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High pressure liquid chromatography in coatings analysis.

High pressure liquid chromatography (HPLC HPLC high-performance liquid chromatography.


high performance liquid chromatography.

HPLC High-performance liquid chromatography Lab instrumentation A highly sensitive analytic method in which analytes are placed
) is an established standard analytical tool for the analysis of coatings and related materials. An extensive body of literature has been published in various analytical and related fields' journals on HPLC methods. These methods have been developed and used in the determination of small to very large molecules ranging from small organic impurities to polymer resins. In addition, fundamentals of HPLC and related methods have also been extensively covered in the literature and the reader is encouraged to review them as needed. The objective of this article focuses mainly on the applications of HPLC and gel permeation chromatography Gel permeation chromatography (GPC) is a separation technique based on hydrodynamic volume (size in solution). Molecules are separated from one another based on differences in molecular size. This technique is often used for polymer molecular weight determination.  (GPC (1) A PC that uses the Linux-based gOS operating system. See gOS.

(2) (GPC Group) Originally the Graphics Performance Characterization committee of the NCGA, the GPC Group is now part of Standard Performance Evaluation Corporation (SPEC) and oversees the following
) methods in the analysis of coatings.



Liquid chromatography (LC) is probably the most commonly used method of separation and analysis of chemical compounds and ions in solution. Increased understanding of liquid chromatography since its invention in 1906 has led to fascinating and dramatic development and usage of the technique in a variety of applications. (1-7) For example, instrumental advances, automation, and miniaturization min·i·a·tur·ize  
tr.v. min·i·a·tur·ized, min·i·a·tur·iz·ing, min·i·a·tur·iz·es
To plan or make on a greatly reduced scale.

 of the technique has brought about the development of significant methods utilizing complex separation mechanisms, such as electrophoresis, supercritical fluid chromatography Supercritical Fluid Chromatography (SFC) is a robust and easy-to-use form of normal phase chromatography ideally suited to the analysis and purification of low to moderate molecular weight, thermally labile molecules. It is especially suited to the separation of chiral compounds. , microscale separations, and numerous powerful hyphenated hy·phen·at·ed  
1. Having a hyphen: a hyphenated adjective.

2. Often Offensive Of or relating to naturalized citizens or their descendants or culture.
 techniques such as online and off-line liquid chromatography/nuclear magnetic resonance (LC-NMR), and very efficient multi-detector analysis systems for extremely complex analyses.

High pressure liquid chromatography (HPLC) is an analytical technique used for separation of low-to-moderate molecular weight compounds of resins. The instrumentation for HPLC and size exclusion (SEC) or gel-permeation chromatography (GPC) is similar, but the columns differ. A schematic diagram of HPLC components is shown in Figure 1.

HPLC components consist of the following: (a) solvent and/or solvent mixture, (b) solvent delivery system (pumps and controller), (c) sample delivery system (manual or automatic injection), (d) a column suitable for the separation of interest, (e) a detector system, and (f) a data system.



There are numerous types of separation mechanisms in liquid chromatography. The most important ones include: affinity chromatography, ion chromatography, capillary electrophoresis, normal phase chromatography, reverse phase chromatography, supercritical Adj. 1. supercritical - (especially of fissionable material) able to sustain a chain reaction in such a manner that the rate of reaction increases
critical - at or of a point at which a property or phenomenon suffers an abrupt change especially having enough mass
 chromatography, and microscale chromatography.

Affinity Chromatography

Affinity chromatography is a technique that uses like-like interactions for effective separation and analysis of sample components in solution. The column is designed to retain specific or similar analytes while allowing everything else through the column. The retained species can then be selectively eluted from the column by appropriate solvent. A schematic example of affinity chromatography is shown in Figure 2.

Ion Chromatography

Ion chromatography is a liquid chromatography method that can be used for the separation and quantification of anions and cations. For example, anions and cations that be separated and quantified are given in Table 1. A schematic anion anion (ăn`ī'ən), atom or group of atoms carrying a negative charge. The charge results because there are more electrons than protons in the anion.  separation and identification is shown in Figure 3.

In the context of this article, ionic chromatography is used in the determination of acid species in the acid/thermal decomposition of polymeric materials and ionic contamination in polymers and paints.

