11.2 Collecting Bacteriological, Biological, Chemical & Physical Samples

11.2 Collecting Bacteriological, Biological, Chemical & Physical Samples

2025 WPI alignment: This section teaches collecting samples for bacteriological, biological, chemical, and physical analyses in Laboratory Analysis, the 15-question area containing 3 recall, 12 application, and 4 calculation items.

Why this laboratory task matters

Representative sampling begins before the bottle is filled. Operators choose the permit/SOP location and sample type, prevent contamination, use correct containers and preservatives, measure field parameters promptly, maintain custody, and meet method-specific transport conditions.

Analytical foundation

Laboratory workflow

1. Read the permit and SOP for location, type, frequency, timing, container, volume, preservation, and analysis deadline.
2. Inspect the site for safety, representativeness, backwater, solids deposits, turbulence, and access.
3. Label containers before collection with unique identity and required information without contaminating them.
4. Collect in the specified order and manner, preserve immediately when required, and record field measurements and actual conditions.
5. For composites, verify aliquot delivery, flow/time pacing, ice/refrigeration, tubing cleanliness, total volume, and start/stop times.
6. Seal, cool/protect, document custody, transport promptly, and notify the laboratory of unusual matrices or deviations.

Quality and diagnostic evidence

Calculation and interpretation

WPI’s formula table gives a composite sample single portion relationship using instantaneous flow, total desired sample volume, number of portions, and average flow. Use a consistent flow basis and verify that the sum of portions will fill the container without overflow. Flow-proportional compositing changes aliquot volume with flow; time-proportional collection uses equal time spacing when allowed.

Worked laboratory scenario

A 24-hour composite bottle contains only half the expected volume because the suction line clogged overnight. Dividing the existing result by two does not create the missing time periods. The operator documents the sampler failure, protects the sample record, informs the responsible laboratory/compliance staff, and follows the permit/SOP decision for recollection or qualification.

Common exam traps

• No single container or preservative is correct for every analyte.
• Refrigerating a late sample does not reverse elapsed holding time.
• A composite with missed portions may be unrepresentative even if enough volume remains for analysis.
• Never rinse a sterile bacteriological bottle that contains a prepared dechlorinating agent.

Field-to-exam checklist

• Sampling requirements come from the permit, approved method, and SOP.
• Match grab/composite type, location, container, preservation, and time to the analyte.
• Verify automatic sampler function, not just programming.
• Document deviations honestly and preserve chain of custody.

Representativeness before precision

A perfectly analyzed unrepresentative sample answers the wrong question. Examine whether the sampling point is well mixed, whether deposits or a side stream bias collection, and whether the sampler draws during every required condition. Automatic equipment should be challenged with measured aliquot volumes and a time or flow record. When process flow changes rapidly, record the exact relationship between each grab or composite portion and the flow data used to interpret it.

Compositing arithmetic and container practice

Composites are built one of three ways, and only the permit says which is acceptable: constant time interval with aliquot volume proportional to flow, constant volume of flow between constant-size aliquots, or constant time interval with constant aliquot volume. The first two are flow-proportional; the third is time-paced and represents the operating period only when flow is relatively steady.

WPI's formula table supplies the single-portion relationship: portion volume = (instantaneous flow x total desired sample volume) / (number of portions x average flow).

Worked portion calculation. Instantaneous flow is 6.0 MGD, average flow is 4.0 MGD, the program takes 24 portions, and the composite bottle is to hold 4,000 mL. Portion = (6.0 x 4,000) / (24 x 4.0) = 24,000 / 96 = 250 mL. A time-paced program would have drawn 4,000 / 24 = 167 mL at that same moment, so the flow-proportional program weights the high-flow hour by 50 percent more — which is the entire point when load, not concentration, is what the permit limits.

Container practice ruins more samples than analysis does.

• Never pre-rinse a bottle that contains a preservative or a sterile dechlorinating tablet; rinsing discards the reagent the method depends on.
• Fill BOD and dissolved-oxygen containers gently and without entraining air, since air is exactly what the test measures.
• Leave the headspace a method calls for, and leave none where volatile constituents are involved.
• Ice, not merely a closed cooler, is what reaches the required temperature during transport; record the temperature at laboratory receipt, because that record is what demonstrates the condition later.

Where a sample point is not well mixed — a quiescent corner, a backwater, a spot immediately downstream of a side stream — no amount of correct bottle handling makes the result representative.