The authors thank Drs. Bobby J. Presley, Robert Tayloy, Paul Boothe, Mr. Bryan Brattin, and Mr. Mike Metcalf for their assistance during the laborious field sampling and lab work for the practical development and application of "clean techniques".

1.サンプラーの準備<ol><li>サンプラー<ol><li>サンプラーの集合<ol><li> 1.5メートル耐薬品性シリコーン圧送管（OD 0.635センチメートル）に4メートルの長いフッ化エチレンプロピレン（FEP）チューブ（ID 0.635センチメートル、OD 0.95センチメートルまたは類似）を接続します。 </li><li>ポンピングチューブにポリプロピレンYコネクタを挿入し、他の一つの出口、及び（20 cmの圧送管によって）0.45μmのカプセルフィルターに50 cmのポンピングチューブを接続します。 </li><li>それらが清掃された後、クリーンルーム（ベンチ）にチューブを組み立てます（下記参照）、およびポリエチレン袋の2層に集合体を格納します。 </li></ol></li><li>サンプラークリーニング<ol><li>ラボ用洗剤液でチューブセット（蠕動ポンプの上に1.5メートルのポンプチューブを取り付けることにより）を記入し、24時間のためにそれを浸します。 10％（v / v）の塩酸（試薬グレード）とそれを記入し、48時間浸し、その後、脱イオン水でセットチューブを洗い流します。 </li><li>徹底的に脱イオンワットで設定したチューブをフラッシュ数回ATER、プラスチック袋に集合体を格納します。 24時間50％（v / v）のHNO 3（試薬グレード）溶液中でそれらを浸すことによって、Yコネクタと短い揚水管を清掃してください。 </li></ol></li><li>カプセルフィルター<ol><li>最初の脱イオン水で48時間の浸漬工程のため、その後、10％（v / v）のHCl溶液を、それらをフラッシュすることによって、カプセルフィルターを清掃してください。 </li><li>酸浸漬した後、脱イオン水でフィルターをフラッシュし、酸を中和するために、各フィルターに21％（重量）NH 4 OH（サブゆで）溶液1mlを追加します。 </li><li>ポリエチレンジッパー付きの袋に30-cmのクリーンポンプ入口と出口を接続管、および店舗のループで個々のフィルタをシール。 </li></ol></li></ol></li><li>サンプル容器<ol><li>ポリエチレン（PE、1,000ミリリットル）とFEP（500ミリリットルと千ミリリットル）微量金属決意用容器のためのボトルを使用してください。 <ol><li> （1％）第一の界面活性剤でそれらを浸すことによってボトルをきれいにし、その後、50％（v / v）のHNO 3（試薬グレード）、および24、48および24時間、10％（v / v）の塩酸（試薬グレード）の溶液を、それぞれ、および2つの浸漬工程の間に脱イオン水でボトルをすすぎます。 </li><li>最終塩酸浸漬した後、完全に脱イオン水（DIW）でボトルをすすぎ、クリーンルームやクラス100クリーンベンチ内（密封されたキャップ付き）ボトルを乾燥させます。 </li><li>シールは、輸送のためのポリエチレン袋にポリエチレンジッパー付きの袋にペットボトルや二重袋にそれらを掃除しました。 </li></ol></li><li>溶存有機炭素（DOC）決意のためにきれいなガラスびん<ol><li> 48時間、10％のHClで、溶存有機炭素（DOC）の分析のために、琥珀色ホウケイ酸ガラスボトル（40 ml）に浸し。脱イオン水で洗浄したガラスボトルをすすぎ、そして使用前に2時間480℃でそれらを燃焼。輸送のためのジッパー付きポリエチレン袋に個別のボトルを密封します。 </li></ol></li></ol></li></ol> 2.サンプリング<ol><li>水サンプル採取<ol><li> UPOn個のサンプリングサイトに到着、外袋のマークサンプル識別番号と、元の袋にボトルを保持します。 </li><li>川の土手で、またはボートに乗って、1人はサンプラーで袋を開けていると3メートルのポリプロピレンポール（洗浄）に4メートルのFEPチューブを取り付けます。できるだけ銀行からポールを伸ばし、30cm程度のポンプがオンになっている前に、河川の流水の表面下のFEPチューブの入口を保ちます。 </li><li>別の人は、蠕動ポンプ（内蔵バッテリ付きサンプリングポンプ）のポンプヘッドにポンプチューブを取り付ける必要があります。ポンプを起動し、水（下流側）総サンプラー容積の少なくとも3倍のドレイン。パウダーフリーの手袋を着用して、サンプル瓶を充填開始するボトルとキャップの袋を開きます。 <ol><li>別の人が利用可能な場合あるいは、第三者がインナーバッグやサンプルのキャップを開け、aとbに試料を排出するサンプリングチューブを保持する担当してきましたottle。 </li></ol></li><li>フィールド内の導電率、温度およびpHの測定のための125ミリリットルのペットボトルでフィルタリングされていないサンプルを収集します。 </li><li> （1（フィルタなしコンセントを介して）最初の（500-mlまたは粒子状のサンプル収集のための千ミリリットル）フィルタ処理されていないサンプルを収集した後、ろ過水のサンプルを収集するためにカプセルフィルターを通過する水を強制的にプラスチック製のクランプでコンセントを閉じます溶解した微量金属決意用-L）。 </li><li> DOCおよびEDTA測定のための40ミリリットルのアンバーガラスびんに（フィルター付きコンセントを介して）、濾過したサンプルを収集します。 </li><li> 1,000ミリリットル/分に近い500ml /分の流速で水サンプルを収集します。