Capillary Electrophoresis

The capillary electrophoresis technique is based on the migration of charged molecules in an electrical field. The units are composed of a power source with a voltmeter and voltage regulator, an electrophoresis tank that holds the electrophoresis buffer, an anode anode (ăn`ōd), electrode through which current enters an electric device. In electrolysis, it is the positive electrode in the electrolytic cell.

Terminal or electrode from which electrons leave a system.
 and a cathode connected to the power source, a glass plate that holds the gel which is submerged into the electrophoresis buffer, and a comb sampler. These units are based on the simple relationship between the applied electric field (V), applied current (I), and resistance (R)--the Ohm's law:

V = I R

The applied electric field is measured in volts (V) and is proportional to the current (I) measured in milliamps. For a given voltage, a constant amount of current flows through the circuit and the voltage decreases across any element as its resistance increases. Molecules which have electrical charge due to groups capable of dissociating electrolytically (for example proteins), will migrate in an electrical field with a certain velocity and direction, which will depend on their electrical mobility. It has been shown that electrical mobility of a molecule is dependent on a variety of factors, such as electrical charge, coefficient of friction coefficient of friction
n. pl. coefficients of friction
The ratio of the force that maintains contact between an object and a surface and the frictional force that resists the motion of the object.
, shape, and molecular weight.


While capillary electrophoresis is a very important tool in the separation and identification of bio-molecules, it also finds important application in polymer analysis. (8)

Normal Phase Liquid Chromatography

Normal phase liquid chromatography (NPLC NPLC National Private Leased Circuit
NPLC Number of Power Line Cycles
NPLC National Pedigree Livestock Council
) a powerful separation technique, relies on the usage of polar stationary phase and nonpolar nonpolar

not having poles; not exhibiting dipole characteristics.
 liquid mobile phases for the separation of polar species. See the Type of Column Phases section below.

Reverse Phase Liquid Chromatography

Reverse phase liquid chromatography (RPLC RPLC Reverse-Phase Liquid Chromatography ) is the most popular method of liquid chromatography. The name is derived from the reverse elution elution /elu·tion/ (e-loo´shun) in chemistry, separation of material by washing; the process of pulverizing substances and mixing them with water in order to separate the heavier constituents, which settle out in solution, from the  order of most solutes when compared to NPLC. It relies on the usage of nonpolar stationary phases with polar mobile phases. Typical mobile phases may include water with added polar organic solvent, such as acetonitrile acetonitrile /ac·e·to·ni·trile/ (as?e-to-ni´tril) a colorless liquid with an etherlike odor used as an extractant, solvent, and intermediate; ingestion or inhalation yields cyanide as a metabolic product. . See the Type of Column Phases section.

Supercritical Fluid Chromatography

Supercritical fluid chromatography (SFC SFC
sergeant first class
) is a technique that uses mobile phases as either gases or liquids taken above their critical point. The advantages of this approach include the solvating power of liquid mobile phases and diffusion coefficients closer to gases. The lower viscosity of SFC allows for higher flow rates compared to LC, and the solvating power can be adjusted by changing the pressure. A supercritical fluid chromatograph chromatograph /chro·mato·graph/ (kro-mat´o-graf)
1. the apparatus used in chromatography.

2. to analyze by chromatography.


1. to analyze by chromatography.

 consists of a gas supply (C[O.sub.2]), a pump, a column in a temperature controlled oven, a restrictor to maintain high pressure in the column, and a detector. The flame ionization detector A flame ionization detector (FID) is a type of detector used in gas chromatography. Principle
The Flame Ionization Detector (FID) is one of the many methods by which to analyze materials coming off of gas chromatography column.
 (FID) is most common, but other GC or LC detectors can also be used. SFC finds many applications in polymer additives analysis. (9)


Microscale Liquid Chromatography

Microscale liquid chromatography is simply the miniaturization of liquid chromatography technique. The main advantages are the significant reduction of solvents, shorter analysis times, and overall improved sensitivities.


There are two main types of phase separation: (1) the normal phase HPLC which consists of a polar column material and relatively nonpolar solvents, and (2) reversed phase HPLC which consists of a nonpolar column material and relatively polar solvents.


