Chapter 18 : Chemical Analysis

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The methods selected for this chapter are most commonly used in research relating to microbial components and metabolism, but most are also suited to advanced undergraduate and graduate teaching laboratories. The incorporation of radioactively labeled compounds into bacterial cells is used as a means of estimating the rates and amounts of synthesis as well as the distribution of the main small molecule and macromolecule fractions of the cells. Macromolecules are precipitated by cold dilute solutions of trichloroacetic acid. The precipitate fraction of macromolecules is sequentially extracted with organic solvents for lipids, alkali for RNAs, and hot trichloroacetic acid for DNAs; the precipitate remaining after these extractions contains the cell proteins and peptidoglycan. Some macromolecules contain both carbohydrate and noncarbohydrate components (lipopolysaccharides, peptidoglycans, teichoic acids, lipoteichoic acids, teichuronic acids, nucleic acids, glycoproteins). In some cases gas-liquid chromatography can be useful when coupled with mass spectrometry. For nitrite analysis, the most widely accepted method involves modification of the diazotization and coupling reactions. The methods described in this section differ mainly in how the DNA is purified and prepared. A number of short-chain organic acids and alcohols are formed in bacterial cells as intermediates or end products of the citric acid, glycolytic, and other metabolic pathways.

Citation: Daniels L, Hanson R, Phillips J. 2007. Chemical Analysis, p 462-503. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch18

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Bacteria and Archaea
Organic Nitrogen Compounds
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Flow diagram of fractionation procedure.

Citation: Daniels L, Hanson R, Phillips J. 2007. Chemical Analysis, p 462-503. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch18
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1. Kennell, D. 1967. Use of filters to separate radioactivity in RNA, DNA, and protein. Methods Enzymol.: Nucleic Acids 12A:686692.
2. Roberts, R. B.,, D. B. Cowie,, E. T. Bolton,, P. H. Abelson,, and R. J. Britten. 1955. Studies of biosynthesis in Escherichia coli. Publ. 607. Carnegie Institute of Washington, Washington, DC.
3. Sutherland, I. W.,, and J. F. Wilkinson,. 1971. Chemical extraction methods of microbial cells, p. 345383. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5B. Academic Press, Inc., New York, NY.
4. Chytil, F.,, and D. B. McCormick (ed.). 1986. Vitamins and coenzymes. Methods Enzymol. 122:3425.
5. Daniels, L.,, N. Bakhiet,, and K. Harmon. 1985. Widespread distribution of a 5-deazaflavin cofactor in Actinomycetes and related bacteria. Syst. Appl. Microbiol. 6:1217.
6. Eirich, L. D.,, G. D. Vogels,, and R. S. Wolfe. 1978. Proposed structure for coenzyme F420 from Methanobacterium. Biochemistry 17:45834593.
7. Escalente-Semerena, J. C.,, K. L. Rinehart, Jr.,, and R. S. Wolfe. 1984. Tetrahydromethanopterin, a carbon-carrier in methanogenesis. J. Biol. Chem. 259:94479455.
8. Gorris, L. G.,, and C. vander Drift. 1994. Cofactor contents of methanogenic bacteria reviewed. Biofactors 4:139145.
9. Hausinger, R. P.,, W. H. Orme-Johnson,, and C. Walsh. 1985. Factor 390 chromophores: phosphodiester between AMP or GMP and methanogen factor 420. Biochemistry 24:16291633.
10. Isabelle, D. W.,, D. R. Simpson,, and L. Daniels. 2002. Large-scale production of coenzyme F420 by using Mycobacterium smegmatis. Appl. Environ. Microbiol. 68:57505755.
11. McCormick, D. B.,, J. W. Suttie,, and C. Wagner (ed.). 1997. Methods in Enzymology, Part I. 279:3502.
12. McCormick, D. B.,, J. W. Suttie,, and C. Wagner (ed.). 1997. Methods in Enzymology, Part J. 280:3496.
13. McCormick, D. B.,, J. W. Suttie,, and C. Wagner (ed.). 1997. Methods in Enzymology, Part K. 281:3469.
14. McCormick, D. B.,, J. W. Suttie,, and C. Wagner (ed.). 1997. Methods in Enzymology, Part L. 282:3505.
15. Purwantini, E. 1991. Coenzyme F420: factors affecting its purification from Methanobacterium thermoautotrophicum. M.S. Thesis. University of Iowa, Iowa City.
16. Schonheit, P.,, H. Keweloh,, and R. K. Thauer. 1981. Factor F420 degradation in Methanobacterium thermoautotrophicum during exposure to oxygen. FEMS Microbiol. Lett. 12:347349.
17. Colowick, J.,, and N. Kaplan (ed.). 1966. Complex carbohydrate. Methods Enzymol. 8:1759.
18. Ginsburg, V. (ed.). 1989. Complex carbohydrates. Methods Enzymol. 179:3287.
19. Herbert, D.,, P. J. Phipps,, and R. E. Strange,. 1971. Chemical analysis of microbial cells, p. 209344. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5B. Academic Press, Inc., New York, NY.
20. Work, E., 1971. Cell walls, p. 361418. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5A. Academic Press, Inc., New York, NY.
21. Bera, A.,, R. Biswas,, S. Herbert,, and F. Gotz. 2006. The presence of peptidoglycan O-acetyltransferase in various staphylococcal species correlates with lysozyme resistance and pathogenicity. Infect. Immun. 74:45984604.