圧力が（フィルタ」仕様に応じて）構築を開始したときにカプセルフィルターを交換してください。サンプルの種類ごとに、品質管理アリコートとして機能するように選択された位置で、余分なサンプルだけでなく、フィールドブランクを収集します。 </li><li>ドライアイスと保存されている私に40ミリリットルのガラスボトルを置きます氷の胸、アイスチェスト中のポリエチレンボトルのn。 </li></ol></li><li>浮遊粒子状物質の集合（SPM） <ol><li>プラスチック製濾過漏斗および容器を用いて、真空濾過により0.4μmのポリカーボネート（PC）膜フィルター（酸洗浄し、予め秤量した）にSPMを集めます。 </li><li> SPM濃度を生じ、粒状微量金属決意するための粒子サンプルを提供するために、実験室で乾燥した膜フィルタを凍結。 </li></ol></li></ol> 3.サンプルの前処理（溶存微量金属） <ol><li>溶解した微量金属の決意について、サンプル瓶に2mlの濃サブ煮HNO 3（1-Lあたりのサンプル）を追加します。 FEPボトルに溶解した微量金属試料（酸性化）を転送します。あるいは、サンプルは、FEPボトルに直接回収することができます。 24時間（8〜15ワットのUVランプ）用のFEPボトル内のサンプルをUV-照射。 </li></ol>微量金属分析のための4.前濃縮および治療<ol><li&gt;の前濃縮樹脂<ol><li>小さ ​​な（10-30 mLの容量）のプラスチックカップに陽イオン交換樹脂を2g秤量し、カップに2 N HNO 3溶液を少量加えます。 10ミリリットルの容量クロマトグラフィーカラムに樹脂を注ぎます。二回（サブゆで）5-mlの2 N HNO 3でカラムを洗浄することにより樹脂を清掃し、かつ高純度の水（HPW）で3回すすいでください。 </li><li> NH 4 +型に樹脂を変換するために、カラムに1 M NH 4 OH（サブゆで）の10ミリリットルを追加します。 </li></ol></li><li>緩衝液（1 M酢酸アンモニウム） <ol><li> HPWの約800ミリリットル中（サブゆで）氷酢酸の57ミリリットルを追加します。 〜（サブ煮21％）水酸化アンモニウムの60ミリリットルを加え、酢酸で混ぜます。 5.5への最終pHと千ミリリットルへの最終的な音量を調整します。 </li></ol></li><li>予備濃縮手順（微量金属を溶解しました） <ol><li> 1 MアンモニアACET 30mlのを添加することによって5.5±0.3に酸性化し、UV照射サンプルのpHを調整します試料に緩衝液と一部（〜2.8ミリリットル）NH 4 OH（サブゆで）を食べました。 </li><li> 〜30センチメートル陽イオン交換樹脂（4.1節）を充填したカラム上にサンプル瓶を置き、~60センチFEPチューブとクロマトグラフィーキャップとコネクタ（メスルアー）により試料瓶と前濃縮カラムを接続します。 </li><li>列の上に接続された2方コックを使用して3〜5ミリリットル/分の流量を制御します。サンプルは前濃縮カラムを通過することを許可します。サンプルがカラムを通過した後、カラムからのチューブとキャップを外してください。 </li><li> 2×5 mlのHPWと1 M酢酸アンモニウム（pHは5.5）の4×5ミリリットル、および他の微量金属からの主要な陽イオンを分離するためのHPWの2×5ミリリットルの列を扱います。 </li><li>ちょうど列の下に30 mlの酸洗浄PE瓶を置き、PEボトルに7×1-mlの2 N HNO 3（サブゆで）でカラムを洗浄します。 </li><li>重量aを取得することにより、2 N HNO 3の流出物（〜8.0ミリリットル）の容積を決定しますND 2 N HNO 3流出液の比重。 </li></ol></li><li>浮遊粒子状物質の消化<ol><li> SPMサンプルとドライPCフィルターを凍結乾燥後、それらの重量を量ります。予め計量したペルフルオロアルカン（PFA）船舶（60-mlの容量）に、フィルタと、SPMのサンプルを配置し、容器に濃HNO 3の3ミリリットルを追加します。 </li><li> 2.5キロメートルの一定のトルクに血管を締めて、12時間130℃で、従来のオーブンで消化しました。冷却後、血管を開き、容器にHFの2ミリリットルを追加します。 </li><li> 2.5キロメートルの一定のトルクに血管を締めて、12時間130℃で、従来のオーブンで消化しました。冷却後、血管を開き、容器に4.5％ホウ酸溶液の16ミリリットルを追加します。 </li><li> 2.5キロメートルの一定のトルクに血管を締めて、12時間130℃で、従来のオーブンでダイジェスト。各容器を計量し、それぞれの消化液の比重を決定します最終ダイジェスト体積を得ました。 </li><li>各フィルタ上のSPMの最終体積と重量（フィルタ上に試料の重量で割った最終ダイジェストボリューム）からダイジェスト用の希釈係数を計算します。 </li></ol></li></ol> 5.サンプルの分析<ol><li>微量金属<ol><li>事前に濃縮サンプルとフレーム原子吸光分析、黒鉛炉原子吸光分析法を用いて、微粒子からの溶液に溶解し、および/または誘導結合プラズマ中に微量金属（カドミウム、CoとCuの、鉄、マンガン、ニッケル、鉛、およびZn）濃度を決定質量分析計。 </li><li>誘導結合プラズマ発光分光分析により、前浮遊粒子状物質の予備濃縮および消化に描かれたサブサンプル中の主要イオン濃度といくつかの微量金属を、決定します。 </li></ol></li><li>補助パラメータ<ol><li>ポータブルデバイスを使用して、フィールド内の試料温度、pH、塩分濃度と導電率を決定します。 </li><li>決定溶解または赤外分光法21による二酸化炭素の検出と湿式酸化に基づいて、全有機炭素分析によりganic炭素（DOC）濃度、。確立された手順22,23以下（SPD-M10AV Diodearray検出器付き）高速液体クロマトグラフィーによる総EDTAの濃度を決定します。 </li></ol></li></ol>