Reversed phase HPLC is the preferred type of HPLC as it has proven to be the best for analysis of many types of resins. The most common column packing materials are nonpolar hydrocarbons, [C.sub.8], [C.sub.18] and phenyl phenyl (fĕn`əl), C6H5, organic free radical or alkyl group derived from benzene by removing one hydrogen atom.  bonded phases columns. The most common polar solvents for resins include acetonitrile (ACN ACN Accenture (stock symbol)
ACN Accenture
ACN Australian Company Number
ACN Automatic Collision Notification (US DOT)
ACN Acetonitrile
ACN Anglican Communion Network
)/water, methanol/water, and tetrahydrofuran tetrahydrofuran: see furfural.  (THF THF tetrahydrofolic acid.


tetrahydrofolic acid.
)/water. They are normally used in the gradient elution mode because they allow one to start with low content of organic component in the eluent eluent

the solution used in elution.
 (ACN or THF) then the strongly retained components will move faster for improved resolution. Performance of gradient elution is strongly dependent on the instrumentation. Solubility of the material to be separated is a very important consideration in HPLC. For example, ACN/water is good for low molecular weight resins and is limited by solubility in higher molecular weight resins. THF/water is better and sometimes a mixture of ACN/THF/water may be the best for analyzing low to high molecular weight resins.


Samples are injected into the HPLC via an injection port. The injection port of an HPLC commonly consists of an injection valve and the sample loop. The sample is typically dissolved in the mobile phase before injection into the sample loop. The sample is then drawn into a syringe and injected into the loop via the injection valve. A rotation of the valve rotor closes the valve and opens the loop in order to inject the sample into the stream of the mobile phase. Loop volumes can range between 10 [micro]l to over 500 [micro]l. In modern HPLC systems, the sample injection is typically automated.


Detection methods are critical to the successful application of HPLC methods in separation. Some of the conventional and newer detection methods are: UV/Vis, fluorescence, conductivity, and luminescence luminescence, general term applied to all forms of cool light, i.e., light emitted by sources other than a hot, incandescent body, such as a black body radiator. , Fourier, transform infrared spectroscopy (FTIR FTIR Fourier Transform Infrared (spectroscopy)
FTIR Frustrated Total Internal Reflection
FTIR Fourier Transfer Ir
), mass spectrometry (MS), and nuclear magnetic resonance nuclear magnetic resonance: see magnetic resonance.
nuclear magnetic resonance (NMR)

Selective absorption of very high-frequency radio waves by certain atomic nuclei subjected to a strong stationary magnetic field.
 (NMR NMR: see magnetic resonance. ). Exhaustive discussion of these newer detection methods has been reviewed in the literature. (10)

An example of HPLC Run conditions for polymer emulsion and/or paints can be as follows:

SAMPLE PREPARATION: The sample can be dissolved in THF at roughly 1% (wt/wt) and centrifuged at 40,000 rpm for 15 minutes. The supernatant is then filtered through 0.2 [micro]m filters prior to injection on the HPLC system.

CHROMATOGRAPHIC chro·mat·o·graph  
An instrument that produces a chromatogram.

tr.v. chro·mat·o·graphed, chro·mat·o·graph·ing, chro·mat·o·graphs
To separate and analyze by chromatography.

Column: PLRP-S 4000[Angstrom] 8[micro] analytical column (Polymer Labs), 4.6 m X 15 cm, 8 [micro]m [d.sub.p].

Mobile Phase: 25:75 THF/methanol initially. Linear gradient to 75:25 THF/methanol over 35 min. Back to initial conditions in 5 min. Equilibrate e·quil·i·brate  
v. e·quil·i·brat·ed, e·quil·i·brat·ing, e·quil·i·brates

To be in or bring about equilibrium.
To maintain in or bring into equilibrium.
 for 10 min.


Flow Rate: 1.0 ml/min

Injection Volume: 10 [micro]l

Column Temperature: Ambient

Detector 1: UV@270 nml

Detector 2: Evaporative Light Scatter-ing Detector (ELSD ELSD Evaporative Light Scattering Detector
ELSD Electronic Limited Slip Differential (automotive)
ELSD electronic limited slip device
ELSD Electric Limited Slip Differential


Raw materials that go into the manufacture of paints are sometimes more complex than the supplier or formulators realize. Components such as surfactants may be sold as "pure" or within certain specifications but may often contain several by-products that may play an important role in the chemistry of the formulated paint. The following examples illustrate how HPLC can be used for raw materials and fully formulated paint analyses.