22. Blumenkrantz, N.,, and G. Asboe-Hansen. 1976. An assay for total hexosamine and a differential assay for glucosamine. Clin. Biochem. 9:269274.
23. Brade, H.,, C. Galanos,, and O. Luderitz. 1983. Differential determination of the 3-deoxy-D-mannooctulosonic acid residues in lipopolysaccharides of Salmonella minnesota rough mutants. Eur. J. Biochem. 131:195200.
24. Bryan, B. A.,, R. J. Linhardt,, and L. Daniels. 1986. Variation in composition and yield of exopolysaccharides produced by Klebsiella sp. strain K32 and Acinetobacter calcoaceticus BD4. Appl. Environ. Microbiol. 51:13041308.
25. Cadmus, M. C.,, C. A. Knutson,, A. A. Lagoda,, J. E. Pittsley,, and K. A. Burton. 1978. Synthetic media for production of quality xanthan gum in 20 liter fermentors. Biotechnol. Bioeng. 20:10031014.
26. Caroff, M.,, S. Lebbar,, and L. Szabo. 1987. Detection of 3-deoxy-2-octulosonic in thiobarbiturate-negative endotoxins. Carbohydr. Res. 161:c4c7.
27. Chen, W.-P.,, J.-Y. Chen,, S.-C. Chang,, and C.-L. Su. 1985. Bacterial alginate produced by a mutant of Azotobacter vinelandii. Appl. Environ. Microbiol. 49:543546.
28. Cox, A. D.,, J. C. Wright,, M. A. J. Gidney,, S. Lacelle,, J. S. Plested,, A. Martin,, E. R. Moxton,, and J. C. Richards. 2003. Identification of a novel inner-core oligosaccharide structure in Neisseria meningitidis lipopolysaccharide. Eur. J. Biochem. 270:17591766.
29. Dische, Z. 1953. Qualitative and quantitative colorimetric determination of heptoses. J. Biol. Chem. 204:983997.
30. Dische, Z. 1962. Color reactions of carbohydrates. Methods Carbohydr. Chem. 1:477514.
31. Elson, L. A.,, and W. T. Morgan. 1933. A colorimetric method for the determination of glucosamine and chondrosamine. Biochem. J. 27:18241828.
32. Ford, J. D.,, and J. C. Haworth. 1964. The estimation of galactose in plasma using galactose oxidase. Clin. Chem. 10:10021006.
33. Fox, A.,, and G. Black,. 1994. Identification and detection of carbohydrate markers for bacteria: derivatization and gas chromatography-mass spectrometry, p. 107131. In C. Fenselau (ed.), Mass Spectrometry for the Characterization of Microorganisms. American Chemical Society, Washington, DC.
34. Fox, G. E.,, L. J. Magrum,, W. E. Balch,, R. S. Wolfe,, and C. R. Woese. 1977. Classification of methanogenic bacteria by 16S ribosomal RNA. Proc. Natl. Acad. Sci. USA 74:45374541.
35. Fox, K. F.,, A. Fox,, M. Nagpal,, P. Steinberg,, and K. Heroux. 1998. Identification of Brucella by ribosomalspacer- region PCR and differentiation of Brucella canis from other Brucella spp. pathogenic for humans by carbohydrate profiles. J. Clin. Microbiol. 36:32173222.
36. Fukagawa, K.,, H. Yamaguchi,, D. Yonezawa,, and S. Murao. 1974. Isolation and characterization of polysaccharide produced by Rhodotorula glutinis K-24. Agr. Biol. Chem. 38:2935.
37. Ghuysen, J.-M.,, D. J. Tipper,, and J. L. Strominger. 1966. Enzymes that degrade bacterial cell walls. Complex Carbohydr. 8:685699.
38. Gillespie, J.,, S. T. Weintraub,, G. G. Wong,, and S. C. Holt. 1988. Chemical and biological characterization of the lipopolysaccharide of the oral pathogen Wolinella recta ATCC 33238. Infect. Immun. 56:20282035.
39. Hacking, A. J.,, I. W. F. Taylor,, T. R. Jarman,, and J. R. W. Govan. 1983. Alginate biosynthesis by Pseudomonas mendocina. J. Gen. Microbiol. 129:34733480.
40. Hadzija, O. 1974. A simple method for the quantitative determination of muramic acid. Anal. Biochem. 60:512517.
41. Ito, M.,, K. Ikeda,, Y. Suzuki,, K. Tanaka,, and M. Saito. 2002. An improved fluorometric high-performance liquid chromatography method for sialic acid determination: an internal standard method and its application to sialic acid analysis of human apolipoprotein E. Anal. Biochem. 300:260266.
42. Kaplan, N.,, E. Rosenberg,, B. Jann,, and K. Jann. 1985. Structural studies of the capsular polysaccharide of Acinetobacter calcoaceticus BD4. Eur. J. Biochem. 152:453458.
43. Karkhanis, Y. D.,, J. Y. Zeltner,, J. J. Jackson,, and D. J. Carlo. 1978. A new and improved microassay to determine 2-keto-3-deoxyoctonate in lipopolysaccharide of gram-negative bacteria. Anal. Biochem. 85:595601.
44. Kosinski, E. D. 1981. The indirect determination of lactose using a glucose analyzer with particular reference to solutions containing low levels of lactose in high levels of glucose and galactose. J. Soc. Dairy Technol. 43:2831.
45. Lee, Y. C. 1990. High-performance anion-exchange chromatography for carbohydrate analysis. Anal. Biochem. 189:151162.
