Cluster for Molecular Chemistry

As will be explained in more detail in 3.1. Preparing an experiment and 3.2. Health risks, these safety regulations revolve around reducing risk when working with hazardous substances by using the Personal Protection Equipment described in 2.3 . If during preparations for an experiment it becomes evident that one or more of the substances being used is highly toxic, the person conducting the experiment should consider whether replacing the substance with a less toxic alternative would affect the results. Alternatively, consider using smaller amounts of the substance. Whatever the circumstances, all experiments involving chemicals must be carried out in a fume cupboard, all solutions, reagents and products must be clearly and correctly labelled, and proper PPE should be worn at all times.

• Eating and drinking are prohibited in the lab areas, with the exception of the designated areas (no chewing gum and no sucking/chewing pencils etc). Smoking is prohibited throughout the building.
• Do not put food or drink in a fridge or any other area or container intended for chemicals.
• Never drink from glassware used in the lab.
• Always wash your hands before eating or drinking and before and after using the toilet, but also during your work whenever necessary.
• Never wipe your hands on your lab coat; change your coat for a clean one regularly.
• Sucking liquid into a pipette is prohibited, unless it is clearly stated that the liquid being drawn up is entirely safe; use a pipette balloon (do not let the liquid touch the balloon), a micropipette or a dispenser.

• Use high-quality gloves whenever necessary (Appendix 1).
• Do not wash your hands with organic solvents (they remove the natural oils from your skin).
• Never work with an open wound; always cover it with a band aid that will not come off easily.
• Ensure good personal hygiene, i.e. clean hands and short nails. - Wash your hands when leaving the lab (always before eating and drinking) and during your work if necessary.

• Wear safety glasses

• Always use a fume cupboard when working with chemicals and keep it closed as much as possible. Never work in a fume cupboard which is out of order and/or gives an alarm.
• Close bottles of chemicals immediately after use.
• Never smell substances directly from bottles; waft a little vapour or smell the stopper carefully.
• Avoid dispersing particles.

• Hair: Long hair must be tied back and no headwear may be worn. This is to avoid contamination and fire risk.
• Transporting chemicals: Use special trays with handles for transporting bottles of chemicals from one lab to another.
• Needles: avoid walking around with noncapped injection needles. Discard needles properly (in a "SharpSafe", see section 3.5) immediately after use. If this is no option, cap them (carefully!)

The main message of the detailed safety regulations shown above is that good personal hygiene and meticulous, careful, orderly and tidy working practices will help to prevent accidents. This is not only the responsibility of the person carrying out the work, but also of the people working around him/her. Many accidents are not caused by the victim, but by someone else working in the same area.

A large proportion of accidents in a laboratory are caused by falling, stumbling, or bumping into things. Order and tidiness in the lab are vital. Never work on the edge of the bench or fume cupboard. Place heat sources (burners, heating mantles and hot plates) where they are least dangerous. All laboratory workers must conduct experiments in a way that will keep the fire risk to a minimum. When working with flammable substances, welding or other activities with a high fire risk, extinguishers should always be within easy reach. Make sure that benches and fume cupboards are clean. Clear up spillage immediately (including water). Label all spray bottles. Wash and tidy up equipment after use. Make sure that the lab is clean and tidy at the end of every working day. Plan weekly and six-monthly cleaning sessions together with other workers in the laboratory.

• fire and smoke protection doors are not propped open
• corridors, emergency exits and escape routes are kept clear of obstacles
• fire extinguishers and alarm buttons are always visible and accessible, and can be used without delay if necessary
• no rubbish, packaging material or paper is left lying around
• power connections are sound, electric wires are properly fused, and approved adapters are used
• equipment is switched off when not in use
• no household appliances (coffee machines, toasters etc.) are used in the rooms or laboratories
• defects and irregularities are reported immediately
• no superfluous chemicals or cleaning agents are left in the lab areas.

Before embarking on experiments, new laboratory workers must feel confident in the laboratory and be made aware of a few important aspects.

• exits and emergency escape routes
• the first-aid boxes, and the qualified first-aiders (list available for every department). There is at least one first-aid box per department (and per lab in some departments); ask the office staff or the first-aiders from the department and/or Internal Emergency Team if necessary.
• Fire extinguishers and hose reels.
  Find out where they are (next to fume cupboards and in the lab areas and corridors) and how to use them. Read the instructions on the fire extinguisher.
  • CO₂ extinguisher: can be used in case of most types of fire, but will be empty within ~20 seconds! Never use on persons, as liquid CO₂ is so cold that it causes burn-wounds. Never use on metal fires because carbon dioxide reacts with for example LiAlH₄ and elementary magnesium, sodium, potassium and aluminium.
  • Hose reel: general extinguishing equipment; never use in case of:
    
