7.4 Sample Preservation, Data Recording, Shipping, and Chain of Custody

Sample Preservation, Holding Times, and Quality Control Blanks

In CESSWI General Principles Review Manual Section 9.7 (SAOP 8 Test Objective 6), sample integrity between field collection and laboratory analysis relies on three pillars: thermal/chemical preservation, holding time compliance, and field QA/QC blanks and duplicates.

1. Thermal, Chemical, and Physical Preservation Methods

• Thermal Preservation ($4^\circ\text{C}$ on Wet Ice): Immediately after collection and capping, laboratory samples requiring cooling (such as $\text{TSS}$, laboratory turbidity, nutrients, oil and grease, and bacteria) must be placed inside an insulated cooler packed with double-bagged wet ice to chill and maintain the samples at $4^\circ\text{C}$ (accepted regulatory receipt range $> 0^\circ\text{C}$ to $\le 6.0^\circ\text{C}$ without freezing) in the dark.
  • Why Wet Ice Instead of Dry Ice or Loose Blue Ice Packs: Wet ice reliably maintains a $0^\circ\text{C}$ to $4^\circ\text{C}$ melt equilibrium. Dry ice (solid $\text{CO}_2$ at $-78.5^\circ\text{C}$) is prohibited for water samples because it freezes and shatters glass bottles and can infiltrate caps to carbonate and acidify water samples. Unassisted blue gel packs warm rapidly in hot vehicles and frequently fail the $\le 6.0^\circ\text{C}$ lab receipt gate.
  • Temperature Blank: Every cooler must contain a dedicated temperature blank—a small unpreserved bottle of water packed among the sample bottles. When the cooler arrives at the laboratory, the sample custodian measures the temperature blank to verify that samples remained at $\le 6.0^\circ\text{C}$ during transit without opening and contaminating the actual analytical samples.
• Chemical Preservation:
  • Nitric Acid ($\text{HNO}_3$) to $\text{pH} < 2$: Added to Total Recoverable Metals samples in acid-cleaned HDPE bottles. Acidification keeps metal ions in solution so they do not precipitate as metal hydroxides or adsorb onto the plastic bottle walls. (Note: When analyzing Dissolved Metals, the sample must be filtered in the field through a $0.45\text{ }\mu\text{m}$ pore-size membrane filter before adding $\text{HNO}_3$; adding acid before filtering dissolves suspended clay-bound metals and ruins the dissolved measurement!)
  • Sulfuric Acid ($\text{H}_2\text{SO}_4$) to $\text{pH} < 2$ (plus cooling to $4^\circ\text{C}$): Added to Nutrient samples (Total Phosphorus, Ammonia, Total Kjeldahl Nitrogen, combined Nitrate+Nitrite), Chemical Oxygen Demand ($\text{COD}$), and Oil and Grease (which may use $\text{H}_2\text{SO}_4$ or $\text{HCl}$) to halt bacterial metabolism.
  • Sodium Thiosulfate ($\text{Na}_2\text{S}_2\text{O}_3$): Pre-dosed in sterile bacteria bottles to neutralize any trace residual chlorine that would otherwise continue killing indicator bacteria inside the bottle during transit.
• Physical Zero-Headspace Preservation: Samples for Volatile Organic Compounds ($\text{VOCs}$) are collected in $40\text{ mL}$ glass VOA vials with PTFE-lined septa caps (preserved with $\text{HCl}$ and cooled to $4^\circ\text{C}$). The vial is filled to a convex dome meniscus and capped so that zero headspace (zero air bubbles) remains when inverted; even a pea-sized air bubble allows volatile organics to partition out of the water phase.

