Manthan Automated Analysis

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M.Pharm -2nd sem mpa


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Content:- Introduction Need for automation Automated device Objective of automation Classification of automatic analysers Automation in various methods of analysis Automation approach in analysis Features and advantages of automation systems Readout devices and computers Summary Reference 2


Introduction IUPAC define automation as “the use of combinations of mechanical and instrumental devices to replace, refine, extend or supplement human effort and facilities in the performance of a given process, in which at least one major operation is controlled without human intervention by a feedback mechanism .” Automation may involve operation like the preparation of samples, the measurements of responses, and the calculation of results. 3

“Complete automation will lead to human prohibition in pharma industry” :

“Complete automation will lead to human prohibition in pharma industry” 4

Need for automation.:

Need for automation. The partial or complete replacement of human participation in laboratory process Increasingly stricter control of growing number of samples in which a large number of analytes are to be determined at increasingly low concentration Cost reduction In research for increased accuracy, precision, and productivity, the pharmaceutical analyst seeks to select optimal measurement and to automate as appropriate 5

Objective of Automation:

Objective of Automation Automation is used for: - Facilitating an analytical technique or method Processing of large number odd samples Determination of several components in the same sample Reduction of human participation to Avoid error Cut costs Increasing sample throughout Process (industrial or otherwise ) control Lowering consumption of sample and/or reagent(s) Samples , occasionally dealt with in large number or valuable to deal with manually Analytes , which are sometime present in very dissimilar or low concentration in sample. 6

Objective of Automation:

Objective of Automation Reagents , some of which are rare or expensive even unstable. Rapidity , frequently essential in large laboratories such as those in hospitals, urgently requiring the analytical result, and of industrial and other laboratories require in constant availability of data for process controls. Economy , in personnel and material expenditure. Precision , closely related to the elimination of both definite and indefinite errors arising from the so-called ‘human factor’ Data generation , some analytical technique are based on the acquisition of a large number do data, especially in the drug discovery and development stages. 7


ADVANTAGES OF AUTOMATIC ANALYSES Automated instruments offer a major economic advantage because of their savings in labor costs . Their speed , which is frequently significantly greater than that of manual devices. So, the number of determination per day can be much higher than with manual methods. A well-designed analyzer can usually produce more reproducible results over a long period of time than can an operator employing a manual instrument. The ability to process samples in situation that would be dangerous for humans. 8

Automated device :

Automated device They are defined as those encasing automation. They are conceived to make decisions with the aid of feedback system, without human intervention. There is a different operation sequence for each situation (sample).Some system are self-monitoring and self-adjusting ,have greater independence than automatic devices and are sometimes called ‘ completely automatic ’. 9

Classification of automatic analysers::

Classification of automatic analysers: According to the degree of automation Automatic semi-automatic According to the way in which samples and reagents are transported Batch (discrete) Continuous Segmented Unsegmented Robotics According to the number of analytes per sample One parameter Multi-parameter 10

PowerPoint Presentation:

According to flexibility Specific Flexible According to source Commercial Hand-made According to the state of aggregation of sample Gas analyser Liquid analyser Solid analyser According to sample frequency One-off Periodic Continuous 11


TYPES OF AUTOMATED ANALYSIS SYSTEMS Automatic analytical systems are of two general types: Discrete analyzers Continuous-flow analyzers 12

1) Discrete analyzers :

1) Discrete analyzers In this , individual samples are maintained as separate entities and kept in separate vessels throughout each unit operation. The system has many moving parts. Ex. Discrete automatic analyzer Reagents are combine with sample in discrete cuvettes where mixing, incubation and final color measurement occurs. These samples may be analyzed sequentially or in parallel. 13

Advantage: :

Advantage: Cross contamination among samples is totally eliminated . Inexpensive and reliable Changeover from one method to the next is automatic . Less operator training Multiple tests on each sample 14

2) Continuous-flow analyzers.. :

2) Continuous-flow analyzers.. In this , the sample becomes a part of a flowing stream where several of the steps take place. Ex . Flow injection analysis Here, interaction among samples are always concern. So, special precaution are required to minimize sample contamination. 15

Continuous-Flow Analysis:

Continuous-Flow Analysis There are generally two kind of analysis Segmented flow analysis Developed by Skeggs in 1957. Samples are carried through the system to a detector by flowing aqueous solution that contain closely spaced air bubbles. 16

Segmented flow analysis…:

Segmented flow analysis… It includes a peristaltic pump that continuously aspirates sample and reagent, a variable no. of tubes constituting a manifold to circulate liquid and a detector system. Aspirated sample are segmented by injecting air bubbles that should be remove before they can be reaching to the detector. At detector air bubble are removed and thus each sample is separated by washing solution. Thus a square shaped detector response is obtained, the height of rectangle is directly proportional to conc. of analyte . 17

Unsegmented flow analysis :

Unsegmented flow analysis FLOW-INJECTION ANALYSIS... Flow-injection methods are an outgrowth of segmented-flow procedures , which were widely used in clinical laboratories in the 1960s and 1970s for automatic routine determination of a variety of species in blood and urine samples for medical diagnostic purposes. The absence of air bubbles imparts several advantages like higher analysis rates , enhanced response times. 18


Instrumentation... In this, a peristaltic pump moves colorimetric reagent directly into a valve that permits injection of samples into the flowing stream. The sample and reagent then pass through a 50 cm reactor coil where the reagent diffuses into the sample plug and produces a colored product by sequence of reactions. From the reactor coil, the solution passes into a flow-through photometer and the signal output from this system for a series of standards . 19

Sample and reagent transport system:

Sample and reagent transport system Ordinarily, the solution in a flow-injection analysis is moved through the system by a peristaltic pump, a device in which a fluid (liquid or gas) is squeezed through plastic tubing by rollers. 20

Sample injectors and detectors:

Sample injectors and detectors The injectors and detectors employed in flow-injection analysis are similar in kind and performance requirements to those used in HPLC . Sample size for flow injection procedure ranges from 1 μ L to 200 μ L. For successful analysis, injectors must not disturb the flow of the carrier system. The most common detectors in flow injection are spectrophotometer, photometer and fluorometer . 21

Separations in FIA:

Separations in FIA Separations by dialysis, by liquid/liquid extraction, and by gaseous diffusion are readily carried out automatically with flow-injection systems. Dialysis and gas diffusion Extraction 22

1) Dialysis and gas diffusion:

1) Dialysis and gas diffusion Dialysis is often used in continuous-flow methods to separate inorganic ions , such as chloride or sodium or small organic molecules, such as glucose, from high-molecular-weight species such as proteins. It is used for determination of ions and small molecules in whole blood stream or serum . Gas diffusion from a donor stream containing a gaseous analyte to an acceptor stream containing reagent that permits its determination. 23

2) Extraction:

2) Extraction Another common separation technique readily adapted to continuous-flow methods is extraction . Ex. A system for the colorimetric determination of an inorganic cation by extracting an aqueous solution of the sample with chloroform containing a complexing agent such as 8-hydroxyquinoline. It is important to reiterate that none of the separation procedures in FIA methods are complete. 24

Principles of flow-injection analysis :

Principles of flow-injection analysis After injection with a sampling valve, it moves through tubing, so band broadening or dispersion takes place. In this, convection arising from laminar flow and creating a parabolic front. Diffusion also causes band broadening . Two types of diffusion occur: radial ( perpendicular the flow direction) and longitudinal(parallel to the flow direction). At low flow rate, radial diffusion is the major source of dispersion.So , dispersion by both convection and radial diffusion occur. 25


Dispersion Dispersion D is defined by the equation D = c o /c where , c o = analyte concentration of the injected sample and c = peak concentration at the detector. Dispersion is influenced by three interrelated and controllable variables: sample volume, tube length, and pumping rate. 26

Applications of flow-injection analysis:

Applications of flow-injection analysis 1) Low-dispersion applications: Low-dispersion flow-injection techniques have found considerable application for high-speed feeding of such detector systems as flame atomic absorption and emission as well as inductively coupled plasma. 2)stopped flow methods: It is used for kinetic measurement. 3)flow injection titration: Titration can also be performed continuously in a flow injection apparatus. 27

Automation Approach in Analysis: :

Automation Approach in Analysis: A serial (or sequential ) automation process A parallel automation process 28

A serial (or sequential ) automation process:

A serial (or sequential ) automation process The general characteristic of a serial automation process is that a given determination reaches completion before the next determination begins, although with some serial automation systems, the second determination may be started before the first is completed Serial automation could also be configured in an on-line arrangement, where tandem processes in the chemical determination are performed by collection from a flowing stream. 29

PowerPoint Presentation:

Such an on-line system could be used to acquire a sample, process it, and Perform the associated chemical measurement step Eg . in the case of on- line microdialysis /LC/MS or on-line solid-phase extraction/LC/MS/MS 30

PowerPoint Presentation:

Schematic representation of a generalized serial automation system, including on-line sampling, optional sample concentration/preparation (C1), optional sample separation step (C2), detection (D), and data system (DS) for acquisition and control. Here, W indicates waste. 31