46. Linhardt, R. J.,, R. Bakhit,, L. Daniels,, F. Mayerl,, and W. Pickenhagen. 1989. Microbially produced rhamnolipid as a source of rhamnose. Biotechnol. Bioeng. 33:365368.
47. Lott, J. A.,, and K. Turner. 1975. Evaluation of Trinder’s glucose oxidase method for measuring glucose in serum and urine. Clin. Chem. 21:17541760.
48. Lysenko, E.,, J. C. Richards,, A. D. Cox,, A. Stewart,, A. Martin,, M. Kapoor,, and J. N. Weiser. 2000. The position of phosphoryl-choline on the lipopolysaccharide of Haemophilus influenzae affects binding and sensitivity to C-reactive protein-mediated killing. Mol. Microbiol. 35:234245.
49. Mason, M. 1982. A new method for ethanol measurement utilizing an immobilized enzyme. J. Am. Soc. Brew. Chem. 40:7879.
50. Mason, M. 1983. Determination of glucose, sucrose, lactose, and ethanol in foods and beverages, using immobilized enzyme electrodes. J. Assoc. Off. Anal. Chem. 66:981984.
51. Mukumoto, T.,, and H. Yamaguchi. 1977. The chemical structure of a mannofucogalactan from the fruit bodies of Flammulina velutipes (Fr.) Sing. Carbohydr. Res. 59:614621.
52. Nesser, J.-R.,, and T. F. Schweizer. 1984. A quantitative determination by gas-liquid chromatography of neutral and amino sugars (as O-methyloxime acetates), and a study on hydrolytic conditions for glycoproteins and polysaccharides in order to increase sugar recoveries. Anal. Biochem. 142:5867.
53. Osborn, M. J. 1963. Studies on the Gram-negative cell wall. I. Evidence for the role of 2-keto-3-deoxyoctanoate in the lipopolysaccharide of Salmonella typhimurium. Proc. Natl. Acad. Sci. USA 50:499506.
54. Pfeffer, J. M.,, H. Strating,, J. T. Weadge,, and A. J. Clarke. 2006. Peptidoglycan O acetylation and autolysin profile of Enterococcus faecalis in the viable but nonculturable state. J. Bacteriol. 188:902908.
55. Rocklin, R. D.,, and C. A. Pohl. 1983. Determination of carbohydrates by anion exchange chromatography with pulsed amperometric detection. J. Liquid Chromatogr. 6:15771590.
56. Rondle, C. J. M.,, and W. T. J. Morgan. 1955. The determination of glucosamine and galactosamine. Biochem. J. 61:586590.
57. Roth, H.,, S. Segal,, and D. Bertoli. 1965. The quantitative determination of galactose—an enzymic method using galactose oxidase, with applications to blood and other biological fluids. Anal. Biochem. 10:3252.
58. Sempre, J. M.,, C. Gancedo,, and C. Asensio. 1965. Determination of galactosamine and N-acetylgalactosamine in the presence of other hexosamines with galactose oxidase. Anal. Biochem. 12:509515.
59. Smith, I. H.,, and G. W. Pace. 1982. Recovery of microbial polysaccharides. J. Chem. Technol. Biotechnol. 32:119129.
60. Stewart-Tull, D. E. S. 1968. Determination of amino sugars in mixtures containing glucosamine, galactosamine, and muramic acid. Biochem. J. 109:1318.
61. Stork, A. D. M.,, H. Kemperman,, D. W. Erkelens,, and T. F. Veneman. 2005. Comparison of the accuracy of the HemoCue Glucose analyzer with the Yellow Springs Instrument glucose oxidase analyzer, particularly in hypoglycemia. Eur. J. Endocrinol. 153:275281.
62. Sweeley, C. C.,, W. W. Wells,, and R. Bentley. 1966. Gas chromatography of carbohydrates. Methods Enzymol.: Complex Carbohydr. 8:95107.
63. Taylor, P. J.,, E. Kmetec,, and J. M. Johnson. 1977. Design, construction, and applications of a galactose selective electrode. Anal. Chem. 49:789794.
64. Tipper, D. J. 1968. Alkali-catalyzed elimination of Dlactic acid from muramic acid and its derivatives, and the determination of lactic acid. Biochemistry 7:14411449.
65. Weadge, T. J.,, J. M. Pfeffer,, and A. J. Clarke. 2005. Identification of a new family of enzymes with potential O-acetylpeptidoglycan esterase activity in both grampositive and gram-negative bacteria. BMC Microbiol. 5:49.
66. Weisbach, A.,, and J. Hurwitz. 1959. The formation of 2-keto-3-deoxyheptanoic acid in extracts of E. coli B. J. Biol. Chem. 234:705712.
67. Dittmer, J. C.,, and M. A. Wells. 1969. Quantitative and qualitative analysis of lipids and lipid components. Methods Enzymol.: Lipids 14:482530.
68. Kates, M., 1972. Techniques of lipidology isolation, analysis, and identification of lipids, p. 275327. In T. S. Work, and E. Work (ed.), Laboratory Techniques in Biochemistry and Molecular Biology, vol. 3. American Elsevier Publishing Co., Inc., New York, NY.
69. O’Leary, W., 1974. Chemical and physical characterization of fatty acids, p. 275327. In A. I. Laskin, and H. A. Lechevalier (ed.), Handbook of Microbiology, vol. 2. Chemical Rubber Co., Cleveland, OH.
70. Bird, C. W.,, J. M. Lynch,, S. J. Pirt,, W. W. Reid,, C. J. Brooks,, and B. S. Middleditch. 1971. Steroids and squalene in Methylococcus capsulatus grown on methane. Nature (London) 230:473474.
