r i f u g a t i o n procedures i n response to the requirements of the i n v e s t i g a t o r . SpinPro runs on the IBM PC/XT. The program i s based on techniques from the f i e l d of A r t i f i c i a l I n t e l l i g e n c e (AI) and expert systems: the powerful c a p a b i l i t i e s of the Lisp programming language; an inferencing procedure capable of drawing conclusions from a complex knowledge base; and a knowledge base derived from the expertise of u l t r a centrifugation experts. Indeed, SpinPro's use can be compared to the advice any person might seek from an expert. The i n v e s t i g a t o r and SpinPro enter into a question and answer dialogue i n which the investigator describes the research goals and sample characteri s t i c s . At the conclusion of the dialogue, SpinPro produces the following reports: 1. 2.

3. 4.

The Design Inputs Report i s a summary of the SpinProinvestigator dialogue. The Optimal Plan Report describes an optimal u l t r a c e n t r i f u g a t i o n procedure designed to solve the problem described i n the dialogue. I t uses the most appropriate rotor from the entire l i n e of Beckman r o t o r s . The Lab Plan Report i s s i m i l a r to the Optimal Plan, but i t describes a procedure based e x c l u s i v e l y on the u l t r a c e n t r i f u g e s and rotors a v a i l a b l e i n the investigator's laboratory. The Plan Comparisons Report compares the Optimal Plan and Lab Plan, i d e n t i f y i n g s i g n i f i c a n t differences and trade-offs between the two plans.

The reports constitute a complete set of recommendations f o r the u l t r a c e n t r i f u g a t i o n problem posed to SpinPro. Thus, SpinPro performs the advisory role of an u l t r a c e n t r i f u g a t i o n expert: interviewing the i n v e s t i g a t o r f o r the problem d e s c r i p t i o n , o f f e r i n g expert advice on the most appropriate c e n t r i f u g a t i o n procedure, and f i n a l l y , comparing a l t e r n a t i v e procedures. Major Functions SpinPro has four major functions: CONSULTATION, INFORMATION, CALCULATION, and CONFIGURATION. The CONSULTATION function performs the role of expert advisor. I t i s the main topic of t h i s paper. The INFORMATION function provides a database of u l t r a c e n t r i f u g a t i o n


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techniques, centrifuges, rotors, and l i t e r a t u r e references. The CALCULATION function performs a variety of routine c a l c u l a t i o n s including rotor speed reductions, k f a c t o r s , and p e l l e t i n g time. The CONFIGURATION function records the ultracentrifuges and rotors i n the investigator's laboratory. This information i s used by the CONSULTATION function when designing a run using the equipment from the laboratory.


User Interface A l l user inputs are made by pointing at text on the computer screen with a "mouse" controlled cursor. The mouse i s a hand-held pointing device, which when moved by the investigator over a f l a t surface, controls the movement of a cursor or pointer on the computer screen. To run the CONSULTATION function, the user points at the text "CONSULTATION" on the screen and c l i c k s the mouse button. When using SpinPro, the keyboard i s not required. In our observations, novice users of the program have been able to design u l t r a c e n t r i fugation procedures w i t h i n minutes of using the program. The CONSULTATION Function The primary goal of the CONSULTATION function i s to provide the best advice possible on precisely how to set up and run an u l t r a c e n t r i fugation procedure that i s s p e c i f i c a l l y designed f o r the i n v e s t i gator's research. SpinPro addresses v i r t u a l l y a l l problems i n the u l t r a c e n t r i f u g a t i o n of b i o l o g i c a l samples excluding whole c e l l s . To t h i s end, SpinPro i s "knowledgeable" about d i f f e r e n t i a l , rate-zonal, and isopycnic methods. I t addresses the separation of proteins, glycoproteins, proteoglycans, l i p o p r o t e i n s , subcellular f r a c t i o n s , nucleic a c i d s , and v i r u s e s . SpinPro's rotor knowledge includes swinging bucket, f i x e d angle, v e r t i c a l tube, zonal, and continuous flow rotors. Operation The CONSULTATION function i s run by using the mouse to select the text "CONSULTATION" from the computer screen. The f i r s t question of the dialogue, "Please enter the class of your sample of i n t e r e s t " , appears on the screen. The pop-up menu l i s t s the sample types to chose from. The i n v e s t i g a t o r then uses the mouse to select the appropriate response from the pop-up menu. This question and answer procedure continues u n t i l SpinPro has enough information, t y p i c a l l y 10 to 15 questions, from which to i n f e r a l l of the relevant parameters. The dialogue i s directed by SpinPro i n response to answers to previous questions. Thus, i f the sample i s a p r o t e i n , SpinPro requests the sedimentation c o e f f i c i e n t ; i f the sample i s a n u c l e i c a c i d , SpinPro requests the type of nucleic a c i d . At the conclusion of the dialogue, the reports are w r i t t e n to the disk. Using the pop-up menu, the reports can be read or saved. The dialogue includes c a p a b i l i t i e s to increase i t s f l e x i b i l i t y . F i r s t , the i n v e s t i g a t o r can change an answer to a previous question without d i s r u p t i n g the course of the dialogue. This c a p a b i l i t y i s useful when describing a problem that d i f f e r s only s l i g h t l y from a previously described problem. Second, the i n v e s t i g a t o r can ask why