    • Burning organic solvents, especially oil, as they will float on the water and the fire will spread quickly.
    • Metal fires, as water reacts with for example LiAlH₄ and elementary magnesium, sodium, potassium and aluminium.
    • Fires involving GMO’s, because the GMO’s will be spread.
    • Persons, as the hose is really strong.
    • Electrical equipment (unless power is absolutely off), because water causes short circuits.
  • Sand: preferred to extinguish metal fires. Can also be used for organic solvents
  Laboratory workers can familiarise themselves with the various extinguishing devices during the periodical drills arranged by the Department of Occupational Health & Safety and Environmental Service (see paragraph 2.2).
• Emergency showers
  Find out where they are and how they work.
  Be careful:
  1. they are connected to the mains and are under high pressure;
  2. the area can soon flood if they are used so try not to leave them on for too long.
  Emergency showers can be better than fire blankets as they not only extinguish the fire, but they also have a cooling effect on the victim.
• Fire blankets
  Find out where they are and how to use them. Use emergency showers in preference to fire blankets.
  IMPORTANT: a person needs to be laid down on the floor and rolled into the blanket. When the victim is standing upright, the fire will not be extinguished
• The location of the nearest green telephone
  These telephones continue to work even if the main switchboard is out of order; only to be used to report emergencies if the other telephones are down.
• The emergency and alarm buttons. There are three sorts:
1. Large round buttons near the emergency exits of all lab areas;
   pressing the button cuts off the electricity in the wall sockets in the laboratory concerned (or several labs in a wing).
2. Square red boxes with a glass front, which can be activated by breaking the glass (see figure 4 in paragraph 4.5 ). These are fire alarms; they set off an alarm.
3. Square green boxes with a glass front, which can be activated by breaking the glass. They release the lock on the emergency exit if this does not happen automatically as part of an evacuation alarm.

Working alone means that a person is working in a situation whereby he/she cannot continually be seen or heard by others. Situations like this can occur during regular working hours; laboratory workers should try to anticipate periods in which most of the group is attending a conference, for example. The guidelines for working alone are as follows:

• Working alone in a laboratory in the ‘medium’ (SL-2) or ‘low’ (SL-1) category is strictly prohibited. Another member of staff must be in sight or within hearing distance of the workplace/fume cupboard at all times.
  When working outside regular working hours, the overtime clock (see Figure 1) must be activated in order to restore the ventilation in the lab. Without proper exhaustion it is not safe to work in the lab.
• Laboratory workers may work alone in one of the offices on library work, computer work or reading and writing.

Laboratory, practical or workshop activities are not allowed outside regular working hours unless explicit permission has been given by the manager concerned; working alone is still prohibited, even if permission for working outside regular working hours has been granted. The group leader is authorised to make decisions in these matters and is therefore ultimately responsible.

2.7. End of the working day, night-time experiments

1. Close the fume cupboard at the end of the day and switch off the lights.
   IMPORTANT: fume cupboards must be properly closed as they only work at 30% of their capacity at night!
2. Switch off all equipment at the end of the day.
3. Take special care that all water cooling is turned off if it’s not needed for an experiment. In case you do need water cooling, please take care that all your water hoses are properly secured (only use transparent/plastic ones!) and that the water flow is not too high. Be aware that water pressure might increase significantly during the evening and at night.

The Lab Steward (see 2.2) is responsible for experiments that continue into the night; regulations may vary per department. For each overnight experiment a card that shows information about the specific risks of the experiment must be placed at the fumehood window. This card can be red, green or yellow, depending on the risk level of the experiment. The general rules regarding the cards are as follows:
Green card: for reactions that do not require heating/reflux, pressure, vacuum or hydrogen
Red card: for reactions that require heating/reflux, pressure, vacuum, or hydrogen; also an overnight running vacuum pump requires a red card.
Yellow card: for continuously operating setups (drying liquids under nitrogen or argon.

Also the filling in of the cards should happen according to strict rules:

• Name, phone number, location
• In case of green or red card: date of the experiment (from [day] to [day])
• Type of experiment
• Volume of the reaction solvent
• Write down all solvents and chemicals that you use in the reaction, including their hazards, on the card in full: that means that you must not use abbreviations like THF, KOH, DCC, Pd/C, etc.

IMPORTANT: laboratory workers carrying out night-time experiments are personally responsible for taking the card back to the fume cupboard with the experiment the next morning. The Safety Steward can withdraw the card (or cards) if the laboratory worker misuses them.

Before every experiment, examine the specific risks of the chemicals you are using by looking at the label, consulting Chemwatch⁵ and reading the supplier’s information. Do not simply take note of the risks if everything goes according to plan, but look at what can happen if something goes wrong. Think about the possible repercussions, or what would happen if the agitator, reaction flask, cooling or heating were to break or fail. Precautions must be taken for these risks.
The safety regulations shown in the previous section (2. General safety) are aimed at limiting all the risks involved in working with hazardous substances. The following paragraph (3.2. Health risks) shows exactly why everything must be done to minimise the risk of a hazardous substance being ingested, absorbed via the skin, penetrating the eyes or being inhaled. If the risk inventory indicates that one or more of the substances being used in an experiment is highly toxic, the laboratory worker should consider whether less-toxic chemicals could be used without affecting the results. Another useful measure is to keep the amount of the substance used to a minimum. Whatever else, all experiments involving chemicals must be carried out in a fume cupboard, all solutions, reagents and products must be correctly labelled, and good personal protection equipment must be worn.

⁵For links, see paragraph 5.2 at the end of this document.

3.2 Health risks

The GHS-CLP (Globally Harmonised System of Classification, Labelling and Packaging of Chemicals) system⁵ introduced throughout Europe in 2009 distinguishes between the following hazard risks and categories of hazardous substances and mixtures, using the pictograms shown below:

• Physical hazards (flammable, corrosive, explosive, compressed gases, oxidising);
• Health hazards (corrosive, toxic, irritating/sensitising, long-term health risks, oxidising);
• Environmental hazards (hazard to aquatic environment).
  

Any particular substance may belong to several risk classes and categories.

Figure 2: GHS categories for hazardous substances and the relevant pictograms.