<ol>
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T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparison+of+methods+for+the+measurement+of+dissolved+organic+carbon+in+natural+waters.">A comparison of methods for the measurement of dissolved organic carbon in natural waters.</a> Marine Chemistry. 54, 85-96 (1996).</li><li>Nowack, B., Kari, F., Hilger, S. U., Sigg, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+dissolved+and+adsorbed+EDTA+species+in+water+and+sediments+by+HPLC.">Determination of dissolved and adsorbed EDTA species in water and sediments by HPLC.</a> Analytical Chemistry. 68 (3), 561-566 (1996).</li><li>Bergers, P. J. M., de Groot, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+analysis+of+EDTA+in+water+by+HPLC.">The analysis of EDTA in water by HPLC.</a> Water Research. 28 (3), 639-642 (1994).</li></ol>

The authors have no conflict of interests to disclose.

天然水で信頼性の高い微量金属のデータを取得する汚染を低減することを目指して、サンプル収集、処理、前処理、および分析中に強調したよう細心の注意が必要です。濃度は、以前に報告されたものより数桁低いことができることを見出し、最後の二十年で「クリーン技術」を用いて得られた天然水中の金属濃度をトレースします。微量金属のレベルを正確にヒトおよび高等生物に有害な?...

一般的に天然水に対して得られたいくつかの微量金属の結果は、サンプル収集、トリートメントと決意1,2の間に適用される不十分な技術に起因するアーチファクトのため不正確であることが認識されてきました。溶解した微量金属の真の濃度は、（地表水3におけるnM範囲にサブnM単位）以前に発表された値よりも2桁低いまでになりました。同じ状況が改善されたサンプリングと分析方法が導入されているとして海洋水域で受け入れられた溶存微量金属濃度は過去40年ほどかけて桁違いに減少しており、海洋化学で発見されました。努力は、微量金属の分析4-8のすべての段階を通して微量金属汚染の低減又は排除を目的と「クリーン技術」の開発とデータ品質を改善する試みがなされています。周囲における微量金属濃度の決意するためレベルは、予備濃縮がしばしば要求されます。イオン交換技術8-12は、一般的に効率的な予備濃縮のために適用されています。 汚染は、容器の壁、容器の洗浄、サンプラー、サンプルの取り扱いや保管、およびサンプルの保存と分析7,13から生じ得ます。さらに最近行っきれいな方法を用いて、全ての研究は、天然水中の微量金属の濃度は、一般的によく日常的な方法7の検出限界未満であることを示しています。その水質モニタリングが15を投影するために1990年代初めの容疑者の微量金属データの認識ので、クリーンな方法は、米国EPA（環境保護庁）に組み込まれている微量金属の決意14と米国地質調査所のためのガイドラインは、クリーンな方法を採用しています。微量金属の研究のためのクリーンな方法は、しっかりと正確なデータ・ベースを作成するために、すべてのプロジェクトで採用される必要があります。 <pクラス原則として= &quot;jove_content&quot;&gt;、微量金属決意に使用する水のサンプルは、適切なサンプリング特定の材料のギアや組成物を収集、保存し、機器分析を進める前に、適当な容器および装置を用いて、適切に扱われるべきです。浮遊粒子状物質（SPM）は、サンプル保管期間中に変化を受け、水の組成を変化させることができるので、水試料からのSPMの迅速な分離は、水環境中の微量金属の研究のための一般的な方法です。天然水中の溶存微量金属濃度の決意ため、濾過が必要であり、インライン濾過技術が適切かつ効率的です。 そのような表面とグラウンド水域などの水生環境における微量金属の分布と挙動が（ 例えば 、風化）自然と人為的（ 例えば 、廃水流出物）の要因、ならびに再など他の環境条件によって影響を受けることができますgional地質学、形態、土地利用や植生、気候16-19。これは、その後、浮遊粒子状物質（SPM）、溶存有機炭素（DOC）、人為的なリガンド（ 例えば 、エチレンジアミン四酢酸、EDTA）、塩、酸化還元電位およびpH 17-20の濃度などの物理化学的パラメータの違いにつながることができます。したがって、正確な、関連する微量金属の研究は、適切な微量金属分析のための試料の採取ならびにための関連要因とパラメータの決意を必要とします。

<table border="0" cellpadding="0" cellspacing="0" width="782">
	<tbody>
		<tr height="22">
			<td height="22" style="height:22px;width:249px;">Nitric Acid</td>
			<td style="width:216px;">Seastar Chemicals</td>
			<td style="width:129px;"></td>
