The first four installments of this series dealt with fluorescent proteins found within various anthozoans. An understanding of fluorescence is needed before we examine another phenomenon – that of reflective pigments within coral tissues, which in turn is a prerequisite to understanding spectral data collected from various corals (presented later in this series).

There is considerably less attention paid to these pigments by scientists. Since they only reflect light (and do not fluoresce and hence are not particularly useful in biomedical research), these pigments have received the attention of far fewer researchers. However, these colored pigments are of interest to those reef aquaria hobbyists wishing to exhibit vibrantly colored corals within reef aquaria.

It is a common mistake to contribute all vivid coloration of corals to fluorescence – many corals’ beautiful blues, purples, pinks or reds due to the presence of chromoproteins.

 

Chromoproteins

Chromoproteins (from the Greek words ‘khroma’ (color) and ‘prote’ (primary)), for our purposes, are organic pigment-containing compounds that simply absorb and reflect radiation – they are normally not fluorescent or only weakly fluorescent. Their coloration is generally purple-blue (Labas et al., 2002), although violet, blue or pink/red chromoprotein pigments are known to exist. Generally, these pigments absorb light in the bandwidth of ~560- ~590nm (though there are exceptions). See Table 2.

Different terminology is used when discussing non-fluorescent chromoproteins. These are some terms with which we should be familiar:

Absorb: To retain light without reflection or transmission.

Absorption: The process in which incident radiation is retained without reflection or transmission on passing through a medium.

Absorbance: The ability of a solution or layer of a substance to absorb radiation. Expressed mathematically (for those who want to know) as the negative common logarithm of the transmittance of the substance and abbreviated as ‘Abs’.

FWHM = Full Width at Half Maximum. The spectral width (in nanometers) at the level that is 50% of the value at the peak.

Pocilloporan – Chromoproteins originally isolated from Pocillopora stony corals.

Reflect: To ‘bounce back’ light without alteration of wavelength.

Reflection: The process in which incident radiation is returned without alternation of wavelength, in others words; reflected light is not absorbed or fluoresced.

Reflectance: The ratio of the total amount of radiation (such as light) reflected by a surface to the total amount of radiation incident to the surface.

In the cases of non-fluorescent chromoproteins, that light energy that is not absorbed by a substance (coral tissue, host pigments, photopigments) is reflected, and apparent color is a combination of absorbed and reflected light. One of the most apparent examples of apparent color is that of green leaves containing chlorophylls. Since chlorophylls absorb red and blue light, the majority of the light reflected is green. Corals containing zooxanthellae are often brown, since the zooxanthellae contain chlorophylls and peridinin which absorb blue, green and red light, thus making them appear brown. Corals containing chromoproteins will usually appear purple-blue, but can be almost any color of the rainbow.

 

Color Definitions

Defining color by bandwidth would seem a straightforward proposition. However, few references agree. For clarification, see Table 1 for a listing of bandwidth and associated colors used in this article.

Table 1. Color and associated wavelengths.

Color	Wavelength (nm)
Violet	400 – 430
Blue	431 – 480
Green-Blue	481 – 490
Blue-Green	491 – 510
Green	511 – 530
Yellow-Green	531 – 570
Yellow	571 – 580
Orange	581 – 600
Red	601 – 700

Table 2. A concise listing of chromoproteins and hosts.

	Host	Color	Reference
CP-480	Pocillopora	Red	Shibata, 1969
CP-500	Acropora	Yellow	Shibata, 1969
CP-560	Pocillopora damicornis	Pink	Dove et al., 1995
CP-560	Seriatopora hystrix	Pink	Dove et al., 1995
CP-560	Stylophora pistillata	Pink	Dove et al., 1995
CP-562	Anemonia sculata	Reddish	Wiedenmann et al., 2000
CP-568	Anemonia sculata	Violet	Labas et al., 2002
CP-571	Condylactis gigantea	Red	Labas et al., 2002
CP-571	Condylactis passiflora	Red	Labas et al., 2002
CP-575	Anemonia sculata	Purple	Verkusha & Lukyanov, 2004
CP-578	Heteractis crisperi	Violet/Purple	Labas et al., 2002
CP-578	Acropora digitifera	Violet/Purple	Dove et al., 1995
CP-580	Goniopora tenuidens	Purple/Red	Martynov et al., 2003
CP-586	Acropora	Violet	Shibata, 1969
CP-588	Rhizostoma anemone	Blue	Bulina et al., 2004
CP-588	Cassiopea xamachana	Blue	Bulina et al., 2004
CP-588	Acropora formosa	Blue	Dove et al., 1995
CP-590	Montipora efflorescens	Blue	Nienhaus et al., 2005
CP-595	Anemonia sculata	‘Kindling’ Protein	Bulina et al., 2004
CP-597	Anemonia equina	Blue	Shkrob et al., 2005
CP-610	Cnidopis japonica	Blue	Chan et al., 2006