ANALYSIS OF RESIDUAL ALCOHOLS AND ALKANES The following is a list of straight-chain alkanes and their common names, sorted by number of carbon atoms.

Number of C atoms Formula Common name Synonyms
1 CH4 Methane marsh gas; methyl hydride; natural gas
2 C2H6
 IN SURFACTANTS: Figure 4 shows HPLC analysis of two batches of surfactants for residual alcohol impurities. In Figure 5, the HPLC of two batches of a sulfated surfactant Surfactant Definition

Surfactant is a complex naturally occurring substance made of six lipids (fats) and four proteins that is produced in the lungs. It can also be manufactured synthetically.
 that are supposed to be chemically identical are presented.

ANALYSIS OF BINDER TYPE IN FULLY FORMULATED PAINT SYSTEMS: Compositional analysis of paints can be a daunting daunt  
tr.v. daunt·ed, daunt·ing, daunts
To abate the courage of; discourage. See Synonyms at dismay.

[Middle English daunten, from Old French danter, from Latin
 task for analysts. The paint components can be dissolved in appropriate solvent (THF) and separated by either normal phase HPLC (polar column and nonpolar solvents) or reverse phase HPLC (nonpolar column and polar solvents). For example, a failed coating can be extracted by THF; however, depending on the composition of the formulated paint, analysts may encounter very challenging situations where answers to specific questions may depend very much on many variables. For example, the use of organic opacifiers has many benefits including increased opacity, low binder demand, and partial Ti[O.sub.2] replacement, which significantly reduces cost. The challenge of determining binder types and composition in fully formulated paints which may or may not contain organic opacifiers (mainly styrene, such as Ropaques from Rohm and Haas Rohm and Haas Company (NYSE: ROH), a Philadelphia, Pennsylvania based company, manufactures miscellaneous materials. A Fortune 500 Company, Rohm and Haas employs more than 17,000 people in 27 countries. The annual sales revenue of Rohm and Haas stands at about USD 8.2 billion.  Company), can be very difficult. We have demonstrated in our labs that such systems can be analyzed by HPLC and/or HPLC/FTIR and/or off-line HPLC/Pyrolysis-GC/MS methods (see Figures 6 and 7).





Counteracting or preventing the building up of deposits on underwater surfaces, such as the undersides of boats: antifouling paint. 
 ADDITIVES IN PAINTS: Antifouling biocides are important in paints and coatings. Some of these preservatives include chlorothanolin, dichlorofluanid, diuron diuron

a phenylurea herbicide of low toxicity but capable of poisoning animals if given in very large amounts. Causes anorexia, weight loss and muscular weakness.
, Irgarol 1051, and SeaNine 211. These are relatively small molecules and the most common method for analysis of their purity, efficacy, level, and fate in the environment when associated with paint particles is by HPLC. (11) HPLC allows for the separation, identification, and quantitative analysis.

ANALYSIS OF EPOXY COATINGS RESINS: Sheih and Benton have provided an in-depth overview of how HPLC methods can be used in the analysis of epoxy coatings resins. (12) Epoxy resins are used widely in coatings due to their superior balance of properties such as good adhesion, impact resistance, chemical resistance, and flexibility.


Gel permeation chromatography (GPC), or size exclusion chromatography Size exclusion chromatography (SEC) is a chromatographic method in which particles are separated based on their size, or in more technical terms, their hydrodynamic volume. It is usually applied to large molecules or macromolecular complexes such as proteins and industrial polymers.  (SEC), is a branch of HPLC whose primary role is to provide molecular weight data on polymers and resins. GPC allows polymer synthesis chemists and paint formulation and application scientists to isolate and characterize low molecular weight additives and medium-to-high molecular weight polymers and resins in polymeric systems. The two most common detectors are ultraviolet (UV) and refractive index (RI). Only samples that absorb in the UV region (with UV chromophore chromophore /chro·mo·phore/ (kro´mo-for) any chemical group whose presence gives a decided color to a compound and which unites with certain other groups (auxochromes) to form dyes. ) can be detected by UV detection; for example, aromatic rings, or conjugated double bonds. If a polymer has no UV chromophore or is unknown, the RI detector is the best choice. The RI detector functions by monitoring the change in refractive index between the pure mobile phase and that containing the sample. The difference is the signal of the peak(s) of interest.