			<td style="width:188px;">Baseline grade</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:249px;">Ammonium hydroxide</td>
			<td style="width:216px;">Seastar Chemicals</td>
			<td style="width:129px;"></td>
			<td>Baseline grade</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:249px;">Acetic Acid</td>
			<td style="width:216px;">Seastar Chemicals</td>
			<td style="width:129px;"></td>
			<td>Baseline grade</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:249px;">Nitric Acid</td>
			<td style="width:216px;">J. T. Baker</td>
			<td style="width:129px;">9601-05</td>
			<td>Reagent grade</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:249px;">Hydrochloric acid</td>
			<td style="width:216px;">J. T. Baker</td>
			<td style="width:129px;">9530-33</td>
			<td>Reagent grade</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">Chromatographic columns</td>
			<td style="width:216px;">Bio-Rad</td>
			<td>7311550&#160;</td>
			<td>Poly-Prep</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">Column stack caps</td>
			<td style="width:216px;">Bio-Rad</td>
			<td>7311555</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">Cap connectors (female luers)</td>
			<td style="width:216px;">Bio-Rad</td>
			<td style="width:129px;">7318223</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">2-way stopcocks</td>
			<td style="width:216px;">Bio-Rad</td>
			<td>7328102</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">Cation exchange resin</td>
			<td style="width:216px;">Bio-Rad</td>
			<td>1422832&#160;</td>
			<td>Chelex-100</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">Portable sampler (sampling pump)</td>
			<td style="width:216px;">Cole Palmer</td>
			<td>EW-07571-00</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">FEP tube</td>
			<td style="width:216px;">Cole Palmer</td>
			<td>EW-06450-07</td>
			<td>6.4 mm I.D., 9.5 mm O.D.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">Pumping tube</td>
			<td style="width:216px;">Cole Palmer</td>
			<td>EW-06424-24</td>
			<td>6.4 mm I.D. C-Flex</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">Capsule filter (0.4 mm)</td>
			<td style="width:216px;">Fisher Scientific</td>
			<td>WP4HY410F0</td>
			<td>polypropylene casing</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">1 L low density polyethylene bottle</td>
			<td>NALGE NUNC INTERNATIONAL</td>
			<td>312088-0032</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:249px;">1 L (or 500 ml) FEP bottle</td>
			<td>NALGE NUNC INTERNATIONAL</td>
			<td>381600-0032</td>
			<td></td>
		</tr>
	</tbody>
</table>

clean sampling and analysis of river and estuarine waters for trace metal studies