2. High-Yield Maximum Holding Times (40 CFR Part 136)

The holding time clock starts at the collection time; for an automated composite, 40 CFR 136 Table II footnote 3 generally starts the holding time at the end of collection, while manual compositing generally starts at the first grab. Immediate field parameters and particular methods have separate requirements:

1. Immediate Field Analysis (Within 15 Minutes): pH, Temperature, Dissolved Oxygen ($\text{DO}$), and Total Residual Chlorine.
2. 6 to 8 Hours (Max 24 Hours for Specific Non-Potable Methods): Microbiological Indicator Bacteria (E. coli, Fecal Coliform, Enterococci) cooled at $< 10^\circ\text{C}$ ($4^\circ\text{C}$).
3. 48 Hours: Laboratory Turbidity (stored in the dark at no more than 6°C), Settleable Solids (cooling is not specified in Table II), Biochemical Oxygen Demand ($\text{BOD}_5$) ($4^\circ\text{C}$), and Unpreserved Nitrate ($\text{NO}_3^-$) ($4^\circ\text{C}$).
4. 7 Days: Total Suspended Solids ($\text{TSS}$) and Total Dissolved Solids ($\text{TDS}$) cooled at $4^\circ\text{C}$ (no chemical preservative).
5. 28 Days: Oil and Grease ($\text{H}_2\text{SO}_4$ or $\text{HCl}$ to $\text{pH} < 2$, $4^\circ\text{C}$), Total Phosphorus / Ammonia / Preserved Nitrate+Nitrite ($\text{H}_2\text{SO}_4$ to $\text{pH} < 2$, $4^\circ\text{C}$), and Mercury by routine CVAA (nitric acid to pH < 2); CVAFS has different preservation and a 90-day holding time.
6. 6 Months: Total and Dissolved Metals (all metals except Mercury, preserved with $\text{HNO}_3$ to $\text{pH} < 2$).

3. Field Quality Assurance / Quality Control (QA/QC) Samples

To prove to regulators that field data are both precise and free from artificial contamination, SAPs incorporate three standard field QA/QC sample types:

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Recording Field Sampling Data and Bottle Labeling

Indelible Bottle Labeling Requirements the approved practice detail

Every sample container must carry a waterproof adhesive label completed in indelible, waterproof ink (permanent waterproof marker or ballpoint pen—never pencil or water-soluble felt pen). Best practice is to write the label and affix it to the dry bottle before immersing the bottle, then fill in the exact minute of collection immediately upon capping. Every bottle label must display seven mandatory data elements:

1. Project / Site Name and Permit Number
2. Sampling Location / Station ID (e.g., Outfall-001, Creek-Upstream)
3. Unique Sample Identification Number (matching the Chain of Custody line item)
4. Date and Exact Time of Collection (written in 24-hour military time, such as 15:20, to eliminate AM/PM confusion and establish the legal holding-time start)
5. Sample Collector's Initials or Full Name
6. Requested Analytical Parameter(s) and Sample Type (Grab vs. Composite)
7. Preservative Used (e.g., HNO3 to pH < 2, H2SO4 to pH < 2, Cool 4C, or None)

Contemporaneous Entries in a Bound Field Logbook

Bottle labels alone are never sufficient. At the time of collection, the CESSWI must record complete sampling metadata in a bound, weatherproof (Rite in the Rain) field logbook with pre-numbered pages using indelible ink:

• Correction Protocol: If a writing error occurs in a field logbook or on a Chain of Custody form, never erase, scribble out, tear out a page, or use correction fluid (white-out). Draw a single horizontal line through the incorrect entry so it remains legible, write the correct value next to it, and sign your initials and the date beside the correction.
• Required Field Notes: Weather conditions (air temperature, wind, precipitation type), current and 24-hour on-site rain gauge depth, storm start time, estimated or gauged outfall discharge rate ($Q$) / flow depth, visual and olfactory water characteristics (color, clarity, odor, foam, floatables, oil sheen), all field meter readings ($\text{NTU}$, $\text{pH}$, temperature, $\text{DO}$) with confirmation of daily calibration/drift checks, and time-stamped photographs of the outfall and labeled bottles.