In GPC, the column is packed with a semi-rigid gel or small organic particles, which function as a physical barrier for separating molecules of different molecular weights. The key function of the stationary phase is to allow the polymer molecules to interact with the solvent molecules in such a way that those molecules that have more physical interaction with the pores of the column packing will take longer to elute e·lute  
tr.v. e·lut·ed, e·lut·ing, e·lutes
To extract (one material) from another, usually by means of a solvent.

[From Latin
 than those that have no physical interaction with the pores. For example, high molecular weight polymers will elute faster. The separation mechanism in a GPC column is represented in Figure 8.

It can be simplistically shown that the smaller molecules can fit into the small pores and therefore take longer to elute through the column. The larger molecules, on the other hand, due to their big size, are unable to enter the pores and simply travel around the gel particles on their way out. Thus, the large polymer molecules are the first to elute out, followed by the medium size, and finally by the smaller molecules. It is important to point out that GPC also separates molecules according to their hydrodynamic hy·dro·dy·nam·ic   also hy·dro·dy·nam·i·cal
1. Of or relating to hydrodynamics.

2. Of, relating to, or operated by the force of liquid in motion.
 volume. (13)

In order for the GPC data to be useful, it must be calibrated with polymers of known molecular weight. Polystyrene standards are often used because they are readily available over a wide range of molecular weights.


Coatings scientists are always interested in determining the composition of polymers as they largely determine the chemical resistance properties of coatings. GPC is one of the best methods that can be used to measure the molecular weight of polymers in coatings. In order to accomplish this, the polymer must be isolated from the paint by using an appropriate solvent. In the example shown in Figure 9, the polymer was isolated from a paint by extraction in THF followed by centrifugation to remove the pigment and other THF insolubles.

In coatings failure analysis, GPC can provide data to show whether the failing paints conform to specification by comparing and finger-printing failed paint to a control paint. Sometimes, failures due to polymer decomposition can easily be detected. For example, GPC performed on a failing and on control paints showed a dramatic difference in molecular weight, with the failing samples being lower than the control. (14) There are numerous examples in the literature of how GPC can be used to investigate coating failures. (15) It is important to point out that GPC has two major limitations in coating analysis: it cannot be used on insoluble coating samples and it cannot provide chemical information. However, the chemical information limitation can be overcome by interfacing GPC to a variety of chemically sensitive tools, such as FTIR, NMR, and MS. (7,12)


A general overview of the application of HPLC techniques for analysis of paints and related materials has been presented. We have discussed how HPLC can be used for raw materials and for fully formulated paint analyses. GPC has been demonstrated as one of the best methods that can be used to measure the molecular weight of polymers in coatings and coatings failure analyses. However, these methods require that the polymer or isolated components must be soluble in appropriate solvent(s).
Table 1 -- Anions and Cations that can be Separated and Quantified


Bromide    [Br.sup.-]
Chloride   [Cl.sup.-]
Fluoride   [F.sup.-]
Nitrate    N[O.sub.3-]
Nitrite    N[O.sub.2-]
Phosphate  P[O.sub.4.sup.3-]
Sulfate    S[O.sub.4.sup.2-]


Ammonium   N[H.sub.4.sup.+]
Calcium    [Ca.sup.2+]
Lithium    [Li.sup.+]
Magnesium  [Mg.sup.2+]
Sodium     [Na.sup.+]
Zinc       [Zn.sup.2+]

Abbreviations and Acronyms

FTIR        Fourier Transform Infrared
GPC         Gel Permeation Chromatography
HPLC        High Performance Liquid Chromatography
LC          Liquid Chromatography
MS          Mass Spectrometry
MWT         Molecular Weight
NMR         Nuclear Magnetic Resonance
RPLC        Reverse-Phase Liquid Chromatography
Pyro-GC/MS  Pyrolysis Gas Chromatography Mass Spectrometry
SEC         Size Exclusion Chromatography
THF         Tetrahydrofuran
UV          Utraviolet


(1) Dorsey, J.G., "Liquid Chromatography" and "Column Theory and Resolution in Liquid Chromatography" Encyclopedia of Analytical Chemistry The Encyclopedia of Analytical Chemistry is an English-language multivolume encyclopedia published by John Wiley & Sons Ltd.