Most of the trace metal concentrations in ambient waters obtained a few decades ago have been considered unreliable owing to the lack of contamination control. Developments of some techniques aiming to reduce trace metal contamination in the last couple of decades have resulted in concentrations reported now being orders of magnitude lower than those in the past. These low concentrations often necessitate preconcentration of water samples prior to instrumental analysis of samples. Since contamination can appear in all phases of trace metal analyses, including sample collection (and during preparation of sampling containers), storage and handling, pretreatments, and instrumental analysis, specific care needs to be taken in order to reduce contamination levels at all steps. The effort to develop and utilize "clean techniques" in trace metal studies allows scientists to investigate trace metal distributions and chemical and biological behavior in greater details. This advancement also provides the required accuracy and precision of trace metal data allowing for environmental conditions to be related to trace metal concentrations in aquatic environments.
This protocol that is presented here details needed materials for sample preparation, sample collection, sample pretreatment including preconcentration, and instrumental analysis. By reducing contamination throughout all phases mentioned above for trace metal analysis, much lower detection limits and thus accuracy can be achieved. The effectiveness of "clean techniques" is further demonstrated using low field blanks and good recoveries for standard reference material. The data quality that can be obtained thus enables the assessment of trace metal distributions and their relationships to environmental parameters.

「クリーン技術」の開発および使用には、現在よく、周囲の海域で正確な微量金属濃度を得るために、水試料中の微量金属の前濃縮が一般的であることが認識されます。天然水中の微量金属のための最も水質基準が低いμgの/ Lの範囲であるが、検出下限は、水生環境における周囲の濃度で微量金属上の地球化学的および生物学的効果を調査するために必要とされてい...

Watch this Scientific Journal Video about Clean Sampling and Analysis of River and Estuarine Waters for Trace Metal Studies at JoVE.com

Clean Sampling and Analysis of River and Estuarine Waters for Trace Metal Studies

Special care using "clean techniques" is required to properly collect and process water samples for trace metal studies in aquatic environments. A protocol for sampling, processing, and analytical procedures with the aim of obtaining reliable environmental monitoring data and results with high sensitivity for detailed trace metal studies is presented.

きれいなサンプリングと微量金属の研究のために川や河口ウォーターズの分析

Most of the trace metal concentrations in ambient waters obtained a few decades ago have been considered unreliable owing to the lack of contamination ...

clean-sampling-analysis-river-estuarine-waters-for-trace-metal

Research

JoVE Journal

Environment

11.3K ビュー.  National Sun Yat-sen University. Special care using "clean techniques" is required to properly collect and process water samples for trace metal studies in aquatic environments. A protocol for sampling, processing, and analytical procedures with the aim of obtaining reliable environmental monitoring data and results with high sensitivity for detailed trace metal studies is presented.

ビデオ: Clean Sampling and Analysis of River and Estuarine Waters for Trace Metal Studies

Integrated Field Lysimetry and Porewater Sampling for Evaluation of Chemical Mobility in Soils and Established Vegetation

Potentially toxic chemicals are routinely applied to land to meet growing demands on waste management and food production, but the fate of these chemicals is often not well understood. Here we demonstrate an integrated field lysimetry and porewater sampling method for evaluating the mobility of chemicals applied to soils and established vegetation. Lysimeters, open columns made of metal or plastic, are driven into bareground or vegetated soils. Porewater samplers, which are commercially available and use vacuum to collect percolating soil water, are installed at predetermined depths within the lysimeters. At prearranged times following chemical application to experimental plots, porewater is collected, and lysimeters, containing soil and vegetation, are exhumed. By analyzing chemical concentrations in the lysimeter soil, vegetation, and porewater, downward leaching rates, soil retention capacities, and plant uptake for the chemical of interest may be quantified. Because field lysimetry and porewater sampling are conducted under natural environmental conditions and with minimal soil disturbance, derived results project real-case scenarios and provide valuable information for chemical management. As chemicals are increasingly applied to land worldwide, the described techniques may be utilized to determine whether applied chemicals pose adverse effects to human health or the environment.

Field lysimetry and porewater sampling allow researchers to evaluate the fate of chemicals applied to soils and established vegetation. The goal of this protocol is to demonstrate how to install required instrumentation and collect samples for chemical analysis during integrated field lysimetry and porewater sampling experiments.

統合フィールドLysimetryと間隙水土壌中の化学物質の移動性の評価のためのサンプリングと設立植生

Potentially toxic chemicals are routinely applied to land to meet growing demands on waste management and food production, but the fate of these chemicals ...

環境学

Soil Sampling and Isolation of Entomopathogenic Nematodes (Steinernematidae, Heterorhabditidae)

Entomopathogenic nematodes (a.k.a. EPN) represent a group of soil-inhabiting nematodes that parasitize a wide range of insects. These nematodes belong to two families: Steinernematidae and Heterorhabditidae. Until now, more than 70 species have been described in the Steinernematidae and there are about 20 species in the Heterorhabditidae. The nematodes have a mutualistic partnership with Enterobacteriaceae bacteria and together they act as a potent insecticidal complex that kills a wide range of insect species.
Herein, we focus on the most common techniques considered for collecting EPN from soil. The second part of this presentation focuses on the insect-baiting technique, a widely used approach for the isolation of EPN from soil samples, and the modified White trap technique which is used for the recovery of these nematodes from infected insects. These methods and techniques are key steps for the successful establishment of EPN cultures in the laboratory and also form the basis for other bioassays that consider these nematodes as model organisms for research in other biological disciplines. The techniques shown in this presentation correspond to those performed and/or designed by members of S. P. Stock laboratory as well as those described by various authors.