A GHS label comprises the following elements:

• one or more pictograms or hazard symbols (see Figure 2), showing the hazard category;
• a signal word (‘danger’ or ‘warning’), depending on the hazard category of the substance;
• one or more hazard statements (‘H statements’), which replace the present ‘R statements’ (risk statements) and describe the risks posed by the substance;
• one or more precautionary statements (‘P statements’), which replace the present ‘S statements’ (safety statements), and explain the safety regulations governing working with the substance concerned. There are five types of safety recommendations: general statements, and statements regarding prevention, response, storage and disposal. Pictograms can also be used to indicate the recommendations.

Home-made mixtures and solutions of chemicals should be labelled according CLP (Classification, Labelling and Packaging) regulation. The bottle or container for general use should contain:

• Name and ratios of the chemicals inside the bottle or container
• Hazard pictograms
• The relevant signal word

For Personal use also:

• Your name
• Date

If for solutions no GHS symbols are applicable, a blank GHS sticker should be used as a proof of checking its properties. If GHS classification of newly synthesized compounds is unknown, no stickers should be used. Stickers with GHS symbols as well as blank stickers are available from Peter van Dijk.

Posters showing GHS symbols and the H and P statements are available from the Department of Occupational Health & Safety and Environmental Service. The measures per hazard category shown below correspond with those in section 2.3. Personal protection equipment and 2.4. Order, tidiness and hygiene.

The hazard categories are:
Flammable:
Solids, aerosols, gases and liquids (such as organic solvents with a high vapour pressure, e.g. diethyl ether, ethanol) are flammable.
Measure: Avoid open flames in a lab where these substances are used, and beware of heat sources, sparks, electric discharge etc. that may cause ignition.

Corrosive:
Corrosive (aggressive) substances (such as concentrated acids, concentrated alkali, strong oxidisers) are dangerous if they come into direct contact with the skin, eyes and mucous membranes.
Measure: Avoid contact by using PPE.

Toxic:
Toxicity: It is best to assume that all chemicals are potentially dangerous to health, even though for some of them (e.g. water) the concentration would have to be very high to pose a serious risk. In chemistry, the term toxic substances is used if even small doses of a substance could damage the body. Toxicity is therefore relative and depends on the dose. This in turn depends on:

• concentration,
• duration of exposure;
• means of entering the body: ingestion, via the skin, via the eyes, inhalation.

The effect of the dose also depends on personal factors such as sensitivity and general condition.
Measure: Avoid contact by using PPE; do not eat or drink in the labs and wash your hands when leaving the lab.

Irritating, Sensitising, Harmful:
Irritating substances are mildly toxic and single exposure causes minor irritation to the skin, eyes or airways without lasting damage. In the case of repeated exposure without time to recover, the cumulative effect can cause a chronic inflammation. Examples: high or low pH, dissolving skin oil due to organic solvents, specific reactions to epoxide resins, for example, with protein in the skin or mucous membranes. Sensitising substances are allergens that cause an allergic reaction by activating the immune system. This can lead to inflammation and allergic conditions. Alongside reactions to biological-based substances, an allergic reaction can also be caused by latex, for example. Take care when using gloves (see Appendix 1).
Measure: Avoid contact by using PPE; do not eat or drink in the labs.

Long-term health risk:
This includes all the CRM substances: carcinogenic (causes cancer), reprotoxic (dangerous to unborn foetus) and mutagenic (causes permanent changes to genetic material).
Measure: Avoid contact by using PPE; do not eat or drink in the labs. If substances in this category are ordered in Labservant, a warning is generated by Chemwatch stating that this is a CRM substance; in extreme cases, certain orders will initially be blocked. The Internal & Housing Affairs department and the head of the department concerned will first look for an alternative to this substance; if this is not available, a permit for working with the substance will have to be applied for via the Department of Occupational Health & Safety and Environmental Service.
IMPORTANT: Pregnant women must avoid all contact with reprotoxic substances. Tell your manager in plenty of time if you are pregnant. Strictly speaking, all laboratory workers (m/f) planning a family should stay away from reprotoxic substances.
Compounds that damage the airways when inhaled (such as asbestos and silica) form another important category and can lead to silicosis (Potter’s rot). Asbestos should only be processed by specialist companies; chromatography columns should be filled in a fume cupboard with good exhaustion (see 3.6).
Measure: See the PPE for breathing protection (dust and gas masks) in the faculty safety regulations⁶.

Hazard to aquatic environment:
Toxic to water organisms.
Measure: Avoid releasing these substances into the environment by for example discharging them through the drains (see also Appendix 3).

Explosive:
Substances and mixtures that may undergo a powerful exothermal discharge without contact with oxygen (air), with a rise in pressure resulting from the accumulation of gases.
Measure: Avoid heat, shocks and high concentrations. Use the fume cupboard when working with substances of this kind.

Oxidising:
Helps to ignite flammable substances.
Measure: Avoid contact with flammable substances.

Compressed gases: see section 3.6

⁶For links, see paragraph 5.2 at the end of this document.

3.3. Threshold limits

Threshold limits have been set for hazardous substances:
The limit of the exposure concentration is linked to a specific duration. The combination of threshold limit and duration should be chosen so that a laboratory worker would not suffer damage to his/her health even after having worked with the defined threshold limit of the substance concerned for forty years. For this reason, the threshold value is defined as a Time Weighted Average over a period of eight hours, the length of an average working day; this is the so-called 8-hour TWA. The time-weighted average for acutely toxic substances is defined as the 15-min TWA. Specific cases require an instantaneous exposure threshold limit, the so-called ceiling value (reference time practically zero). The laboratory worker is at liberty to determine the reference time for the threshold limit, as long as the basic principle is realised: exposure at the threshold limit during an employee’s entire working life must not cause damage to his/her health; this also applies to his/her children (see table 2 for the example of acetone).