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the current question i s being asked. The "Why?" function informs the user what SpinPro i s attempting to i n f e r ( i . e . , the l i n e of reasoning) at any p a r t i c u l a r step, and i t describes the a f f e c t that d i f f e r e n t answers w i l l have on the l i n e of reasoning. Third, when the answer to a question i s not known, the investigator can answer the question with "unknown". Depending on the question, SpinPro responds either by asking a related question or by assuming a reasonable answer and designing the procedures based on this assumption. Any assumptions that have been made are noted i n the reports. F i n a l l y , for the experienced users of SpinPro, there i s the option to request that, during the dialogue, a short form of the question be used. Optimization C r i t e r i a Two of the dialogue questions are of unique importance and are p a r t i c u l a r l y representative of SpinPro's c a p a b i l i t i e s . The f i r s t i s a question of research requirements. Every u l t r a c e n t r i f u g a t i o n procedure should r e f l e c t the investigator's concern f o r purity of the separation or short run time, goals that often run counter to each other. Rarely does any procedure state t h i s trade-off e x p l i c i t l y . The optimization c r i t e r i a question, "Select one of the following optimizations:", not only i d e n t i f i e s the trade-offs involved when designing a procedure, but allows the investigator to control them. The investigator can select the c r i t e r i o n which s a t i s f i e s the s p e c i a l i z e d requirements of the research. The c r i t e r i a are: 1) p u r i t y , 2) minimize run time, 3) minimize cumulative run time, 4) minimize number of runs, 5) continuous flow rotor procedures, and 6) procedures f o r processing many samples of small volume. Based on the optimization c r i t e r i o n , SpinPro can select the most appropriate rotor. For example, suppose the investigator has a r e l a t i v e l y large sample volume, a l l of which needs to be processed as soon as possible. The "minimize cumulative run time" c r i t e r i o n would be the appropriate choice. SpinPro would then i n i t i a t e the following rotor selection procedure: SpinPro determines the t o t a l sample volume based on inputs of the sample volume, the current concentration of the sample, and a correction f o r any pre-run d i l u t i o n s of the sample. Next, consideration i s made f o r whether tubes or bottles w i l l be used. The program then evaluates rotors for the number of tube positions and the amount of sample per tube. At t h i s point, SpinPro w i l l have estimated f o r each rotor the number of runs required to process the sample. SpinPro then estimates the run time f o r each rotor to perform a single run. Based on these estimates, SpinPro selects the rotor that w i l l give the shortest t o t a l run time when the run time i s summed over the t o t a l number of runs. S i m i l a r l y , the investigator can select any of the optimization c r i t e r i a and i n i t i a t e a v a r i e t y of precise rotor s e l e c t i o n procedures. Lab Rotors The second question of unique importance concerns the investigator's s e l e c t i o n of a rotor f o r the Lab Plan. Whereas, i n the Optimal Plan, SpinPro selects the rotor; i n the Lab Plan, the investigator selects the rotor. The i n v e s t i g a t o r , however, i s not required to


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select the rotor b l i n d l y from those available i n the lab. SpinPro a s s i s t s i n the s e l e c t i o n by assigning each of the rotors i n the lab to a category based on how well the rotor s a t i s f i e s the requirements of the problem. The categories are as follows: 1. 2.


3.

4.

Optimal rotors - the rotors that are both best suited to perform the run and to achieve the stated optimization c r i t e r i o n . Alternate rotors - other rotors that are not optimal but can perform the run. Not q u a l i f y i n g rotors - rotors that are not recommended for the problem usually because they are too large or too small for the sample volume, or because they do not generate s u f f i c i e n t l y high c e n t r i f u g a l forces. Not compatible rotors - rotors that are not c l a s s i f i e d , as part of the rotor safety program, for running i n the ultracentrifuge chosen from the lab.