71. Bligh, E. G.,, and W. J. Dyer. 1959. A rapid method of total lipid extraction and purification. Can. J. Biochem. Physiol. 37:911917.
72. Brandl, H.,, R. A. Gross,, R. W. Lenz,, and R. C. Fuller. 1988. Pseudomonas oleovorans as a source of poly-β-hydroxyalkanoates for potential applications as biodegradable polyesters. Appl. Environ. Microbiol. 54:19771982.
73. Brandl, H.,, R. A. Gross,, R. W. Lenz,, and R. C. Fuller. 1990. Plastics from bacteria and for bacteria: poly-β- hydroxy-alkanoates as natural, biocompatible, and biodegradable polyesters. Adv. Biochem. Eng. Biotechnol. 41:7793.
74. Braunegg, G.,, B. Sonnleitner,, and R. M. Lafferty. 1978. A rapid gas chromatographic method for the determination of poly-beta-hydroxybutyric acid in microbial biomass. Appl. Microbiol. Biotechnol. 6:2937.
75. Folch, J. 1957. A simple method for the isolation and purification of total lipids from animal tissue. J. Biol. Chem. 226:497509.
76. Hara, A.,, and N. S. Radin. 1978. Lipid extraction with a low-toxicity solvent. Anal. Biochem. 90:420426.
77. Hopmans, E. C.,, S. Schouten,, R. D. Pancost,, M. T. J. van der Meer,, and J. S. S. Damste. 2000. Analysis of intact tetraether lipids in archaeal cell material and sediments by high performance liquid chromatography/atmospheric pressure chemical ionization mass spectrometry. Rapid Commun. Mass Spectrom. 14:585589.
78. Jahn, U.,, R. Simmons,, H. Sturt,, E. Grosjean,, and H. Huber. 2004. Composition of the lipids of Nanoarchaeum equitans and their origin from its host Ignicoccus sp. Strain KIN4/I. Arch. Microbiol. 182:404413.
79. Jarute, G.,, A. Kainz,, G. Schroll,, J. R. Baena,, and B. Lendl. 2004. On-line determination of the intracellular poly(β-hydroxybutyric acid) content in transformed Escherichia coli and glucose during PHB production using stopped-flow attenuated total reflection FT-IR spectrometry. Anal. Chem. 76:63536358.
80. Johnson, A. R.,, and R. B. Stocks,. 1971. Gas chromatography of lipids, p. 195218. In A. R. Johnson, and J. B. Davenport (ed.), Biochemistry and Methodology of Lipids. Wiley-Interscience, New York, NY.
81. Kates, M., 1993. Membrane lipids of archaea, p. 261295. In M. Kates,, D. J. Kushner, and A. T. Matheson (ed.), The Biochemistry of Archaea. Elsevier, Amsterdam, The Netherlands.
82. Kates, M. 1964. Simplified procedure for hydrolysis or methanolysis of lipids. J. Lipid Res. 5:132135.
83. Kates, M.,, S. C. Kushwaha,, and G. D. Sprott. 1982. Lipids of purple membrane from extreme halophiles and of methanogenic bacteria. Methods Enzymol. 88:98111.
84. Kates, M.,, L. S. Yengoyan,, and P. S. Sastry. 1965. A diether analog of phosphytidyl glycerophosphate in Halobacterium cutirubrum. Biochim. Biophys. Acta 98:252268.
85. Koga, Y.,, and H. Morii. 2005. Recent advances in structural research on ether lipids from Archaea including comparative and physiological aspects. Biosci. Biotechnol. Biochem. 69:20192034.
86. Kushwaha, S. C.,, M. Kates,, G. D. Sprott,, and I. C. P. Smith. 1981. Novel complex polar lipids from the methanogenic archaebacterium Methanospirillum hungatei. Science 211:11631164.
87. Langworthy, T. A. 1977. Comparative lipid composition of heterotrophically and autotrophically grown Sulfolobus acidocaldarius. J. Bacteriol. 130:13261332.
88. Law, J. H.,, and R. A. Slepecky. 1961. Assay of poly-β- hydroxybutyric acid. J. Bacteriol. 82:3336.
89. Lenz, R. W.,, B. W. Kim,, H. W. Ulmer,, K. Fritzche,, E. Knee,, and R. C. Fuller,. 1990. Functionalized poly-β- hydroxyalkanoates produced by bacteria, p. 2335. In E. A. Dawes (ed.), Novel Biodegradable Microbial Polymers. Kluwer Academic Publishers, Dordrecht, The Netherlands.
90. Paton, J. C.,, E. J. McMurchie,, B. K. May,, and W. H. Elliot. 1978. Effect of growth temperature on membrane fatty acid composition and susceptibility to cold shock of Bacillus amyloliquefaciens. J. Bacteriol. 135:754759.
91. Patt, T. E.,, and R. S. Hanson. 1978. Intracytoplasmic membrane, phospholipid, and sterol content of Methylobacterium organophilum cells grown under different conditions. J. Bacteriol. 134:636644.
92. Rilfors, L.,, A. Wieslander,, and S. Stahl. 1978. Lipid and protein composition of membranes of Bacillus megaterium variants in the temperature range of 5 to 70°C. J. Bacteriol. 135:10431052.