PowerPoint Presentation:

Serial automation system based on a single powerful robotic arm. The arm in this example is reticulated and is capable of traversing along a 6-foot monorail to access a wide array of laboratory apparatus and analytical instrumentation. Despite the complexity of this automation system, the overall throughput of the system is generally limited by the serial processing of the robotic arm. 32

Parallel Automation Process :

Parallel Automation Process Parallel Automation Process for chemical analysis is, simply, a process where more than one automated chemical manipulation is performed simultaneously . These parallel manipulations can encompass some or all of the following discrete analytical chemistry operations: Experiment initiation, Sampling (obtaining the samples), Sample preparation, Component separation, Analyte detection and data reduction/reporting 33

Micro scale Liquid–Liquid Extraction on a 96-well liquid handling workstation.:

Micro scale Liquid–Liquid Extraction on a 96-well liquid handling workstation. The procedure is implemented on the six-position stage of a Tomtec Quadra-96 or 384-well liquid handling workstation. Samples plus standards are placed in a 96-well tube plate, which is located in position 2 of the workstation stage, and to this plate are added an internal standard (position 6), a buffer (position 5), and an organic solvent (position 4), using the parallel liquid handling capabilities of the 96-well head. After off-line vortexing and centrifugation, a phase separation allows the organic solvent extract to be transferred to a clean plate located in stage position 1. These extracts are then evaporated to dryness and reconstituted without having ever left the 96-well format. 34

PowerPoint Presentation:

Tomtec Quadra-96 liquid handling workstation. Conceptualized semiautomated 96-well liquid–liquid extraction procedure. 35

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Tomtec Quadra-96 liquid handling workstation. Conceptualized semiautomated 96-well liquid–liquid extraction procedure. 36

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Zymark combinatorial chemistry workstation 37

Hybrid Automation Systems:

Hybrid Automation Systems Hybrid automation systems for chemical analysis combine features of both serial and parallel systems. Some processes are conducted in serial, some are conducted in parallel, and all are integrated into a single system Hybrid Automation = Serial Automation + Parallel Automation. 38

PowerPoint Presentation:

Hybrid Automation Systems In Robotic solid-phase extraction method development system, a central robotic arm (XP) which service various arm peripherals, and a 144-port vacuum manifold, used for solid-phase extraction method development. The system and development of the solid-phase extraction columns, which are in parallel. Although a fully parallel system would have been desirable, it was beyond the capabilities of commercially available system at this time. 39

PowerPoint Presentation:

Robotic solid-phase extraction method development system MLS1 or MLS2 master lab station, SV switching valve,W waste, R1, R2, or R3 rack. No. 1, 2, or 3 S1 or S2 solvent reservoir H1, H2, H3, H4 hand No. 1, 2, 3, or 4 CS capper station TW tip waste, VM vacuum manifold, XP zymate XP robotic arm, PEC1 or PEC2 power and event controller 1 or 2, PT pressure transducer, VS vortex station, TB tip blower, MP metering pump. 40

Selection of automation approach:

Selection of automation approach If off-line batch processing were necessary, parallel automation would be appropriate. In contrast, if time-dependen t sampling of a single experiment or a few samples were in order, such as in-process control sampling, then serial automation would be appropriate. Also, if one-time-use components or difficult -to-create components , such as Affinity chromatography columns or Immobilized automated membrane (IAM) columns , must be employed, then it could be prudent to select a serial on-line automation system. If, however, postprocess characterization of a batch of samples is required, then parallel automation processing is most appropriate. 41

Automated Method in P’Copoeia: :

Automated Method in P’Copoeia : Automated method have been found especially useful in testing the content uniformity of tablets and capsules and in facilitating methods requiring precisely controlled experimental conditions . In addition,the detection system and calculation of results for automated methods are often computerized. Before , an automated method for testing an article is adopted as an alternative, it is advisable to ascertain that the results obtained by this system are equivalent in accuracy and precision to those obtained by the prescribed p’copoeial method. Because of their versatility, this system designed for rapid determination of specified substance often can be readily modified by the addition of suitable modules and accessories to permit the determination of one or more additional substance in dosage form. the automated analysis of articles containing both estrogen & progestogens . 42


FEATURES AND ADVANTAGES OF AUTOMATION SYSTEMS Speed . Complexity . Hardware . Redundancy of hardware Software . Expandability . Unattended operation . Error recovery . 43

Readout Devices And Computers :