The investigator can select any rotor from categories 1 and 2 above. This allows the investigator to experiment with the rotors i n the lab and to design procedures as variations on the theme established i n the Optimal Plan. Ultimately, the rotor selected i n the Optimal Plan by SpinPro and i n the Lab Plan by the investigator are the major source of difference i n the run parameters, p u r i t y , and o v e r a l l effectiveness of the two plans. The Design Inputs Report As noted e a r l i e r , SpinPro writes four reports regarding the recommended procedures. The Design Inputs Report summarizes the questions posed by SpinPro and the answers provided by the i n v e s t i g a t o r . A Design Inputs Report i s shown i n figure 1. The pop-up menu on the r i g h t allows the user to switch between reports, p r i n t the reports, or perform other functions. The report summarizes the problem that i s addressed by the Optimal Plan (Figure 2) and the Plan Comparisons Report (Figure 3). A summary of the report follows: The problem i s to separate proteins. Furthermore, SpinPro should pay p a r t i c u l a r attention to the p u r i t y of the separation. The sample i s not negatively affected by sucrose, has a sedimentation c o e f f i c i e n t of 16 Svedbergs, and i s i n l i q u i d form of 3 mL and a concentration of 1% w/w. The protein of i n t e r e s t should be placed 45% from the top of the gradient at the end of the run. Of the gradient concentrations 10-40% and 5-20%, the 10-40% i s preferred by the investigator. There are no solvents i n the sample that are harmful to the tubes. F i n a l l y , from the lab, SpinPro should use the L2-75B u l t r a c e n t r i f u g e and the SW 41 T i r o t o r , which does not require a speed derating due to i t s age. The Optimal Plan Report The Optimal Plan i s SpinPro's recommendation of how best to perform the run. The Optimal Plan of figure 2 i s underlined and annotated below:




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SpinPro Ultracentrifugation Expert System Design Inputs Experiment: SpinPro Consultation 11-Sept-1985 9:30:00 Particle class: Protein Separation vs Concentration: Separation Optimization criterion: Purity Assoc/Dissoc in sucrose: No Sedimentation coefficient: 16.0 10-40% or 5-20% gradient?: 10-40 Sample form: liquid/semi-solid Total sample volume (mL): 3.0 Sample concentration % w/w: 1.0 Selected final location: 45.0 Solvents: No

Page Forward Page Backward Optimal Plan Lab Plan Comparisons Design Inputs Change Answer Save Reports SpinPro Top Exit to D O S

Selected lab centrifuge: L2-75B Selected lab rotor: S W 41 Ti Rotor derated?: No

Figure 1. The Design Inputs Report f o r the problem described to SpinPro. The Optimal Plan and the Lab Plan are based on t h i s problem. The pop-up menu on the right allows switching to the other reports or performing other functions.


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SpinPro Ultracentrifugation Expert System Optimal Plan Experiment: SpinPro Consultation 11-Sept-1985 9:30:00 This is a complete plan for a protein sample separation Optimization criterion: Purity


Method: Density gradient, Rate-zonal Gradient: 10-40% continuous sucrose Rotor/run conditions: S W 55 Ti rotor at 55000 rpm for approximately 6 hours Potential tube materials: Polyallomer, Ultra-Clear Centrifuge: L8-80M set at 4 degrees C Omega-squared t: 7.132x10 11 Acceleration/deceleration: fast/fast Λ


Prior to the run prepare sample as follows: No special sample preparation is required. Load 0.3 mL of the Protein sample in full tubes at the top position of the gradient. At the end of the run the 16 S particles will be approximately 45% from the top of the gradient. To process the entire sample volume requires approximately 2 centrifuge run(s) with an estimated total run time of 12 hours, 5 minutes.

Figure 2. The Optimal Plan Report f o r the problem described i n the Design Inputs Report of figure 1. The plan gives the recom mended procedure f o r doing the run.

SpinPro Ultracentrifugation Expert System Plan Comparisons Experiment: SpinPro Consultation 11-Sept-1985 9:30:00 Run summaries: Optimal: SW 55 Ti at 55000 rpm for 6 hours per run in 2 run(s). Requiring a total of approximately 12 hours, 5 minutes Lab: S W 41 Ti at 41000 rpm for 15 hours, 45 minutes per run in 2 run(s). Requiring a total of approximately 31 hours, 30 minutes Comparisons: The Optimal Plan requires 38% of the Lab Plan run time for a single run. It requires 38% of the Lab Plan run time when processing the entire sample.


Figure 3. The Plan Comparisons Report compares the Optimal and Lab Plans. The comparison shows that, because the Lab Plan uses the SW 41 T i r o t o r , the run times are dramatically d i f f e r e n t .