Threshold limits can be found in Material Safety Data Sheets. However, the extent of exposure in the lab is hard to estimate. Therefore, always work in your fume cupboard to prevent inhalation.

Table 2. Threshold limits for occupational exposure to acetone.

Legal threshold limit (Netherlands)
Acetone
Threshold limit 8-hr TWA	1210 mg/m3
Threshold limit 15-min TWA	2420 mg/m3
EU (directive 2000/39/EC)
Acetone
Indicative threshold limit 8 hr	500 ppm
	1210 mg/m3

3.4. Storage of chemicals

Chemicals are separated according to their danger categories. In case of multiple categories for a single compound, the priority is:

1. Flammable
2. Oxidizing
3. Toxic
4. Corrosive
5. Dangerous for the aquatic environment

Dangerous combinations can be found in Figure 3. The fifth category is normally combined with the other categories, so no extra precautions are needed.
Don’t store Flammable and Oxidizing compounds together at any time.

Figure 3. Dangerous and "safe" combinations of the different GHS categories.

Solvents: All solvents have to be stored in the safety cupboards ("plofkasten"), in different drip trays according to their safety label. This also applies to the solvent stock. Each drip tray should contain chemicals of only one danger category; its volume should be at least 110% of that of the largest bottle.
Liquid commercial chemicals should also be stored in these safety cupboards. Each tray is assigned to a single danger category and its volume should be at least 110% of that of the largest bottle. Acids and bases: have to be stored in separate cupboards under the fumehoods (this also applies to acids such as SOCl₂). However, if they are flammable (such as KOH in EtOH) bases and acids should be stored in the safety cupboard. This does not apply to HCl solutions as they are corrosive; its ethylacetate and dioxane solutions go in the waste fume cupboard. Nitric acid (oxidizing) should be stored completely separate from all other liquids.
Solid commercial chemicals: Personal chemicals (only solids, registered in your name) can be stored under your own fume hood, as long as they are categorized as mentioned above in different drip trays.
Shared solid chemicals, such as salts, bases and silica, can be stored per category in the apothecary cupboard.
Toxic chemicals should not be stored in the lab but in the logistic centre. If they are used in the lab they should be returned to the logistic centre as soon as possible.
Oxidizing chemicals, however, have to be in a separate drip tray in the safety cupboard.
Freezer/Fridge: Commercial and "home-made" chemicals should not be in the same freezer/fridge. All the different categories have to be in different drip trays.

Liquids of contents 250 ml or less and solids of contents 1 kg or less hasn’t to full fill these rules. However place them always in a drip tray.

During ordering of a chemical a location has to be given were it will be stored. This location should be in agreement with the above rules. After use of a chemical it should be returned to its location that is given on the label that can be found on the container.

"Home-made" chemicals: Home-made mixtures and solutions of chemicals should be labelled according CLP (Classification, Labelling and Packaging) regulation. The bottle or container for general use should contain:

• Name and ratios of the chemicals inside the bottle or container
• Hazard pictograms
• The relevant signal word

• Your name
• Date

For waste originating from our laboratories the following main categories are distinguished: liquid waste, solid waste, glass waste, sharp objects. For a full overview, please refer to the faculty regulations mentioned in the introduction (also available in English)⁷ and see the AMD website⁷ on "Afvalstoffenregeling FNWI"/"Waste Management Regulations" and "Overzicht van de indeling voor gevaarlijke stoffen RU"⁷.
On the whole, paper/cardboard that has not been in contact with chemicals, and the majority of special waste products such a ink cartridges, batteries and adhesives are collected per department in special containers in the technical area near to the lifts.

Labelling: All waste containers must be properly labelled; appropriate stickers are available in every lab area. See faculty waste disposal regulations) for an overview. IMPORTANT: the old labels must be removed or rendered illegible on all empty containers ready for disposal before being taken to the Logistic Transfer area; this also applies to glass bottles and jars in the glass waste.

Liquid waste should be divided into different categories according to the flow chart in Appendix 2 and disposed in properly labelled 5-litre or 10-litre jerry-cans. The main categories relevant for our labs are: inorganic acids (10-litre jerry-can), inorganic lyes (10-litre jerrycan), non-halogenated organic solvents, halogenated organic solvents, ethidium bromide solutions, toxic liquids, watery solutions originating from laboratories.

Solid heavy metals, to prevent ignition, are collected in liquid waste containers according to the liquid waste flow chart (see Appendix 2).

Solid waste, including silica from the chromatography columns, soiled paper and glass contaminated with chemicals, but with the exception of heavy metals, needles and special waste products (see above) such as ink cartridges, batteries and adhesives, are collected in a brown container with a beige lid. The appropriate labels should be attached to the container (label 5.12 and ADR label toxic).

Glass when contaminated with chemicals, can be collected separately in a similar brown container with beige lid or in a solid waste container (See above). The appropriate labels should be attached to the container (label 5.12 and ADR label toxic). Clean glass is collected in a glass waste container in the laboratory. Rinse glass first if it has been contaminated with chemicals, or let solvents evaporate in the fume cupboard. IMPORTANT: remove the original labels or render them illegible.

Needles and other sharp metal objects must be collected in a special waste container, a so-called "SharpSafe" (yellow with a white lid and a narrow opening – make sure they are properly attached). Preferably, this container is only used for needles without the covers, as putting the cover back on the needle is an additional risk and the covers take up extra space. To detach a needle from a syringe in a safe manner, the incisions in the SharpSafe lid can be used. A SharpSafe should NOT be used for glass; neither for syringes, tubing or any other non-sharp thing you might attach to a needle.