Readout Devices And Computers The simplest, is watching a meter or counter and recording the values for later calculation. Next comes the use of Heyrovsky’s invention, the moving pen recorder. collect the data unattended interpret the data interpreted the chart scale. Digital printout Once the readings are available, mathematical manipulations are required and performed & report by automation. A few systems use precalibrated charts and the results can be read off the charts directly. Most direct concentration readouts depend on the system obeying Beer’s law. Results are usually printed out in digital form on a paper tape. 44

PowerPoint Presentation:

The “ off-line ” data acquisition system in which some form of computer readable record is made during analysis time. This might be punched card, punched tape, analog magnetic tape, or digital magnetic tape. These records are subsequently calculated on a remote computer and reported as hard copy to the appropriate submitter. The data can also be easily stored in machine-retrievable form for future reference. In “ on- line .” case, the computer is fed the raw data directly. With essentially immediate access to calibration data and possibly positive sample identification, the computer can identify and calculate the results as soon as the pertinent data has been generated. Again, a hard copy can be made and results can be stored in machine-retrievable form for future reference. 45

PowerPoint Presentation:

Examples of automation in pharmaceutical profiling Property Method Example automation(vendor) Automation value added Integrity LC/UV/MS Autosampler /HPLC/MS(Agilent 1100/MSD)( WatersAquity /Micro) Large sample sets, unattended, integration and software control Permeability PAMPA Evolution (pION) Genesis(Tecan) Multiprobe (Packard) UV Plate Reader Spectra Max (MolecularDevices) Liquid handling and HT analysis, integrated system CellMonolayer (Caco-2,MDCK) Genesis,Incubator,Multiprobe, LC/MS/MS(WatersMicro) Multiple time points, analyze large number Samples from one experiment IAM Autosampler/HPLC Capacity to handle many samples Solubility Turbidimetry NEPHELO star (BMG) Genesis, Multiprobe Solubility Scanner (BDGentest) Read plate in minutes;integratedsystem Read plate in minutes; integrated system DirectUV Evolution( pION ) Integrate the whole experiment and Calculate from model 46

PowerPoint Presentation:

Stability Microsomalstability Genesis, Multiprobe Incubator (JitterBug) LC/MS/MS Autosampler(LEAP) HT sample processing; quantitation S9,Hepatocytes Same CYPInhibition rhCYPisozyme Genesis Incubator Fluo Star Fluorescence Plate Reader (BMG), rhCYP isozymes and fluorescent probes ( BDGentest ) Liquid handling for whole experiment; read plate in minutes Humanlivermicrosomes &LC/MS GenesisLC/MS/MS Robotic sample prep.HT sample analysis LogP Octanol/Water partition LDA(SiriusAnalytical) Integrated system and well developed LogP assay model Capillary Electrophoresis cePRO(CombiSep) 96 parallel CE tracks accelerate analysis pKa Spectral Gradient Analysis SGA(SiriusAnalytical) Integrated system and well developed pKa assay model Capillary Electrophoresis cePRO(Combisep) 96 parallel CE tracks accelerate analysis 47


SUMMARY Due to the on going demand for speed in industrial chemical analysis , parallel automation processes are beginning to replace serial automation processes, especially in those areas where high numbers of assays or samples are expected. For in situ or on-line situations where sparse strategic sampling is the norm, serial automation will likely play a continued role. Expensive robotic arms, while powerful and flexible, do not directly lend themselves to parallel automation systems unless the effort is made to combine them with specialized multichannel hardware. Such hybrid systems are powerful and useful but are extremely complicated to implement and maintain. In the future, it seems likely that automated systems will be miniaturized and that this miniaturized format will lend itself readily to increased use of automation. 48


References Norbert R.Kuzel , Harold E.Roudebush & Charles E.S,(1969),”Automation techniques in pharmaceutical analysis”, Journal of pharmaceutical sciences, vol.58, no.4, 381-406. Edward H.Kerns,Li DI,(2005) “Automation in pharmaceutical profiling” ,Chemical and Screening Science, Wyeth Research,Princeton,NJ,114-123. Berridge J.C,(1990), “Advance in automation of pharmaceutical analysis”, Analytical chemistry department, Pfizer central research, vol.7,No.12, pp.1313-1321. Douglas A. skoog, F. james Holler, Stanley R. Crouch; insrumental analysis; cengage learning india private limited; new delhi; first edition;2007;1015-1036. QUE. Discuss on automated analysis. 49

PowerPoint Presentation:

50 “One machine can do the work of fifty ordinary men. No machine can do the work of one extraordinary man” Thank You For Your Attention.

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