Soil Sampling and Isolation of Entomopathogenic Nematodes Steinernematidae, Heterorhabditidae

Entomopathogenic nematodes are soil-inhabiting roundworms that parasitize a wide range of insects. We demonstrate sampling methods used for the isolation of these nematodes from the soil using two techniques: the insect baiting and the modified White trap, for the recovery of nematodes from soil samples and infected insect cadavers, respectively.

土壌のサンプリングと昆虫病原線虫の分離（Steine​​rnematidae、Heterorhabditidae）

Entomopathogenic nematodes (a.k.a. EPN) represent a group of soil-inhabiting nematodes that parasitize a wide range of insects. These nematodes belong to ...

A Whole Cell Bioreporter Approach to Assess Transport and Bioavailability of Organic Contaminants in Water Unsaturated Systems

Bioavailability of contaminants is a prerequisite for their effective biodegradation in soil. The average bulk concentration of a contaminant, however, is not an appropriate measure for its availability; bioavailability rather depends on the dynamic interplay of potential mass transfer (flux) of a compound to a microbial cell and the capacity of the latter to degrade the compound. In water-unsaturated parts of the soil, mycelia have been shown to overcome bioavailability limitations by actively transporting and mobilizing organic compounds over the range of centimeters. Whereas the extent of mycelia-based transport can be quantified easily by chemical means, verification of the contaminant-bioavailability to bacterial cells requires a biological method. Addressing this constraint, we chose the PAH fluorene (FLU) as a model compound and developed a water unsaturated model microcosm linking a spatially separated FLU point source and the FLU degrading bioreporter bacterium Burkholderia sartisoli RP037-mChe by a mycelial network of Pythium ultimum. Since the bioreporter expresses eGFP in response of the PAH flux to the cell, bacterial FLU exposure and degradation could be monitored directly in the microcosms via confocal laser scanning microscopy (CLSM). CLSM and image analyses revealed a significant increase of the eGFP expression in the presence of P. ultimum compared to controls without mycelia or FLU thus indicating FLU bioavailability to bacteria after mycelia-mediated transport. CLSM results were supported by chemical analyses in identical microcosms. The developed microcosm proved suitable to investigate contaminant bioavailability and to concomitantly visualize the involved bacteria-mycelial interactions.

A whole cell bioreporter assay with Burkholderia sartisoli RP037-mChe was developed to detect fractions of an organic contaminant (i.e., fluorene) available for bacterial degradation after active transport by mycelia bridging air-filled pores in a water unsaturated model system.

水不飽和システムにおける有機汚染物質の輸送および生物学的利用能を評価するために、全細胞バイオレポーターアプローチ

Bioavailability of contaminants is a prerequisite for their effective biodegradation in soil. The average bulk concentration of a contaminant, however, is ...

Sediment Core Sectioning and Extraction of Pore Waters under Anoxic Conditions

We demonstrate a method for sectioning sediment cores and extracting pore waters while maintaining oxygen-free conditions. A simple, inexpensive system is built and can be transported to a temporary work space close to field sampling site(s) to facilitate rapid analysis. Cores are extruded into a portable glove bag, where they are sectioned and each 1-3 cm thick section (depending on core diameter) is sealed into 50 ml centrifuge tubes. Pore waters are separated with centrifugation outside of the glove bag and then returned to the glove bag for separation from the sediment. These extracted pore water samples can be analyzed immediately. Immediate analyses of redox sensitive species, such as sulfide, iron speciation, and arsenic speciation indicate that oxidation of pore waters is minimal; some samples show approximately 100% of the reduced species, e.g. 100% Fe(II) with no detectable Fe(III). Both sediment and pore water samples can be preserved to maintain chemical species for further analysis upon return to the laboratory.

A protocol for sectioning sediment cores and extracting pore waters under anoxic conditions in order to permit analysis of redox sensitive species in both solids and fluids is presented.

無酸素状態下での堆積物コアセクショニングと間隙水の抽出

We demonstrate a method for sectioning sediment cores and extracting pore waters while maintaining oxygen-free conditions. A simple, inexpensive system is ...