It is a comprehensive analytical chemistry reference, covering all aspects from theory and instrumentation through applications and techniques.
, Vol. 15, 11231-11233 and 11334-1134 (2000).

(2) Hage, D.S D.S Drainage Structure (flood protection) ., "Affinity Chromatography," Encyclopedia of Analytical Chemistry, Vol. 15, 11233-11251 (2000).

(3) Dolan, J.W. and Snyder, L.R., "Gradient Elution Chromatography," Encyclopedia of Analytical Chemistry, Vol. 15, 11342-11360 (2000).

(4) Pietrzyk, D.J., "Ion Chromatography," Encyclopedia of Analytical Chemistry, Vol. 15, 11360-11382 (2000).

(5) Cooper, W.T., "Normal Phase Chromatography," Encyclopedia of Analytical Chemistry, Vol. 15, 11428-11442 (2000).

(6) Lough, W.J., "Reverse Phase Liquid Chromatography," Encyclopedia of Analytical Chemistry, Vol. 15, 11442-11450 (2000).

(7) Provder, T., Barth, H.G., and Urban, M.W. (Eds.), in Chromatographic Characterization of Polymers Hyphenated and Multidimensional Techniques, Advances in Chemistry Series 247, American Chemical Society The American Chemical Society (ACS) is a learned society (professional association) based in the United States that supports scientific inquiry in the field of chemistry. Founded in 1876 at New York University, the ACS currently has over 160,000 members at all degree-levels and in , Washington, D.C. (1995).

(8) Simio, C., Cottet, H., Vayaboury, W., Giani, O., Pelzing, M., and Cifuentes, A., "Nonaqueous Capillary Electrophoresis-Mass Spectrometry of Synthetic Polymers," Anal. Chem., 76 (2) pp 335-344 (2004).

(9) Daimon, H. and Hirata, Y., "Directly Coupled Supercritical Fluid Chromatography for Analysis of Polymer Additives," Chromatographia, 32, 549-554 (1991).

(10) Patony, G. (Ed.), in HPLC Detection Newer Methods, VCH VCH Victoria County History
VCH Vertical Clitoral Hood (piercing)
VCH Volunteer Clearing House (University of Colorado)
VCH Vliegclub Hoogeveen
VCH Virtual Channel Handler
 Publishers, Inc., New York, NY (1992).

(11) Thomas, K.V., McHugh, M., Hilton, M., and Waldock, M., "Increased Persistence of Antifouling Paint Biocides When Associated with Paint Particles," Environmental Pollution, 123 (1), 153-161 (2003).

(12) Sheih, D.P. and Benton, D.E., "HPLC Analysis for Epoxy Coatings Resins," Analysis of Paints and Related Materials: Current Techniques for Solving Coatings Problems, ASTM ASTM
American Society for Testing and Materials
 STP STP or standard temperature and pressure, standard conditions for measurement of the properties of matter. The standard temperature is the freezing point of pure water, 0°C; or 273.15°K;.  1119, American Society for Testing and Materials, Philadelphia, 41-46, (1992).

(13) Snyder, L.R. and Kirkland, J.J., Introduction to Modern Chromatography, John Wiley, New York, NY, 512-514 (1979).

(14) Weldon, D.G., in Failure Analysis of Paints and Coatings, John Wiley and Sons, New York, NY, pp. 206, 2001.

(15) Kuo, C. and Provder, T., "High Performance Gel Permeation Chromatography Characterization of Oligomers Used in Coatings Systems," JOURNAL OF COATINGS TECHNOLOGY, 53, No. 683, 69 (1981).

by Francis V. Acholla, Rohm and Haas Company*

*Spring House Technical Center, 727 Norristown Rd., Spring House, PA 19477.
COPYRIGHT 2005 Federation of Societies for Coatings Technology
No portion of this article can be reproduced without the express written permission from the copyright holder.
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Title Annotation:Technology Today
Author:Acholla, Francis V.
Publication:JCT CoatingsTech
Date:Jun 1, 2005
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