Waste is removed from the transfer rooms in the laboratory wings by the Logistic Centre, but only if their instructions as given here are strictly followed up:

• Do not fill the waste disposal jerrycan to more than 80 % of its volume, and make sure that it is not contaminated on the outside.
• Attach an adhesive label describing the contents: halogen-rich, halogen-poor, etc.
• In addition, the appropriate ADR-sticker with the danger symbol, corresponding to the symbol in the bottom left corner of the ‘content‘ label, should be attached. This is supplied by the Logistic Centre with the ‘content‘ label; if necessary additional stickers can be ordered from them (mail to: logistiekcentrum@science.ru.nl).
• Close the lid of a SharpSafe properly (which is: irreversibly) before bringing it to the transfer room.
• For an empty metal container to be taken away as waste, remove any residual liquid, take off the lid and erase the labels including all the barcodes, and in particular the danger symbols, so that it can no longer be mistaken for chemical waste.
• Empty plastic 5 L jerrycans can be re-used as waste vessels and will not be taken away by the Logistic Centre when empty. They can be stored on the lab only if they are clean and dry and the labels/danger signs are removed or erased, in particular the danger symbols, so they can not regarded as full. If an excess of empty jerrycans is piling up in the lab, they can alternatively be discarded in a special "plastic waste container" in the container room (HG03.004). Make sure that the jerrycans are clean and dry and that the labels/danger signs are removed.
• Chemicals without bar code will not be taken away.

Pipes for hydrogen (10 bar), argon, nitrogen, carbon dioxide and helium have been fitted on the lab benches and/or in fume cupboards in line with expected use in the lab areas. If another gas (or hydrogen> 10 bar) is needed regularly for long periods, a large cylinder can be placed in one of the cylinder cupboards outside the lab, and connected using the interchangeable gas feed. Large gas cylinders of this type are available from the Logistics Centre or must be ordered.
IMPORTANT: the gas cylinder must be secured in an upright position. If a cylinder falls (for whatever reason) and the reduction valve is damaged, the cylinder can be launched like a torpedo. Be aware that if large volumes of nitrogen or helium (or any other relatively ‘safe’ gas) suddenly escape from a damaged cylinder or container in a closed, badly ventilated room, this can lead to an oxygen shortage and ultimately to suffocation.
If at all possible, use small cylinders (the so-called) ‘lecture bottles’ in the fume cupboard. Keep these small cylinders in the fume cupboard until they can be removed via the Logistic Transfer area.
Before using either large or small cylinders, check that the inspection date has not elapsed. Listen carefully to your mentor’s instructions about using the reduction valve.
The Matheson Gas Data Book (Library: Geert Grooteplein 15 Application number: MB 56 b 10) is useful reading if you want to know more about the properties of various gases.

Hazardous gases we define as gases or vapors that can cause serious injuries or even death upon inhalation and/or direct exposure to skin. A list of highly toxic gases can be found in paragraph 5.2. When you are planning to perform an experiment involving the use of a hazardous gas, comply with the following rules:

• Avoid the use of hazardous gases when possible. Only use hazardous gases when there is no alternative.
• Look up the MSDS data of the hazardous gas and take the possible precautions one should.
• Inform the co-workers in your lab about the experiment and the possible dangers of the hazardous gas.
• Keep the fume hood sash closed as much as possible during the experiment.
• In case of an accident (sudden outburst of hazardous gas), alert your coworkers and evacuate the lab immediately. Cal 55555 and explain the situation ( see section 4.2: guidelines for accidents). In any case, do not go back into the lab to attend to any possibly injured people!
• After completion of the reaction, follow the right quenching procedure (Hazardous Laboratory Chemicals Disposal Guide) to destroy any excess of hazardous gas.

Hydrogen fluoride (HF, g) is very hazardous to handle because it is highly corrosive and can cause deep tissue damage and systematic toxicity after exposure. Therefore an indicator card should be present on the fume hood where the experiment is performed and at all the lab entrances. Also be sure to warn everybody present on the lab at the start of the experiment.

If there is a need to work with HF (g) always contact Dennis Löwik (office: 03.016) for supervision and accompaniment during all of the experiments performed.

Cyanide is a very toxic substance that is present in salts (eg NaCN), in acid (HCN, which in gas phase smells like almonds) and as reagents (eg cyanogen bromide, CNBr). Hydrogen cyanide is lethal at a concentration of 270 ppm and a concentration of 180 ppm is life threatening within several minutes.

• Place a HCN detector in the fume hood
• Place a sign indicating working with HCN on the fume hood
• Inform the people in the surroundings around you

If an accident with cyanides occurs be sure to:

• Call in the qualified first-aiders
• Call 55555 and report the accident and the number of victims
• Make sure that a BHV-er evacuates the victim out of the area. Do not do this yourself!

Vessels of liquid nitrogen can be ordered from the Logistics Centre and will be delivered to the department via special transport. IMPORTANT: Be aware that liquid nitrogen can escape from a vessel very quickly (if a Dewar container is damaged and loses its cooling capacity). In a closed room, this can cause a lack of oxygen and ultimately suffocation. The laboratories where cryogenic liquids are used must therefore be adequately ventilated. Smaller rooms can be fitted with an oxygen sensor to warn if the oxygen level drops.

Another potential danger when using liquid nitrogen is the fact that oxygen from the air can condense into it. This should be taken into account when handling a cold trap. A cold trap is commonly used to condense solvents, thereby preventing harmful gasses from entering the attached pump (Figure 4). It is important to know how to install and more importantly, remove the cold traps, as formation of liquid oxygen (bp -183°C) and explosion thereof can cause serious injury.