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This i s a complete plan f o r a protein sample separation. A l l of the relevant parameters have been inferred i n a "complete plan". " P a r t i a l plans" indicate that one or more parameters could not be determined.


Optimization c r i t e r i o n : P u r i t y . The report restates the optimizat i o n c r i t e r i o n chosen by the investigator. Method: Density gradient, Rate-zonal. The rate-zonal method i s one of s i x addressed by SpinPro. The other methods are d i f f e r e n t i a l , d i f f e r e n t i a l - f l o t a t i o n , discontinuous, isopycnic, and 2-step isopycnic. These methods d i f f e r dramatically i n t h e i r set up, p r i n c i p l e s of operation, and expected r e s u l t s . The rate-zonal method i s described here b r i e f l y so that the recommendations to follow can be appreciated. P r i o r to the run i n a rate-zonal method, a gradient material i s introduced to the rotor tubes i n steps of increasing density from the top to the bottom of the tube. The sample to be separated i s layered, as a t h i n band, on the top of the gradient. As the run begins, each component i n the sample moves toward the bottom of the tube. Some components sediment f a s t e r than others. This fact i s the basis f o r the separation. I f the run parameters are appropriate, the components w i l l form separate bands within the gradient. At the conclusion of the run, the band representing the component of interest can be removed from the tube. Gradient: 10 - 40% continuous sucrose. SpinPro usually selects the gradient concentration and the gradient material. Here, SpinPro narrowed the choices to the 5-20% or 10-40% gradient, noting i n the dialogue that a trade-off between p u r i t y and run time e x i s t s between the 5-20% and the 10-40% gradient, but e i t h e r w i l l work. The invest i g a t o r selected the 10-40% gradient. The investigator could, i f desired, f i n i s h the plan based on the 10-40% gradient, and then using the change answer function, t r y the 5-20% gradient to f i n d out how the recommendations d i f f e r . Sucrose i s the gradient material of choice here. SpinPro considers a wide v a r i e t y of gradient materials including cesium c h l o r i d e , Nycodenz, Metrizamide, g l y c e r o l , and potassium t a r t r a t e . Rotor/run conditions: SW 55 T i rotor at 55000 rpm f o r approximately 6 hours. These recommendations form the core of any procedure. SpinPro usually considers more factors i n the rotor s e l e c t i o n process than does the expert. In determining the run speed, SpinPro considers every possible reason to reduce the run speed. If there are none, the rotor i s run at f u l l speed. When there are reasons (e.g., when using s a l t gradients, b o t t l e s , d i f f e r e n t i a l p e l l e t i n g , or discontinuous runs), the run speed may have to be reduced dramati c a l l y , from 80,000 rpm to 40,000 rpm, for example. There are many cases of rotors being run too slow for the a p p l i c a t i o n or too fast for safety. Accurate determination of the run time i s a complex problem based on the gradient c h a r a c t e r i s t i c s , c a l c u l a t i o n s , i n t e r polations from numerical tables, and experience. SpinPro employs a l l of these methods i n order to i n f e r run times f o r many s p e c i a l cases.


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P o t e n t i a l tube materials: Polyallomer, Ultra-Clear. SpinPro checks that a l l gradient materials, samples, and solvents are compatible with the tube materials. The a f f e c t s of acids, bases, o i l s , organic solvents, and s a l t s on the tube materials are considered.


Centrifuge: L8-80M set at 4 degrees C. The Optimal Plan recommends the L8-80M u l t r a c e n t r i f u g e . SpinPro selects a temperature that w i l l protect the i n t e g r i t y of the sample. Omega-squared t : 7.132xl0Ell. SpinPro calculates this measure of the t o t a l force applied to the gradient and sample during the run. Acceleration/deceleration: f a s t / f a s t . Many investigators overlook the a f f e c t that improper acceleration or deceleration can have on disrupting the separation, e s p e c i a l l y when re-orientation of the gradient occurs i n f i x e d angle or v e r t i c a l tube rotors. SpinPro addresses many s p e c i a l cases. P r i o r to the run prepare sample as follows: No special sample preparation i s required. Proper sample preparation i s important to prevent overloading the gradient. A sample that i s too concentrated w i l l d r i f t through the gradient before the run i s started. If the sample i s i n a proper form, as i t i s here, then no preparation w i l l be recommended. ,,