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Chain of Custody (COC) Procedures and Sample Shipping

Definition and Legal Purpose of Chain of Custody the approved practice detail

The Chain of Custody (COC) is the unbroken, chronological legal document that tracks the possession, handling, and transfer of environmental samples from the exact moment of field collection through transport, laboratory receipt, internal analysis, and final disposal. Because stormwater sample results can trigger six-figure Clean Water Act civil penalties or criminal prosecution, opposing attorneys routinely scrutinize the COC first: any unexplained gap in custody, missing signature, mismatched bottle ID, or broken custody seal can render the laboratory results legally inadmissible in court.

The Four Legal Conditions of "In Custody"

Under EPA and environmental forensic standards, a stormwater sample is normally documented as under a person's custody when at least one of the following four conditions is satisfied:

1. Actual Physical Possession: The sample is physically held by or carried on the person of the authorized custodian.
2. Direct Line of Sight: The sample is in the custodian's unobstructed, continuous view after having been in their physical possession.
3. Locked in a Controlled Secure Area: The sample was placed by the custodian inside a locked vehicle, locked field cabinet, or locked refrigerator to which only authorized personnel control access.
4. Sealed in a Tamper-Evident Container: The sample is inside a designated container (such as a shipping cooler) sealed with signed and dated tamper-evident Custody Seals across the lid seams so that the container cannot be opened without visibly tearing or breaking the seal.

Executing Custody Transfers and Commercial Courier Shipping

• Person-to-Person Custody Transfer: Whenever samples are handed from one person to another (for example, from the field CESSWI to a courier or directly to the laboratory sample reception desk), both individuals must sign the Chain of Custody form at the moment of transfer:
  • The person giving up the samples signs the "Relinquished by" block, prints their name and company, and records the exact date and 24-hour time.
  • The person taking possession inspects the bottles against the COC lines, signs the "Received by" block, prints their name and organization, and records the exact date and 24-hour time.
• Shipping Samples via Commercial Courier (FedEx / UPS Protocol): When samples are shipped to an off-site laboratory via a commercial parcel carrier (such as FedEx or UPS), the delivery driver will not open the cooler or sign the internal COC form. To preserve an unbroken legal Chain of Custody during commercial shipment, the CESSWI must execute five steps:
  1. Package the Cooler: Wrap glass bottles in bubble wrap, verify all caps are tight, place bottles upright inside a plastic-lined cooler alongside the temperature blank, pack double-bagged wet ice around the bottles (never loose ice that melts into water and soaks off labels), and add absorbent padding.
  2. Sign and Bag the COC Inside the Lid: Complete all sample lines on the COC form, sign the "Relinquished by" box with the date and time, retain the sampler's carbonless copy, seal the original signed COC form inside a waterproof zip-top plastic bag, and tape the bagged COC form to the underside of the cooler lid.
  3. Apply Tamper-Evident Custody Seals: Close and tape the cooler drain plug and lid shut, then affix at least two adhesive paper Custody Seals—signed and dated in indelible ink by the inspector—across opposite lid seams/hinges so the cooler lid cannot be lifted without ripping the seals (cover the seals with clear packing tape to protect the ink from rain while ensuring the paper seal still tears if peeled).
  4. Record the Courier Airbill / Tracking Number: Write the commercial courier's airbill / tracking number on the COC copy and in the field logbook to document transit custody.
  5. Laboratory Receipt Verification: Upon arrival at the lab, the laboratory sample custodian inspects the exterior Custody Seals to confirm they are intact, opens the cooler, measures the temperature blank ($\le 6.0^\circ\text{C}$), verifies headspace and preservative $\text{pH}$, and signs "Received by Laboratory" with the exact date and time.

End-to-End Stormwater Sampling, Calibration, Preservation, and Chain of Custody Workflow

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Apply the analytical method and sampling plan

For the detailed method and approval qualifications, review the related CESSWI explanation.

For the detailed method and approval qualifications, review the related CESSWI explanation.