Modification and Application of a Leaf Blower-vac for Field Sampling of Arthropods

Rice fields host a large diversity of arthropods, but investigating their population dynamics and interactions is challenging. Here we describe the modification and application of a leaf blower-vac for suction sampling of arthropod populations in rice. When used in combination with an enclosure, application of this sampling device provides absolute estimates of the populations of arthropods as numbers per standardized sampling area. The sampling efficiency depends critically on the sampling duration. In a mature rice crop, a two-minute sampling in an enclosure of 0.13 m2 yields more than 90% of the arthropod population. The device also allows sampling of arthropods dwelling on the water surface or the soil in rice paddies, but it is not suitable for sampling fast flying insects, such as predatory Odonata or larger hymenopterous parasitoids. The modified blower-vac is simple to construct, and cheaper and easier to handle than traditional suction sampling devices, such as D-vac. The low cost makes the modified blower-vac also accessible to researchers in developing countries.

Assessment of arthropod abundance in crops is critical for investigating population dynamics and species interactions. Here we describe the modification and application of a leaf blower-vac for suction sampling of arthropods in rice.

節足動物のフィールドサンプリングのためのリーフブロワー-VACの変更と応用

Rice fields host a large diversity of arthropods, but investigating their population dynamics and interactions is challenging. Here we describe the ...

Protocol for Microplastics Sampling on the Sea Surface and Sample Analysis

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of scientific publications address microplastic pollution of the sea surface. The protocol below describes the methodology for sampling, sample preparation, separation and chemical identification of microplastic particles. A manta net fixed on an &#187;A frame&#171; attached to the side of the vessel was used for sampling. Microplastic particles caught in the cod end of the net were separated from samples by visual identification and use of stereomicroscopes. Particles were analyzed for their size using an image analysis program and for their chemical structure using ATR-FTIR and micro FTIR spectroscopy. The described protocol is in line with recommendations for microplastics monitoring published by the Marine Strategy Framework Directive (MSFD) Technical Subgroup on Marine Litter. This written protocol with video guide will support the work of researchers that deal with microplastics monitoring all over the world.

The protocol below describes the methodology for: microplastics sampling on the sea surface, separation of microplastic and chemical identification of particles. This protocol is in line with the recommendations for microplastics monitoring published by the MSFD Technical Subgroup on Marine Litter.

海面とサンプル分析にMicroplasticsサンプリングのためのプロトコル

Microplastic pollution in the marine environment is a scientific topic that has received increasing attention over the last decade. The majority of ...

Extraction of Organochlorine Pesticides from Plastic Pellets and Plastic Type Analysis

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment during manufacturing and transport. Because of their environmental persistence, they are widely distributed in the oceans and on beaches all over the world. They can act as a vector of potentially toxic organic compounds (e.g., polychlorinated biphenyls) and might consequently negatively affect marine organisms. Their possible impacts along the food chain are not yet well understood. In order to assess the hazards associated with the occurrence of plastic pellets in the marine environment, it is necessary to develop methodologies that allow for rapid determination of associated organic contaminant levels. The present protocol describes the different steps required for sampling resin pellets, analyzing adsorbed organochlorine pesticides (OCPs) and identifying the plastic type. The focus is on the extraction of OCPs from plastic pellets by means of a pressurized fluid extractor (PFE) and on the polymer chemical analysis applying Fourier Transform-InfraRed (FT-IR) spectroscopy. The developed methodology focuses on 11 OCPs and related compounds, including dichlorodiphenyltrichloroethane (DDT) and its two main metabolites, lindane and two production isomers, as well as the two biologically active isomers of technical endosulfan. This protocol constitutes a simple and rapid alternative to existing methodology for evaluating the concentration of organic contaminants adsorbed on plastic pieces.

Microplastics act as vector of potentially toxic organic contaminants with unpredictable effects. This protocol describes an alternative methodology for assessing the levels of organochlorine pesticides adsorbed on plastic pellets and identifying the polymer chemical structure. The focus is on pressurized fluid extraction and attenuated total reflectance Fourier transform infrared spectroscopy.

プラスチックペレットからの有機塩素系農薬の抽出と塑性解析

Plastic resin pellets, categorized as microplastics (&#8804;5 mm in diameter), are small granules that can be unintentionally released to the environment ...

Determination of Inorganic Arsenic in a Wide Range of Food Matrices using Hydride Generation - Atomic Absorption Spectrometry.

The European Food Safety Authority (EFSA) underlined in its Scientific Opinion on Arsenic in Food that in order to support a sound exposure assessment to inorganic arsenic through diet, information about distribution of arsenic species in various food types must be generated. A method, previously validated in a collaborative trial, has been applied to determine inorganic arsenic in a wide variety of food matrices, covering grains, mushrooms and food of marine origin (31 samples in total). The method is based on detection by flow injection-hydride generation-atomic absorption spectrometry of the iAs selectively extracted into chloroform after digestion of the proteins with concentrated HCl. The method is characterized by a limit of quantification of 10 &#181;g/kg dry weight, which allowed quantification of inorganic arsenic in a large amount of food matrices. Information is provided about performance scores given to results obtained with this method and which were reported by different laboratories in several proficiency tests. The percentage of satisfactory results obtained with the discussed method is higher than that of the results obtained with other analytical approaches.