93. Shamala, T. R.,, A. Chandrashekar,, S. V. N. Vijayendra,, and L. Kshama. 2003. Identification of polyhydroxyalka- noate (PHA)-producing Bacillus spp. using the polymerase chain reaction (PCR). J. Appl. Microbiol. 94:369374.
94. Skipski, V. P.,, and M. Barlday. 1969. Detection of lipids on thin-layer chromatograms. Methods Enzymol.: Lipids 14:541548.
95. Stein, R. A.,, V. Slawson,, and J. F. Mead,. 1976. Gasliquid chromatography of fatty acids and derivatives, p. 857896. In G. V. Marinetti (ed.), Lipid Chromatographic Analysis, 2nd ed. Marcel Dekker, Inc., New York, NY.
96. Tornabene, T. G.,, and T. A. Langworthy. 1979. Diphytanyl and dibiphytanyl glycerol ether lipids of methanogenic archaebacteria. Science 203:5153.
97. Tornabene, T. G.,, R. S. Wolfe,, W. E. Balch,, C. Holzer,, G. E. Fox,, and J. Oro. 1978. Phytanyl-glycerol ethers and squalenes in the archaebacterium Methanobacterium thermoautotrophicum. J. Mol. Evol. 11:259266.
98. Virtue, P.,, P. D. Nichols,, and P. I. Boon. 1996. Simultaneous estimation of microbial phospholipids fatty acids and diether lipids by capillary gas chromatography. J. Microbiol. Methods 25:177185.
99. Vorbeck, M. L.,, and G. V. Marinetti. 1965. Intracellular distribution and characterization of the lipids of Streptococcus faecalis. Biochemistry 4:296305.
100. Welch, D. F. 1991. Applications of fatty acid analysis. Clin. Microbiol. Rev. 4:422438.
101. Wuthier, R. E. 1966. Purification of lipids from non-lipid contaminants on Sephadex bead columns. J. Lipid Res.
102. Braun, V. 1978. Structure-function relationship of the Gram-negative bacterial cell envelope. Symp. Soc. Gen. Microbiol. 28:111138.
103. Braun, V.,, and K. Hantke. 1974. Biochemistry of bacterial cell envelopes. Annu. Rev. Biochem. 43:89121.
104. Costerton, J. W.,, J. M. Ingram,, and K. J. Cheng. 1974. Structure and function of the cell envelope of gram-negative bacteria. Bacteriol. Rev. 38:87110.
105. Cummins, C. S., 1974. Bacterial cell wall structures, p. 251284. In A. I. Laskin, and H. A. Lechevalier (ed.), Handbook of Microbiology, vol. 2. Microbial Composition. Chemical Rubber Co., Cleveland, OH.
106. Elin, R. J.,, and S. M. Wolff,. 1974. Bacterial endotoxin, p. 674731. In A. I. Laskin, and H. A. Lechevalier (ed), Handbook of Microbiology, vol. 2. Microbial Composition. Chemical Rubber Co., Cleveland, OH.
107. Ghuysen, J. M. 1968. Use of bacteriolytic enzymes in determination of wall structure and their role in cell metabolism. Bacteriol. Rev. 32:425464.
108. Leive, L. (ed.). 1973. Bacterial Membranes and Walls. Marcel Dekker, Inc., New York, NY.
109. Luderitz, O.,, A. M. Staub,, and O. Westphal. 1966. Immunochemistry of O and R antigens of Salmonella and related Enterobacteriacae. Bacteriol. Rev. 30:192255.
110. Milner, K. C.,, J. A. Rudbach,, and E. Ribi,. 1971. General characteristics, p. 165. In G. Weinbaum,, S. Kadis,, and S. Ajl (ed.), Microbial Toxins: Bacterial Endotoxins, vol. 4. Academic Press, Inc., New York, NY.
111. Reaveley, D. A.,, and R. E. Burge. 1972. Walls and membranes in bacteria. Adv. Microb. Physiol. 7:181.
112. Rogers, H. J.,, J. B. Ward,, and I. D. J. Burdett. 1978. Structure and growth of the walls of Gram-positive bacteria. Symp. Soc. Gen. Microbiol. 28:139176.
113. Schleifer, K. H.,, and O. Kandler. 1972. Peptidoglycan types of bacterial cell walls and their taxonomic implications. Bacteriol. Rev. 36:407477.
114. Cox, A. D.,, J. C. Wright,, M. A. J. Gidney,, S. Lacelle,, J. S. Plested,, A. Martin,, E. R. Moxton,, and J. C. Richards. 2003. Identification of a novel inner-core oligosaccharide structure in Neisseria meningitidis lipopolysaccharide. Eur. J. Biochem. 270:17591766.
115. Davidson, A. L.,, and J. Badiley. 1964. Glycerol teichoic acids in walls of Staphylococcus epidermidis. Nature (London) 202:874.
116. Ghuysen, J. M.,, D. Tipper,, and J. L. Strominger. 1965. Structure of the cell wall of Staphylococcus epidermidis strain Copenhagen. IV. The soluble glycopeptide and its sequential degradation by peptidase. Biochemistry 4:22452256.
117. Kandler, O.,, and H. Hippe. 1977. Lack of peptidoglycan in the cell walls of Methanosarcina barkeri. Arch. Microbiol. 113:5760.
118. Kandler, O.,, and H. Konig. 1978. Chemical composition of the peptidoglycan-free cell walls of methanogenic bacteria. Arch. Microbiol. 118:141152.
119. Kandler, O.,, and H. Konig,. 1985. Cell envelopes of archaebacteria, p. 413457. In C. R. Woese, and R. S. Wolfe (ed.), The Bacteria, vol. 8. Academic Press, Orlando, FL.
