Two Urinary Catecholamine Measurement Indices for Applied Stress Research: Effects of Time and Temperature until Freezing

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INTRODUCTION

There is increasing evidence that stress at work can have detrimental effects on performance, well-being, and health (Cooper, 1998; Ganster & Perrewe, 2001; Kahn & Byosiere, 1992; Marmot & Wilkinson, 1999; Sonnentag & Frese, 2005). However, the vast majority of studies in occupational stress research have used self-reports for measures of both independent (stressors) and dependent variables (e.g., strain; see Kahn & Byosiere, 1992; Sonnentag & Frese, 2005; Zapf, Dormann, & Frese, 1996). Thus stressor-strain relationships may be overestimated because of correlated measurement error (common method variance; see Semmer, Zapf, & Greif, 1996).

Many authors therefore recommend measuring independent and dependent variables with different methods (multimethod approach; see Kahn & Byosiere, 1992). Physiological measures are good candidates for such an approach. One cannot regard them as "the" more objective measures, given that they also suffer from typical errors such as artifact susceptibility and measurement error attributable to devices, detection range of analytical procedures, handling of instruments, occasional influences, and so forth (see Beehr, 1995; Fried & Ferris, 1987). Because these errors are not correlated with errors of self-report, relationships may be underestimated, as Semmer et al. (1996) have shown for job observation methods (which are another candidate for alternatives to self-report). Nevertheless, if handled carefully, physiological measures do offer the potential to avoid common method variance, to yield better estimates of relationships, and to increase the understanding of the processes involved.

However, as soon as one leaves the laboratory and moves into the field, a number of pitfalls exist that might lead to serious errors, thus hampering the interpretability of field study results. One of these potential sources of errors concerns the necessity for storage and transportation (Lundberg, Melin, Fredrikson, Tuomisto, & Frankenhaeuser, 1990). Clinical chemistry shows that it is not possible to assign fixed correction factors for such interferences to urine analysis. Therefore it is recommended that samples be acidified immediately and frozen as soon as possible (Shoup, Kissinger, & Goldstein, 1984). What seems to be feasible, however, is to keep the samples at refrigerator temperature for a while before freezing them. Thus Boomsma, Alberts, van Eijk, Man in 't Veld, and Schalekamp (1993) showed that catecholamines (CAs) are stable at 4[degrees]C in unpreserved urine for 1 month, a finding that was recently replicated for a 10-hr storage period by Miki and Sudo (1998). Miki and Sudo also stored samples at room temperatures; here they found (a) a strong decrease in CA values in unpreserved samples and (b) a tendency toward increasing, rather than decreasing, values in acidified samples. These increases were, however, within a range of 10% for a delay of 1 day.

From the study by Miki and Sudo (1998), it seems clear that (a) immediate freezing is essential for unpreserved samples and (b) delays of more than a day at room temperature are risky for both acidified and unpreserved specimens. More refined studies are needed, however, to clarify whether results are robust with regard to delays until freezing of up to 24 hr. This is a crucial time period for field studies: Freezing within 24 hr usually can be guaranteed under field conditions. Delays of several hours, however, are often difficult to avoid in field work in which urine samples are picked up at the participants' homes of workplaces in different areas (Elfering, Grebner, Semmer, & Gerber, 2002; Grebner, 2001).

Because this time lag of up to 24 hr before freezing is so crucial, we tested the role of delay as a potential source of error and compared it with the measurement error that arises from the accuracy of the laboratory analysis itself. In this way, the consequences of different violations of the typical recommendations can be tested, and their effects on the trustworthiness of field results with regard to catecholamines can be estimated.

Measurement Error

High performance liquid chromatography (HPLC) with electrochemical detection has become the standard laboratory procedure of analysis. The inter- and intraassay deviation is usually reported to be lower than 10% when standardized probes are analyzed. Therefore, we expect errors for parallel samples that are immediately acidified (pH 3) and frozen (-20[degrees]C) to be normally distributed and to be below 10%, independent of gender and voiding time. Intraclass correlation coefficients should be greater than r = .80.

Storage until Freezing

Urine samples are usually acidified to a pH level of about 3 as quickly as possible and frozen until laboratory analysis. If they cannot be frozen immediately, the acidified samples should be stored at 2[degrees] to 8[degrees]C until freezing. This procedure is recommended in order to inhibit growth of bacteria and hydrolysis, which could cause a substantial decrease in the concentration of hormones (Colombo, 1994). However, as mentioned, Miki and Sudo (1998) found that storage at room temperature was associated with slight increases in CA levels after 1 day, increasing even more over 1 week, when samples were strongly acidified (either 0.5 or 1.0 pH). This might be attributable to evaporation.

In the present study, we compared 24-hr storage at 5[degrees]C (refrigerator) with storage at room temperature (25[degrees]C, electric light). Bacterial activity, hydrolysis, and evaporation should be more evident at the higher storage temperature. If temperature is an important source of bias within this period, deviations caused by high-temperature conditions should be higher than those caused by the genuine measurement variance. However, compared with a room temperature sample, a refrigerator-stored sample should correspond more closely to immediately frozen samples, given that refrigerator storage has been shown to preserve CA for up to 1 year in acidified specimens (Boomsma et al., 1993). Storage at room temperature was varied in four 8-hr steps (0, 8, 16, and 24 hr) until freezing at -20[degrees]C. Bacterial activity, hydrolysis, and evaporation should cause some systematic linear or curvilinear changes in time. We therefore tested for trends.

Indices of Hormone Concentration

Standardization of urinary CA levels may refer to (a) body weight (or, alternatively, body volume, expressed in square meters of body surface) or (b) concentration of urinary creatinine. The first index is useful when urine volumes and voiding intercepts are controlled (e.g., when all urine in 24 hr is collected) because otherwise there would be a great impact of individual water consumption and metabolism on hormone concentration. Creatinine is a waste product in the blood created by normal breakdown of muscles during activity. Therefore it depends on muscle mass, which is correlated with age, height, and body weight. The kidneys take creatinine out of the blood and into …