The usefulness of an analytical method to determine inorganic arsenic in a wide range of food matrices is demonstrated. The method consists of selective extraction of inorganic arsenic into chloroform with a final determination by hydride generation-atomic absorption spectrometry.

無機ヒ素で、広い範囲の食品行列水素化物発生 - 原子吸光法を使用して分析します。

The European Food Safety Authority (EFSA) underlined in its Scientific Opinion on Arsenic in Food that in order to support a sound exposure assessment to ...

Experimental Column Setup for Studying Anaerobic Biogeochemical Interactions Between Iron (Oxy)Hydroxides, Trace Elements, and Bacteria

Fate and speciation of trace elements (TEs), such as arsenic (As) and mercury (Hg), in aquifers are closely related to physio-chemical conditions, such as redox potential (Eh) and pH, but also to microbial activities that can play a direct or indirect role on speciation and/or mobility. Indeed, some bacteria can directly oxidize As(III) to As(V) or reduce As(V) to As(III). Likewise, bacteria are strongly involved in Hg cycling, either through its methylation, forming the neurotoxin monomethyl mercury, or through its reduction to elemental Hg&#176;. The fates of both As and Hg are also strongly linked to soil or aquifer composition; indeed, As and Hg can bind to organic compounds or (oxy)hydroxides, which will influence their mobility. In turn, bacterial activities such as iron (oxy)hydroxide reduction or organic matter mineralization can indirectly influence As and Hg sequestration. The presence of sulfate/sulfide can also strongly impact these particular elements through the formation of complexes such as thio-arsenates with As or metacinnabar with Hg.
Consequently, many important questions have been raised on the fate and speciation of As and Hg in the environment and how to limit their toxicity. However, due to their reactivity towards aquifer components, it is difficult to clearly dissociate the biogeochemical processes that occur and their different impacts on the fate of these TE.
To do so, we developed an original, experimental, column setup that mimics an aquifer with As- or Hg-iron-oxide rich areas versus iron depleted areas, enabling a better understanding of TE biogeochemistry in anoxic conditions. The following protocol gives step by step instructions for the column set-up either for As or Hg, as well as an example with As under iron and sulfate reducing conditions.

Experimental Column Setup for Studying Anaerobic Biogeochemical Interactions Between Iron OxyHydroxides, Trace Elements, and Bacteria

Fate and speciation of arsenic and mercury in aquifers are closely related to physio-chemical conditions and microbial activity. Here, we present an original experimental column setup that mimics an aquifer and enables a better understanding of trace element biogeochemistry in anoxic conditions. Two examples are presented, combining geochemical and microbiological approaches.

(オキシ) 鉄の水酸化物、微量元素、細菌の嫌気性生物地球化学的相互作用を研究するため実験的列の設定

Fate and speciation of trace elements (TEs), such as arsenic (As) and mercury (Hg), in aquifers are closely related to physio-chemical conditions, such as ...

An Ultra-clean Multilayer Apparatus for Collecting Size Fractionated Marine Plankton and Suspended Particles

The distributions of many trace elements in the ocean are strongly associated with the growth, death, and re-mineralization of marine plankton and those of suspended/sinking particles. Here, we present an all plastic (Polypropylene and Polycarbonate), multi-layer filtration system for collection of suspended particulate matter (SPM) at sea. This ultra-clean sampling device has been designed and developed specifically for trace element studies. Meticulous selection of all non-metallic materials and utilization of an in-line flow-through procedure minimizes any possible metal contamination during sampling. This system has been successfully tested and tweaked for determining trace metals (e.g., Fe, Al, Mn, Cd, Cu, Ni) on particles of varying size in coastal and open ocean waters. Results from the South China Sea at the South East Asia Time-Series (SEATS) station indicate that diurnal variations and spatial distribution of plankton in the euphotic zone can be easily resolved and recognized. Chemical analysis of size-fractionated particles in surface waters of the Taiwan Strait suggests that the larger particles (&#62;153 &#181;m) were mostly biologically derived, while the smaller particles (10 - 63 &#181;m) were mostly composed of inorganic matter. Apart from Cd, the concentrations of metals (Fe, Al, Mn, Cu, Ni) decreased with increasing size.

Plankton and suspended particles play a major role in the biogeochemical cycles in the ocean. Here, we provide an ultra-clean, low stress method for the collection of various sizes of particles and plankton at sea with the capability of handling large volumes of seawater.

サイズ分画した海洋プランクトンや懸濁粒子を収集するための超きれいな積層装置

The distributions of many trace elements in the ocean are strongly associated with the growth, death, and re-mineralization of marine plankton and those ...