Installation:

Make sure to NOT attach the vacuum pump to the top adapter as this will cause the vacuum pump to suck up condensed solvents when it reaches the level of the internal tube.
Make sure the installation is airtight (close the system).
Switch on the vacuum pump.
Immediately place/fill the dewar flask with liquid nitrogen.

Removal:

Remove the dewar flask with liquid nitrogen.
Allow air into the system and switch off the pump.
The cold trap is now safe to be disconnected with appropriate gloves to protect you from the cold.

DANGER: when air is allowed into the cold trap whilst still being cooled by liquid nitrogen, oxygen will condense inside the trap to form extremely dangerous mixtures of organic material and liquid oxygen which can potentially explode.

Figure 4: Cold trap.

Chromatography using silica in glass columns should be carried out in a fume cupboard. Silica is made up of minute particles which can penetrate the lungs causing silicosis (occupational lung disease) (see also hazard category Irritating/Sensitising/Damaging in paragraph 3.2); columns should be filled and emptied in the fume cupboard. Inspect the glass for cracks before it comes under pressure, and protect yourself from glass splinters by packing the column in a net. Use a pressure valve if the column is to be pressurised.

Listen carefully to your mentor‘s instructions every time you distil a new solvent; check that the correct drying agent has been used, and whether the solvent needs to be pre-dried. Make sure that the cooling is switched on, never turn off the nitrogen taps, and let the solvent cool to room temperature before refilling. If in doubt, ask your mentor. All distillation set-ups must be constructed so that the heating mantle can be taken out from under the set-up in an emergency by removing the lab jack.

Cleaning glass filters with a mixture of sulfuric acid and hydrogen peroxide can cause an explosion if the filter is rinsed with an organic solvent such as acetone and then not properly dried. This makes it a very risky procedure, which should only be used if it is the only possible way to clean the glass filter. Even then, it should only be applied by experienced staff.

3.11.1 Procedure for cleaning glass filters with sulfuric acid and hydrogen peroxide
The last few years a number of incidents occurred during cleaning glass filters with a mixture of sulfuric acid and hydrogen peroxide. To avoid such incidents use the following procedure.

Use this procedure only if the glass filter can not be cleaned by any other method and if excess waste is removed. Make sure that all the glassware used in this procedure is free of acetone or any organic solvent. Only a very small amount of acetone is enough to cause a big explosion.

Procedure:

• Place the filter on a filtering flask.
• Rinse the filter with (a lot of/excess) water.
• Place the filter in a beaker filled with a layer of water.
• Make sure the beaker is absolutely free of acetone (rinse it with water if you’re not completely sure)
• Fill the filter with sulfuric acid and a small amount of hydrogen peroxide.
• If the filter is clean dispose the content of the filter in the sink and rinse with water.
• Place the filter on a filtering flask.
• Rinse the filter with water.

Materials that need to be sterilised before use should be sterilised in a special pressure cooker or, if large amounts need sterilising, in an autoclave. All materials originating from safe microbiological procedures must also be sterilised after use in line with the strict regulations governing safe microbiological procedures. Depending on the nature of the materials, they can be sterilised using dry sterilisation (only in the autoclave) or they can be post-sterilised, whereby the temperature and the times are adjusted to correspond with the amount and nature of the materials.
Place all objects to be sterilised in drip-trays or pans. Check glass bottles carefully for tiny cracks before starting. In the case of liquids that are being sterilised in closed bottles check that the screw top has not been screwed on tight, so that the pressure caused by heating can escape. Beware of delayed boiling after the post-sterilisation process. Liquids can easily boil over, particularly if the bottles are full and the autoclave or pressure cooker is opened while the temperature is still high.

Biochemistry and cell biology involve fewer safety risks than organic chemistry, but for some biochemical assays, hazardous chemicals are required and thus it is important that you know how to work with them. Compounds that alter biochemical structures are often toxic or carcinogenic. Moreover, chemicals might be needed for purification or other experimental purposes. Always be aware that biochemical experiments bring possible risks even though performed at SL-1 safety level, and make sure you know how to safely handle the compounds you need.

Ethidium bromide is a DNA stain which binds to DNA through interchelation and can therefore act as powerful mutagen. Always work on the designated bench and make use of gloves when handling ethidium bromide. Only touch contaminated materials with your gloves and discard your gloves immediately after use. Consider wearing only one glove in case you have to handle both contaminated and non-contaminated materials. Ethidium bromide-containing waste needs to be discarded into the solid waste container.

Acrylamide is widely used as a cross linking agent for electrophoresis separation procedures (e.g. SDS-PAGE). Acrylamide is easily absorbed by the skin and acts as a neurotoxin and (possibly) as carcinogen. When handling acrylamide powder, always work in a fume hood. Chances of ingesting or absorbing acrylamide are limited when working with ready-made solutions of the compound (common in most labs). Nonetheless, one should always wear gloves when working with acrylamide and discard your gloves immediately after use. It is recommended to use a dedicated part of the lab for working with acrylamide in order to minimize the risk of cross-contamination. Once FULLY polymerized, acrylamide gels are significantly less toxic. They can be discarded in the solid waste containers.

When using organic solvents for e.g. staining/destaining of protein gels or trizol/phenol purification, handle them in the fume hood as much as possible even when working with low quantities. Also always make use of the proper liquid waste container (halogen rich, i.e. compounds containing F, Cl, I or Br, or halogen poor). Contaminated waste materials should go into the solid waste container. If you are not sure how to handle a certain chemical, consult someone who does.