,,

Load 0.3 mL of the Protein sample i n f u l l tubes at the top p o s i t i o n of the gradient. Applying the correct amount of sample i s important to prevent "overloading" the gradient. The rotor tubes can be run f u l l or half f u l l , or b o t t l e s can be used i n place of tubes. SpinPro determines which option i s most appropriate. A number of parameters are affected by t h i s option, including the run time. Knowing where to load the sample i s important. Samples can be loaded at the top, middle, or bottom of gradients, or mixed homogeneously with them. At the end of the run the 16 S p a r t i c l e s w i l l be approximately 45% from the top of the gradient. In the rate-zonal method, common practice i s to have the component of i n t e r e s t at the 50% p o s i t i o n i n the gradient when the run i s over. SpinPro allows the f i n a l p o s i t i o n to be s p e c i f i e d , giving the investigator the opportunity to adjust the procedure so that components not of i n t e r e s t are widely separated from the component of i n t e r e s t . To process the e n t i r e sample volume requires approximately 2 centrifuge run(s) with an estimated t o t a l run time of 12 hours, 5 minutes. SpinPro determines how many runs are required to process the e n t i r e sample volume. The t o t a l run time i s estimated. When large sample volumes are involved, and thus many runs are required, the investigator can change the optimization c r i t e r i o n to "minimize number of runs" or "minimize cumulative run time" i n order to more e f f i c i e n t l y process the sample. Since two runs are required here, the investigator may want to select a larger rotor for use i n the Lab Plan.


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The Lab Plan Report The Lab Plan provides information s i m i l a r to that of the Optimal Plan except that there i s the additional constraint of using only the ultracentrifuges and rotors available i n the laboratory. This requirement can r e s u l t i n dramatic differences between the Optimal Plan and the Lab Plan. The run times can d i f f e r by hours, f o r example, or the p u r i t y of the separation can be s i g n i f i c a n t l y affected. A completely d i f f e r e n t gradient can be recommended as a function of the rotor selected from the lab. If there are no rotors i n the lab capable of doing the separation, SpinPro reports that the run cannot be done with the available rotors. The Plan Comparisons Report The Plan Comparisons report summarizes the differences between the plans i n terms of run time and number of runs required to process the sample (figure 3). In the figure the Optimal Plan uses the SW 55 T i rotor and the Lab Plan uses the SW 41 T i rotor. The d i f f e r e n t run times r e s u l t i n g from these rotors are compared on a percentage basis. A s i m i l a r comparison i s made f o r the t o t a l run time required to process the entire sample. Each of the rotors requires two runs to process the entire sample. The comparison of the t o t a l run times can help i n i d e n t i f y i n g the slower, but larger capacity, rotors that are more e f f i c i e n t f o r handling large sample volumes. I f warranted, SpinPro makes q u a l i t a t i v e comparisons between the two plans. Expert System Details SpinPro i s a t y p i c a l backward chaining, rule-based expert system. Rule-based systems are systems i n which the expert's knowledge i s encoded primarily i n the form of i f - t h e n r u l e s , i . e . , i f a set of conditions are found to be true then draw a conclusion or perform an a c t i o n . "Backward chaining" refers to the procedure f o r finding a s o l u t i o n to a problem. In a backward chaining system, the inference engine works backwards from a hypothesized solution to f i n d facts that support the hypothesis. Alternative hypotheses are t r i e d u n t i l one i s found that i s supported by the f a c t s . SpinPro's backward chaining inference engine i s c a l l e d "MP". "MP" has been developed by Beckman to support the development of expert systems. I t has several features that have been designed s p e c i f i c a l l y i n response to the requirements of the SpinPro project. Two of these requirements are that SpinPro run on an IBM PC/XT and that the program-user interface be advanced and easy to use. The report generator and the pop-up menu/mouse i n t e r a c t i o n provide the advanced user i n t e r f a c e . To be able to run the program on the IBM PC/XT and s t i l l address the u l t r a c e n t r i f u g a t i o n problem required the development of fact tables, "why responses", rule functions, rule groups, and "constraints". Development of these features has greatly improved the a b i l i t y of "MP" to make complex inferences. Some of these features are demonstrated i n the rule example of figure 4. The r u l e , one of approximately 800 rules i n SpinPro, i s assigned to the rule group 2-STEP.ISOPYCNIC.DNA.RULES. Only those r u l e s , i d e n t i f i e d by the rule group name and pertinent to the s o l u -