120. Knox, K. W. 1966. The relation of 3-deoxy-2-oxooctanoate to the serological and physical properties of a lipopolysaccharide from a rough strain of Escherichia coli. Biochem. J. 100:7378.
121. Kottel, R. H.,, K. Bacon,, D. Clutter,, and D. White. 1975. Coats from Myxococcus xanthus: characterization and synthesis during myxospore differentiation. J. Bacteriol. 124:550557.
122. Osborn, M. J. 1963. Studies on the Gram-negative cell wall. Proc. Natl. Acad. Sci. USA 50:499514.
123. Raetz, C. R. H. 1990. Biochemistry of endotoxins. Annu. Rev. Biochem. 59:129170.
124. Strominger, J. L.,, and D. J. Tipper,. 1974. Structure of bacterial cell walls: the lysozyme substrate, p. 169184. In E. F. Osserman,, R. E. Canfield,, and S. Beychok (ed.), Lysozyme. Academic Press, Inc., New York, NY.
125. Taylor, A.,, K. W. Knox,, and E. Work. 1966. Chemical and biological properties of an extracellular liposaccharide from Escherichia coli grown under lysine-limiting conditions. Biochem. J. 99:5361.
126. Warth, A. W.,, and J. L. Strominger. 1969. Structure of the peptidoglycan of bacterial spores: occurrence of the lactam of muramic acid. Proc. Natl. Acad. Sci. USA 64:528535.
127. Weidel, W.,, H. Frank,, and H. H. Martin. 1960. The rigid layer of the cell wall of Escherichia coli strain B. J. Gen. Microbiol. 22:158166.
128. Westphal, O.,, and K. Jan. 1965. Bacterial lipopolysaccharide extraction with phenol-water and further applications of the procedure. Methods Carbohydr. Chem. 5:8391.
129. Work, E., 1971. Cell walls, p. 361418. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5A. Academic Press, Inc., New York, NY.
130. DeLey, J., 1971. The determination of the molecular weight of DNA per bacterial nucleoid, p. 301311. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5A. Academic Press, Inc., New York, NY.
131. Parish, J. H. 1972. Principles and Practice of Experiments with Nucleic Acids. Longman Group, Ltd., London, United Kingdom.
132. Brunk, C. F.,, K. C. Jones,, and T. W. James. 1979. Assay for nanogram quantities of DNA in cellular homogenates. Anal. Biochem. 92:497500.
133. Burton, K. 1957. A study of the conditions and mechanism of the diphenylamine reaction for the calorimetric estimation of deoxyribonucleic acid. Biochem. J. 62:315323.
134. Cesarome, C.,, C. Bolognesi,, and L. Santi. 1979. Improved microfluorometric DNA determination in biological material using 33258 Hoechst. Anal. Biochem. 179:401403.
135. Gallagher, S., 1989. Quantitation of DNA and RNA with absorption and fluorescence spectroscopy, p. A.3.9A.3.15. In F. A. Ausubel,, R. Brent,, R. E. Kingston,, D. D. Moore,, J. M. Seidman,, J. A. Smith,, and K. Struhl (ed.), Current Protocols in Molecular Biology, supplement 8. Greene Publishing Associates and John Wiley & Sons, Inc., New York, NY.
136. Griswold, B. L.,, E. L. Humoller,, and A. R. McIntyre. 1951. Inorganic phosphates and phosphate esters in tissue extracts. Anal. Chem. 23:192194.
137. Labarca, C.,, and K. Paigen. 1980. A simple, rapid, and sensitive DNA assay procedure. Anal. Biochem. 102:344352.
138. Sambrook, J.,, and D. W. Russell. 2001. Molecular Cloning: a Laboratory Manual, 3rd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
139. Tilzer, L.,, S. Thomas,, and R. F. Moreno. 1989. Use of silica gel polymer for DNA extraction with organic solvents. Anal. Biochem. 183:1315.
140. Allen, S. E.,, H. M. Grimshaw,, J. A. Parkinson,, and C. Quarmby,. 1974. Inorganic constituents: nitrogen, p. 184206. In S. E. Allen (ed.), Chemical Analysis of Ecological Materials. Blackwell Scientific Publications, London, United Kingdom.
141. Black, C. A. (ed.). 1965. Methods of Soil Analysis, part 2. Chemical and Microbiological Properties. American Society of Agronomy, Inc., Madison, WI.
142. Bremner, J. M.,, and D. R. Keeney. 1965. Steam distillation methods for determination of ammonium, nitrate, and nitrite. Anal. Chim. Acta 32:485495.
143. Cooper, T. C. 1977. The Tools of Biochemistry. John Wiley & Sons, Inc., New York, NY.
144. Davis, B. D.,, R. Dulbecco,, H. N. Eisen,, and H. S. Ginsberg. 1990. Microbiology, p. 4549. J. B. Lippincott, Philadelphia, PA.
145. Franson, M. A. (ed.). 1976. Standard Methods for the Examination of Water and Wastewater, 14th ed. American Public Health Association, Washington, DC.
146. Gudinowlcz, B. J.,, M. J. Gudlnowlcz,, and H. F. Martin. 1976. Fundamentals of Integrated GC-MS. Part II. Mass Spectrometry. Marcel Dekker, Inc., New York, NY.
147. Horowitz, W. (ed.). 1975. Official Methods of Analyses of the Association of Official Analytical Chemists, 12th ed. Association of Official Analytical Chemists, Washington, DC.