4. Faults, fire and accidents

• Alarm: First alert all people in your surrounding and then ring 55555 in the event of an accident (fire, electrocution, medical emergency). If the normal telephones are out of order, the green telephone should be used. State your name and department and give a brief account of what has happened. Explain concisely what is going on and more specifically, the type of help you require. Call in the qualified first-aiders where necessary. TIP: If you store 0243655555 in your mobile phone under the name ALARM RU, you will be able to raise the alarm in any situation wherever you are on the campus.
• Faults: In the event of minor problems (faults in one or more fume cupboards, overflows, blown fuses), contact the Department of Property Management (UVB). Ring 33333, state your name, department and explain the problem; it will be resolved as soon as possible. Follow-up your telephone call with an e-mail (33333@science.ru.nl), sending a copy to the PAM-mers (Peter van Dijk, Theo Peters or Helene Amatdjais-Groenen).
• Personal accidents must always be reported to the PAM-mers (Peter van Dijk, Theo Peters or Helene Amatdjais-Groenen) and the Department of Occupational Health &Safety and Environmental Service.
• Accidents resulting in material damage: report immediately to the PAM-mers (Peter van Dijk, Theo Peters or Helene Amatdjais-Groenen), internal & Housing Affairs and the Department of Occupational Health & Safety and Environmental Service.
• The use of fire extinguishers must be reported immediately to the UVB. The UVB will make sure that all used equipment is replaced.
• Dangerous situations: report them to the Lab steward or Safety steward (see the lists in the various departments) or directly to your manager and the PAM-mers (Peter van Dijk, Theo Peters or Helene Amatdjais-Groenen).
• Dangerous activities: make the person concerned aware of what he/she is doing and explain why it is dangerous. Alert the mentor, Safety Steward and the PAM-mers (Peter van Dijk, Theo Peters or Helene Amatdjais-Groenen).
• Minor inconveniences: (noise, dirt, bad smells in the lab areas) contact the PAM-mers (Peter van Dijk, Theo Peters or Helene Amatdjais-Groenen) or internal & Housing Affairs (tel. 52600) or the UVB service desk (tel. 11332).
• Faulty equipment: warn the person responsible for the equipment.

During a power failure there is not enough ventilation inside the labs and surrounding areas to provide a safe working environment. But also after a power failure safety cannot be immediately guaranteed. Everything might look normal inside the labs while certain systems are not running. For example, point ventilation and air-conditioning can be still non-operational, which causes dangerous situations even when the fume hoods are running again.

So in case of a power failure, even if everything seems to be back operational immediately, handle as follows:

• Close any bottles containing chemicals, especially those in fume hoods.
• Turn off equipment that can be harmed or do harm when the power comes back. Pay special attention to equipment cooled with water since the water pressure can fall during a power failure. Turn off heating and/or vacuum while leaving the cooling itself running.
• Close all fume hoods (and cupboards).
• Make sure everyone leaves the lab and office spaces, assemble in the 3rd floor corridor.
• Contact your safety steward or one of the PAM-mers; one of them will call 33333. Together they will decide when the labs can be entered again.

For chemical spills, the ‘Hazardous Laboratory Chemicals Disposal Guide’ can be consulted here: Hazardous Laboratory Chemicals Disposal Guide. A supply of the Bentonite absorber material frequently recommended in this Guide is available on each wing, please make sure of its location before you start chemical work.

There are several classes of chemical spills:

• Minor Chemical Spill
  Defined as: spillage of small quantities of chemicals. Act as follows:
  • Alert people in immediate area of spill.
  • Wear protective equipment, including safety goggles, gloves, and long-sleeve lab coat.
  • Avoid breathing vapors from spill.
  • Confine spill to small area.
  • Use appropriate kit to neutralize and absorb inorganic acids and bases. Collect residue, place in container, and dispose as chemical waste.
  • For other chemicals, use appropriate kit or absorb spill with vermiculite, dry sand, or diatomaceous earth. Collect residue, place in container and dispose as chemical waste.
  • Clean spill area with water.
• Major Chemical Spill
  Defined as: spillage of large quantities of chemicals (i.e. solvents >1 L, solids > 1 kg), or when anyone is injured or contaminated. Act as follows:
  • Attend to injured or contaminated persons and remove them from exposure.
  • Alert people in the laboratory to evacuate.
  • If spilled material is flammable, turn off ignition and heat sources.
  • Call 55555, state place and nature of the spill.
  • Close doors to affected area.
  • Have person knowledgeable of incident and laboratory assist emergency personnel.
• Special spills
  
  • Spills in the eye:
    
    • Immediately rinse eyeball and inner surface of eyelid with water continuously for 15 min, using the eye showers or the eye wash bottles available in every lab.
    • Forcibly hold eye open to ensure effective wash behind eyelids.
    • Obtain medical attention.
    • Report incident to your mentor and to one of the PAM-mers.
  • Mercury spills
    Remember: mercury vapors are odorless, colorless, and tasteless. Because of the health effects of mercury, the extremely difficult and time-consuming procedures required to properly clean spills, every effort should be taken to prevent accidents involving mercury. Always store mercury in unbreakable containers and stored in a well-ventilated area. Wear protective clothing: lab coat, gloves and safety goggles!

• Dam the mercury (using rags or other disposable items) to prevent spreading. Divert the mercury from drains, cracks and crevices.
• Keep persons who are not involved in the clean-up away from spill area to limit exposures and to prevent the spread of contamination.
• If you have come in contact with the mercury, avoid spreading the contamination to other areas. Put contaminated clothing/shoes into a trash bag and wipe off any visible mercury beads into the bag. After cleaning up shampoo and shower yourself.
• Use a chemical to coat the mercury or form an amalgam with the mercury, this will keep the mercury from vaporizing and being released into the air. Use sulfur powder for this. As an alternative a mercury collector can be used. Ask one of the PAM-mers.
• Report incident to mentor and to one off the PAM-mers.