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t i o n of a p a r t i c u l a r problem, are applied to that problem. This breakdown of rules into rule groups i s one of the methods used to f a c i l i t a t e putting a complex expert system on a microcomputer with r e l a t i v e l y limited memory and processing power. The o v e r a l l e f f e c t of the rule i n figure 4 i s to s e l e c t , from a set of rotors, those rotors that are best for minimizing the run time when using the 2-step isopycnic method to separate DNA. The i n i t i a l set of rotors i s called USERS.MATCHED.ROTORS. The f i n a l set of rotors i s c a l l e d the MINIMIZE.RUN.TIME.ROTORS. The body of the rule applies tests to the i n i t i a l set of rotors and concludes that the rotors passing the tests are the MINIMIZE.RUN.TIME.ROTORS. In greater d e t a i l , Clause 1 of the rule tests the value of the parameter VERTICAL.TUBE.ROTORS. The value of this parameter t e l l s SpinPro whether v e r t i c a l tube rotors should be considered f o r the run. Often this can be deduced by SpinPro, but when i t can't, the question "Do you want to consider using v e r t i c a l tube rotors i n t h i s run" i s posed to the user. The parameter VERTICAL.TUBE.ROTORS has a set of properties that define i t s c h a r a c t e r i s t i c s including the prompt used to request the information, the "expect" property used to specify the acceptable responses to the prompt, and the "Why Response" property used i n response to the investigator's input of "Why?". If the value of VERTICAL.TUBE.ROTORS i s found to be true (or "yes") then clause 2 of the rule i s evaluated. The references to " f a c t " i n clause 2 cause the system to refer to a table that contains the facts for p a r t i c u l a r rotors. References to the f a c t s ROTOR.DESIGN, TUBE.VOLUME, and K.FACTOR are applications of p a r t i c u l a r constraints to the rotors. For example, two constraints are that the rotor must have a tube volume greater than 1 mL and a k factor less than 50. Clause 3 further pares the set of rotors on the basis of k factor by taking only the best rotor and any rotor with a k factor within 50% of the k factor of the best r o t o r . The Other Functions SpinPro includes two other functions that enhance i t s role as an expert advisor. This i s i n recognition that an expert provides more than expert advice. An u l t r a c e n t r i f u g a t i o n expert serves i n many r o l e s : a teacher of centrifugation p r i n c i p l e s , a describer of standard procedures, and a source of l i t e r a t u r e references. The INFORMATION function contains an extensive database of u l t r a c e n t r i f u g a t i o n information organized i n a h i e r a r c h i c a l fashion (Figure 5). The primary purpose of the INFORMATION function i s to provide an on-line reference to separation techniques, gradient materials, r o t o r s , tubes, and centrifuges. For example, INFORMATION can be used to get information on the Type 70.1 T i rotor, the c o m p a t i b i l i t y of polyallomer tubes with c e r t a i n chemicals, a description of rate-zonal separations, and references to isopycnic methods. The subjects i n the information hierarchy can be expanded to give a more detailed breakdown of the subject. For example, expanding the "Fixed Angle" subject y i e l d s a d e t a i l e d breakdown of the f i x e d angle rotors. The investigator could now select one of the rotor names on the screen and get information about that r o t o r . The INFORMATION function includes the subject "SpinPro", which i s a complete on-line manual of the SpinPro system.


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RULE 2667: (Rulegroup: 2-STEP.ISOPYCNIC.DNA.RULES) If: 1) V E R T I C A L . T U B E . R O T O R S , and 2) Find all instances of T H A T . R O T O R among the value of such that:

USERS.MATCHED.ROTORS

1) the ROTOR.DESIGN fact of T H A T . R O T O R = one of:

SWINGING.BUCKET,

FIXED.ANGLE, or VERTICAL.TUBE, and


2) the T U B E . V O L U M E fact of T H A T . R O T O R > 1, and 3) the K . F A C T O R fact of T H A T . R O T O R < = 50 (saving those in C O L L E C T E D . R O T O R S ) , and 3) Find all instances of T H A T . R O T O R among C O L L E C T E D . R O T O R S for which: the K . F A C T O R fact of T H A T . R O T O R is within 50% of the smallest value so computed (saving those in C O L L E C T E D . R O T O R S ) Then: 1) Conclude that MINIMIZE.RUN.TIME.ROTORS

is each of C O L L E C T E D . R O T O R S .

Figure 4. A rule that selects rotors to minimize the run time i n a plasmid DNA separation. The r u l e examines a set of rotors c a l l e d USERS.MATCHED.ROTORS, s e l e c t i n g those rotors that s a t i s f y c r i t e r i a based on the rotor design, tube volume, and k f a c t o r .