148. Jackson, M. L. 1958. Soil Chemical Analysis. Prentice- Hall, Inc., Englewood Cliffs, NJ.
149. Safe, S.,, and O. Hutzinger. 1973. Mass Spectrometry of Pesticides and Pollutants. CRC Press, Inc., Cleveland, OH.
150. Strickland, J. D. H.,, and T. R. Parsons. 1960. A Practical Handbook of Seawater Analysis. Fisheries Research Board of Canada, Ottawa, Canada.
151. Waller, G. R. (ed.). 1972. Biochemical Applications of Mass Spectrometry. John Wiley & Sons, Inc., New York, NY.
152. Armstrong, F. A. J. 1963. Determination of nitrate in water by ultraviolet spectrophotometry. Anal. Chem. 35:1292.
153. Baca, P.,, and H. Freiser. 1977. Determination of trace levels of nitrates by an extraction-photometric method. Anal. Chem. 49:22492250.
154. Belay, N.,, R. Sparling,, B.-S. Choi,, M. Roberts,, J. E. Roberts,, and L. Daniels. 1988. Physiological and 15NNMR analysis of molecular nitrogen fixation by Methanococcus thermolithotrophicus, Methanobacterium bryantii, and Methanospirillum hungatei. Biochim. Biophys. Acta 971:233245.
155.Bio-Rad Laboratories. 1994. Bio-Rad DC Protein Assay. Bulletin 1731. Bio-Rad Laboratories, Richmond, CA.
156.Bio-Rad Laboratories. 2005. RC DC Protein Assay. Bulletin 2610. Bio-Rad Laboratories, Richmond, CA.
157.Bio-Rad Laboratories. 2003. Colorimetric Protein Assays. Tech note 1069. Bio-Rad Laboratories, Richmond, CA.
158. Bradford, M. M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72:248254.
159. Bremner, J. M. 1960. Determination of nitrogen in soil by the Kjeldahl method. J. Agric. Sci. 55:1133.
160. Canney, P. J.,, D. E. Armstrong,, and J. H. Wiersma. 1974. Determination of Nitrite and Nitrate Ions in Natural Waters Using Aromatics or Diamines as Reagents. Technical Report. University of Wisconsin Water Resources Center, Madison.
161. Chial, H. J.,, H. B. Thompson,, and A. G. Splittgerber. 1993. A spectral study of the charge forms of Coomassie blue G. Anal. Biochem. 209:258266.
162. Chow, T. J.,, and M. S. Johnstone. 1962. Determination of nitrate in sea water. Anal. Chim. Acta 27:441446.
163. Coakley, W. T.,, and C. J. James. 1978. A simple linear transformation for the Folin-Lowry protein calibration curve to 1.0 mg/ml. Anal. Biochem. 85:9097.
164. Gornall, A. G.,, C. S. Bardawill,, and M. M. David. 1949. Determination of serum protein by means of the Biuret reaction. J. Biol. Chem. 177:751756.
165. Guiraud, G.,, J. C. Fardeau,, G. Llimous,, and M. A. Barral. 1977. Determination of nitrate in soils and plants by the Kjeldahl method. Ann. Agron. 28:329333.
166. Hardy, R. W. F.,, and A. H. Gibson (ed.). 1977. A Treatise on Dinitrogen Fixation. Section IV. Agronomy and Ecology. John Wiley & Sons, Inc., New York, NY.
167. Hill, H. D.,, and J. G. Straka. 1988. Protein determination using bicinchoninic acid in the presence of sulfhydryl reagents. Anal. Biochem. 170:203208.
168. Janssen, F. W.,, A. J. Lund,, and L. E. Anderson. 1957. Colorimetric assay for dipicolinic acid in bacterial spores. Science 127:2627.
169. Kalb, V. F., Jr.,, and R. W. Bernlohr. 1977. A new spectrophotometric assay for protein in cell extracts. Anal. Biochem. 82:362371.
170. Kalckar, H. M. 1947. Differential spectrophotometry of purine compounds by means of specific enzymes. I. Determination of hydroxypurine compounds. J. Biol. Chem. 167:429475.
171. Kruse, J. M.,, and M. G. Mellon. 1953. Colorimetric determination of ammonia and cyanate. Anal. Chem. 25: 11881192.
172. Layne, E. 1957. Spectrophotometric and turbidometric methods for measuring proteins. Methods Enzymol. 3:447454.
173. Lowry, O. H.,, N. J. Rosebrough,, A. L. Farr,, and R. J. Randall. 1951. Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193:265275.
174. Markwell, M. A.,, S. M. Haas,, L. L. Bieber,, and N. E. Tolbert. 1978. A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Anal. Biochem. 87:206210.
175. Martin, H. H.,, and J. W. Foster. 1958. On the chromatographic behavior of dipicolinic acid. Arch. Mikrobiol.31:171178.
176. Matsunaga, K.,, and M. Nishimura. 1969. Determination of nitrate in sea water. Anal. Chim. Acta 45:350353.
177. McClard, R. W. 1981. Removal of sulfhydryl groups with 1,3,4,6-tetrachloro-3a-6a-diphenylglycoluril: application to the assay of protein in the presence of thiol reagents. Anal. Biochem. 112:278281.
178. Mubarak, A.,, R. A. Howald,, and R. Woodriff. 1977. Elimination of chloride interferences with mercuric ions in the determination of nitrates by the phenol disulfonic acid method. Anal. Chem. 49:857860.