The guidelines on how to deal with an accident are shown below:

• Alert your surroundings
• If external assistance is needed ring 55555 otherwise call for a first aider or PAM-mer.
• Once you have called the alarm number, the Rapid Intervention Team/Internal Emergency Team will arrive within 4 minutes.
• Follow instructions given by the Internal Emergency Team.
• Report the accident to the Department of Occupational Health & Safety and Environmental Service so that an internal investigation into preventive measures can be carried out if necessary (see Section 5.2 for the link to the report form).

4.5. Guidelines in the event of fire

There are various ways of raising the alarm in the event of fire,: By telephone: ring the alarm number 55555. Manually: push the button on the manual fire alarm (red box with glass front, see Figure 5). Automatically: the smoke alarms are activated.
	Figure 5. Manual fire alarm

The alarm centre (manned 24 hours per day, 7 days per week for the entire university and the hospital) will respond to the alarm. An automatic emergency call will be put through to the Internal Emergency Team during working hours (8.30-16.30 hrs.), and outside office hours, the Nijmegen fire service will be alerted. If necessary, the evacuation alarm will sound on the floor of the wing where the alarm was raised and any other wings/floors affected. The emergency escape route doors will automatically unlock; they can always be opened by activating the nearest green box. The partitioning fire doors along the central corridors will close automatically if necessary.

The guidelines for what to do if you discover a fire are as follows:

First phase:

• Warn everyone in the vicinity and ask for help
• Close the doors to the fire
• Ring the alarm number 55555
• Clearly state:

- your name

- the location (room number) of the fire

- what is on fire

- special details

• Close the doors to the fire
• Warn everyone in the vicinity
• Escort people at risk to a safe place (behind fire-proof doors in the corridors)
• If possible, try to extinguish the fire
• Be aware of your own safety

- Never enter a room filled with smoke

- Do not use water to extinguish a fire in electrical equipment

- Close all doors and windows when leaving the laboratory

- Never use the lift if you suspect fire in the building, or if the evacuation alarm sounds

- Follow instructions given by the Internal Emergency Team (BHV) and the external emergency services.

Second phase:

• Once the alarm has been raised (first phase), the Internal Emergency Team will arrive within 4 minutes. Follow their instructions once they are in place.
• The Internal Emergency Team (BHV, also known as Company Emergency Response Organization, CERO) is made up of people trained to work with pressurized air and in first-aid. They fulfil a coordinating role to save lives, evacuate the danger zone and bring the fire under control.
• The Internal Emergency Team is in close in contact with the internal and external emergency services that have been called in.
• Coordinated emergency help is laid down the Internal Emergency Team emergency plan, and the evacuation procedure in the Internal Emergency Team evacuation plan (see also IM emergency plan 1)

Third phase:

This phase is not directly relevant to laboratory workers

• The external emergency services, fire service, GGD and police arrive on site.
• On arrival, the fire brigade takes charge.
• The Internal Emergency Team (BHV) already in attendance shows the external emergency services the way and assists.

Evacuation can be initiated by an alarm or by members of the Rapid Intervention Team/Internal Emergency Team. The evacuation alarm is a continuous signal (a "slow whoop"), which is tested every first Monday of the month at 12:00h.

In the event that the premises must be evacuated, the following supplementary guidelines apply:

• Switch off equipment, gas and power in the lab if it is safe to do so. Calmly leave the fire compartment (every area has several escape routes, see figure 6) and assemble in the restaurant. If the evacuation alarm is sounding there too, the whole building must be evacuated; leave the building calmly and assemble in the Linnaeus building.
• Do not use the lift when evacuating; use the stairs instead.
• Tell the emergency services anything they want to know about the lab area concerned.

IMPORTANT: it is vital that all staff and students forced to evacuate a lab during a fire register their names at the assembly point so that the emergency services know who has been brought to safety.

5. Important information

5.1. List of abbreviations

. Important internet links

Information concerning Arbo at the intranet from FNWI
http://www.radboudnet.nl/fnwi/fnwi/arbo/.

Information concerning Health, Safety and Sustainability at the intranet from AMD
http://www.radboudnet.nl/amd/english/.

Laser safety
http://www.microscopyu.com/articles /fluorescence/lasersafety.html.

Waste management
The faculty waste management regulation was devised with the RU Department of Occupational Health & Safety and Environmental Service and can be found on
https://www.radboudnet.nl/amd/english/sustainability/environment/

Information on substances (Chemwatch)
http://jr.chemwatch.net/

Accidents
Accident report form for reporting accidents and incidents to the Department of Occupational Health & Safety and Environmental Service
https://www.radboudnet.nl/amd/english/safety/occupational/
Don't forget to inform the PAM’mers.

Overview of highly toxic gases
http://en.wikipedia.org/wiki/List_of_highly_toxic_gases

Hazardous Laboratory Chemicals Disposal Guide (advice in case of chemical spills)
Hazardous Laboratory Chemicals Disposal Guide.pdf

Durability at the University
http://www.ru.nl/duurzaamheid/duurzaamheid/duurzaamheidsagenda
http://www.ru.nl/sustainability/

Appendix 1 Breakthrough times for various glove materials.

Appendix 2. Flow diagram for liquids in laboratories; decision chart to determine whether a substance may be discharged into the drain