=

SpinPro = Information Top Level

Fixed Angle^ Vertical Tube

==

Sample Materials and Particles Separation Methods

=

Swinging Bucket

=

Tubes and Bottles

=

Continuous Flow

=

Ultracentrifuge Rotors

=

Zonal

=

Ultracentrifuges

=

Glossary

=

Table of Rotors by Use Accessories

===== Rotor Maintenance ===== Rotor Warranties Point at a n Information Item a n d c l i c k a n y m o u s e b u t t o n f o r O p t i o n s M e n u .

Figure 5 . The information hierarchy of SpinPro showing the categories of information a v a i l a b l e . The positions i n the h i e r archy can be expanded to give a more d e t a i l e d breakdown of each subject.


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The CALCULATION function provides a variety of routine c a l c u l a tions performed i n most u l t r a c e n t r i f u g a t i o n laboratories. Included are d i l u t i o n calculations f o r sucrose, a p e l l e t i n g time c a l c u l a t i o n , and a calculation for determining rotor speed reductions f o r s a l t gradients. As with the INFORMATION function, the CALCULATION funct i o n i s a support tool i n the e f f o r t to e f f i c i e n t l y design and carry out a separation.


Development of SpinPro There i s much concern about the length of time required to develop expert systems, p a r t i c u l a r l y since so many have achieved various stages of prototype, but few have been completed. Our experience with SpinPro has led to many i n s i g h t s , more than can be f u l l y d i s cussed here. Nevertheless, a few major points are worth mentioning. It i s not p a r t i c u l a r l y clear to us why SpinPro has succeeded i n achieving product status and other expert systems have not, although we suspect that an early decision to produce a product rather than to do AI research has been important. The problem domain of u l t r a centrifugation appears to have been a good choice. The domain has proven to be f a i r l y well bounded, even though the 800 rules required has exceeded early estimates by a factor of four. When considering the various stages of prototyping, debugging, and refinement, over 25,000 rules have been w r i t t e n , and tossed out. Perseverance, sustained by having a concrete goal of "completeness" rather than a more indeterminate goal of "demonstrating f e a s i b i l i t y " or "prototyping", was c r u c i a l to the success of the project. In some ways expert systems programming i s l i t t l e d i f f e r e n t from more " t r a d i t i o n a l " programming. For example, s i m i l a r to most software programs, about 50% of the code i n SpinPro i s f o r the user i n t e r f a c e ; debugging has been very time consuming; and miscommunication was the source of a great deal of additional e f f o r t . Since these problems are a part of t r a d i t i o n a l programming as w e l l , techniques designed to a s s i s t t r a d i t i o n a l programmers, such as organi z a t i o n p r i n c i p l e s , s p e c i f i c a t i o n , and e f f e c t i v e communication also apply to expert systems. In other ways expert systems programming i s much d i f f e r e n t . T r a d i t i o n a l p r i n c i p l e s of s p e c i f i c a t i o n and organization are tested, i n part, because the program undergoes evolutionary and sometimes revolutionary revisions as an understanding of the problem domain grows. Despite early detailed s p e c i f i c a t i o n , the tendency of the s p e c i f i c a t i o n and the project to evolve toward i t s f i n a l d e f i n i t i o n seems to be unavoidable. From i t s inception to completion, the development of SpinPro has taken about s i x person years. The development team has included a manager, two knowledge engineers, one primary expert, four experts for review, and two people responsible f o r the content of the INFORMATION function. During this time, we have completed the following major a c t i v i t i e s : 1. 2. 3. 4. 5.

s p e c i f i c a t i o n and prototyping knowledge a c q u i s i t i o n from the expert knowledge coding into rules and debugging of rules design and implementation of the "MP" inference engine design and implementation of the user interface