179. Murrell, W. G., 1969. Chemical composition, p. 249251. In G. W. Gould, and A. Hurst (ed.), The Bacterial Spore. Academic Press, Inc., New York, NY.
180. Murrell, W. G.,, and A. D. Warth,. 1965. Composition and heat resistance of bacterial spores, p. 124. In L. L. Campbell, and H. O. Halvorson (ed.), Spores III. American Society for Microbiology, Washington, DC.
181. Nakamura, H.,, and J. J. Pisano. 1976. Sensitive fluorometric assay for proteins. Use of fluorescamine and membrane filters. Arch. Biochem. Biophys. 172:102105.
182.Pierce Chemical Co. 2003. Bicinchoninic Acid (BCA) Method. Pierce Chemical Co., Rockford, IL.
183.Pierce Chemical Co. 2003. The Lowry Method. Pierce Chemical Co., Rockford, IL.
184.Pierce Chemical Co. 2003. Coomassie Dye Binding Method. Pierce Chemical Co., Rockford, IL.
185. Pierce, J.,, and C. H. Suelter. 1977. An evaluation of the Coomassie brilliant blue G-250 dye-binding method for quantitative protein determination. Anal. Biochem. 81:478480.
186. Postgate, J. R. (ed.). 1971. The Chemistry and Biochemistry of Nitrogen Fixation. Plenum Press, New York, NY.
187. Raganowicz, E.,, and A. Niewiadomy. 1976. Colorimetric determination of the nitrites content in water by means of 2-sulfanilamidothiazole method. Pol. Arch. Hydrobiol. 23:14.
188. Rexroad, P. R.,, and R. D. Cathey. 1976. Pollutionreduced Kjeldahl method for a crude protein. J. Assoc. Off. Anal. Chem. 59:12131217.
189. Russel, J. A. 1944. The colorimetric estimation of small amounts of ammonia by the phenol-hypochlorite reaction. J. Biol. Chem. 156:457461.
190. Sedmak, J. J.,, and S. E. Grossberg. 1977. A rapid, sensitive, and versatile assay for protein using Coomassie brilliant blue G-250. Anal. Biochem. 79:544552.
191. Smith, P. K.,, R. I. Krohn,, G. T. Hermanson,, A. K. Mallia,, F. H. Gartner,, M. D. Provenzano,, E. K. Fujimoto,, N. M. Goeke,, B. J. Olson,, and D. C. Klenk. 1985. Measurement of protein using bicinchoninic acid. Anal. Biochem. 150:7685.
192. Snell, F. D.,, and C. T. Snell. 1945. Colorimetric Methods of Analysis. D. VanNostrand Co., Inc., New York, NY.
193. Stickland, H. L. 1951. The determination of small quantities of bacteria by means of the Biuret reaction. J. Gen. Microbiol. 5:698703.
194. Warburg, O.,, and W. Christian. 1942. Isolierung und Kristallisation des Garungsferments enolase. Biochem. Z. 310:384421.
195. Warth, A. D.,, D. F Ohye,, and W. C. Murrell. 1963. The composition and structure of bacterial spores. J. Cell Biol. 16:579592.
196. Work, E. 1963. α,ϵ-Diaminopimelic acid. Methods Enzymol. 6:624634.
197. Christian, G. D.,, and F. J. Feldman. 1970. Atomic Absorption Spectroscopy. Applications in Agriculture, Biology and Medicine. Wiley-Interscience, New York, NY.
198. Heirman, R.,, and C. T. J. Allkemade,. 1963. Chemical Analysis by Flame Photometry, 2nd ed. Translated by P. T. Gilbert, Jr. Interscience Publishers, Inc., New York, NY.
199. Morrison, G. H. (ed.). 1956. Trace Analysis. Interscience Publishers, Inc., New York, NY.
200. Snell, F. D.,, and C. T. Snell. 1963. Colorimetric Methods of Analysis, vol. 2, 3rd ed. D. Van Nostrand Co., Inc., Princeton, NJ.
201.U.S. Environmental Protection Agency. 1974. Methods for Chemical Analysis of Water and Wastes. Office of Technology Transfer, Washington, DC.
202. Weinberg, E. D. (ed.). 1977. Microorganisms and Minerals. Marcel Dekker, Inc. New York, NY. Specific References
203. Stewart, M.,, A. P. Somlyo,, A. V. Somlyo,, H. Shuman,, J. A. Lindsay,, and W. C. Murrell. 1980. Distribution of calcium and other elements in cryosectioned Bacillus cereus T spores, determined by high-resolution scanning electron probe X-ray microanalysis. J. Bacteriol. 143:481491.


Generic image for table

Comparison of analytical techniques for analysis of nonprotein nitrogen compounds

For further information and sensitivity, see analysis listed under appropriate nitrogen class.

Citation: Daniels L, Hanson R, Phillips J. 2007. Chemical Analysis, p 462-503. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch18
Generic image for table

Comparison of analytical techniques available for protein assay

Sensitivity is given in micrograms per ml of solution in the assay. This can be converted easily to micrograms per ml of original solution by accounting for the dilution factors. When a range is given, both the extrasensitive microassay and the routine, less sensitive assays are described. Interfering substances may reduce this sensitivity but still allow accurate assays in an appropriate range.

DTT, dithiothreitol.

Glucose, Tris buffer, and ammonium sulfate can cause interference but can be corrected with proper blanks or pH adjustment.

Citation: Daniels L, Hanson R, Phillips J. 2007. Chemical Analysis, p 462-503. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch18

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