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6. c o l l e c t i n g and w r i t i n g the contents of the INFORMATION function 7. converting from Interlisp-D on the Xerox 1108 AI workstation to Gold H i l l Common Lisp (GCLISP) on the IBM PC/XT Of these a c t i v i t i e s , task 3 (knowledge coding) and task 4 (inference engine) were the major e f f o r t s . Knowledge coding and debugging required at least f i v e times as much e f f o r t as task 2, the knowledge a c q u i s i t i o n from the expert. Task 7, converting from the development environment to the product proved to be one of the major hurdles. There are two notable AI enhancements that are not a part of SpinPro. F i r s t , the "MP" inference engine does not include uncert a i n t y reasoning. The problem domain has only a l i m i t e d use f o r i t , and where i t i s required, uncertainty i s handled w i t h i n the c a p a b i l i t i e s of "MP". Second, "MP" does not include an a b i l i t y to explain i t s reasoning beyond the "Why?" function discussed e a r l i e r . An explanation c a p a b i l i t y was not implemented because the usual form of presenting a trace of the rules that have f i r e d i s inadequate and p o t e n t i a l l y confusing to the user. Why? Because rules t y p i c a l l y encode "shallow" knowledge (the expert's experience and rules of thumb) and i n a rule trace, are inadequate for communicating the r e a l , "deep" knowledge, reasons for making a decision. SpinPro and the Expert How does SpinPro compare to the expert i n solving u l t r a c e n t r i f u g a t i o n problems? For most problems, SpinPro designs procedures as good as the expert, i f not better. The inherent c a p a b i l i t i e s of computers are responsible f o r this achievement; they are consistent, they don't forget, and they are precise. For example, SpinPro contains a vast amount of knowledge that i s not a part of the expert's active memory. Many of the rules are an integration of the expert's knowledge and procedures reported i n the l i t e r a t u r e . Other rules are derived from l i t e r a t u r e references only. This vast amount of knowledge i s immediately available to SpinPro, but not to the expert. For the new problems, the ones never described to SpinPro, the expert i s f a r superior. The expert has i n t e l l i g e n c e , c r e a t i v i t y , common sense, and an understanding of the p r i n c i p l e s of u l t r a c e n t r i f u g a t i o n . These are human tools that the expert can bring to bear on new problems. At t h i s stage i n AI a p p l i c a t i o n s , and despite the goal of AI to recreate these human a b i l i t i e s , SpinPro, l i k e other expert systems, i s lacking. From the SpinPro project emerged a strong SpinPro-expert r e l a t i o n s h i p . Early i n the project the expert was doubtful about the prospects of capturing years of education and experience i n a software program. Also the expert f e l t threatened by the expectat i o n that h i s role would be subsumed by a computer. These problems soon disappeared as the challenge of creating SpinPro became more important. As the project neared completion, the expert took personal r e s p o n s i b i l i t y f o r the accuracy of SpinPro and pride i n i t s l e v e l of achievement. SpinPro's future development remains c l o s e l y t i e d to the expert. SpinPro required that the expert c r i t i c a l l y review the science of u l t r a c e n t r i f u g a t i o n and h i s knowledge of i t . For example, SpinPro sometimes designed a procedure using a rotor that was not



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expected or recommended an exceptionally short run time that was shorter than thought possible. These procedures required careful review. Sometimes they were accepted as v a l i d improvements to e x i s t i n g procedures. Isopycnic runs are one example, where SpinPro found that procedures t y p i c a l l y requiring 12-16 hours, could be run for 7-9 hours with the same r e s u l t s . Thus, SpinPro i s i n d i r e c t l y responsible f o r advancing the expert's understanding of u l t r a c e n t r i fugation and f o r improving u l t r a c e n t r i f u g a t i o n techniques. SpinPro promoted a degree of rigorousness that had never before been applied to u l t r a c e n t r i f u g a t i o n . Updates to SpinPro continue as new rotors and new techniques are developed or as inadequacies are found. New expert systems techniques, such as the a b i l i t y to incorporate the p r i n c i p l e s of a problem domain, rather than just the experience of the expert, should give SpinPro the a b i l i t y to design procedures for novel problems and to explain i t s reasoning. The updates insure that SpinPro w i l l be a repository of knowledge about the current state of u l t r a c e n t r i f u g a t i o n ; SpinPro's expertise should continue to improve. Furthermore, the expert remains g a i n f u l l y employed as a f i n a l a r b i t r a t o r on the i n c l u s i o n or exclusion of any new knowledge. Conclusion The SpinPro U l t r a c e n t r i f u g a t i o n Expert System provides an integrated package of expert advice, information, and c a l c u l a t i o n functions. Its purpose i s to allow investigators to f u l l y exploit the c a p a b i l i t i e s of u l t r a c e n t r i f u g a t i o n , thereby improving the e f f i c i e n c y of the u l t r a c e n t r i f u g a t i o n laboratory. I t uses AI techniques to provide the a b i l i t y to advise on the best selection of run parameters that s a t i s f y the investigator's requirements. Our experience with SpinPro has shown that i t e f f e c t i v e l y performs the role of an expert advisor: designing e f f i c i e n t u l t r a c e n t r i f u g a t i o n procedures that can reduce run times and improve the q u a l i t y of separations. Acknowledgments For t h e i r contributions to the SpinPro Ultracentrifugation Expert System, the authors thank Gertrude Burguieres, Mike Brown, P h y l l i s Browning, Marsha Chase, Judy Cummings, Manny Gordon, Mary Jane MacDwyer, Edna Podhayny and Bruce Wintrode. R E C E I V E D January 14,

1986


Artificial Intelligence Applications in Chemistry - American Chemical

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