Table 3. An early summary of absorption bandwidths of water-soluble
chromoproteins extracted from various corals. This work still has
relevance. From Kawaguti, 1944.

Pink, Red and Purple Pigments
Coral	1st Band	1st Max Abs	2nd Band	2nd Max Abs	3rd Band
Fungia repunda	552-583	*	521-535	*	502-510
Pocillopora eydouxi	552-583	*	521-535	*	502-510
Pocillora damicornis	552-583	*	521-535	*	502-510
Stylophora sp.	552-583	*	521-535	*	502-510
Acropora grandis	552-583	*	521-535	*	502-510
Acropora pacifica	552-583	*	521-535	*	502-510
Acropora variaviris	552-583	*	521-535	*	502-510
Acropora hyacynthus	552-583	*	521-535	*	502-510
Porites sp.	552-583	*	521-535	*	502-510
Pocillopora acuta	552-583	*	521-535	*	502-510
Hydnophora microconus	552-583	*	521-535	*	502-510
Echinopora lamellosa	552-583	*	521-535	*	502-510
Goniopora sp.	568-600	~588nm	527-537	~533nm	*
Porites (arboreal)	566-596	~581	530	*	*
Porites (massive)	566-596	~581	530	*	*
Acropora echinata	615	583nm	*	*	*
Acropora sp.	567-605	583nm	531-541	*	*
Porites sp.	567-605	583nm	531-541	*	*
Favites sp.	567-605	583nm	531-541	*	*
Montipora ramosa	567-605	583nm	531-541	*	*
Montipora verrucosa	567-605	583nm	531-541	*	*
Goniopora sp.	567-605	583nm	531-541	*	*
Echinopora lamellosa	567-605	583nm	531-541	*	*
Acropora leptocyanthus	567-605	589nm	*	*	*
Porites sp.	557-608	584nm	*	*	*
Acropora formosa	567-615	583nm	531-541	537nm	*
Acropora echinata	567-615	583nm	531-541	537nm	*
Porites sp.	557-608	584nm	*	*	*

CP-480

• Maximum Absorption (wavelength): Actually in the violet range, with a shoulder absorption at 480nm.
• Full Width at Half Maximum (FWHM): Not determined
• FWHM Bandwidth: Not determined
• Apparent Color: Red
• Shibata (1969) reports this pigment was isolated from a red Pocillopora specimen. Figure 2 demonstrates the absorption of a water-soluble pigment extracted from this stony coral.

CP-500

• Maximum Absorption: 400nm (violet)
  

  Figure 3. Many Pocillopora specimens (such as this P. damicornis) exhibit an attractive reddish coloration due to the presence of at least one chromoprotein.

• Full Width at Half Maximum (FWHM): Not determined
• FWHM Bandwidth: Not determined
• Apparent Color: Yellow

CP-500 is reportedly found in a yellow Acropora specimen from Australia’s GBR (Shibata, 1969). See Figure 4 for absorption of a water-soluble pigment that appears yellow.

CP-560

Chromoproteins with absorption maxima are found in a variety of corals including (but likely not limited to) Pocillopora damicornis, Stylophora pistillata and Acropora species.

Pocillopora damicornis

• Maximum Absorption (Pocillopora damicornis): 560nm (yellow-green)
  

  Figure 4. A chromoprotein found in a yellow Acropora (from Shibata, 1969).

• Full Width at Half Maximum (FWHM): 80nm
• FWHM Bandwidth: 520-600nm
• Apparent Color: Pink
• See Figure 5.

Stylophora pistillata

• After Dove et al., 1995.
• Maximum Absorption (Stylophora pistillata): Actually at 400nm (violet), but listed as 560nm (yellow-green).
• Full Width at Half Maximum (FWHM): Not determined
  

  Figure 5. Absorbance of chromoprotein 560 from a pink Pocillopora damicornis. It is little wonder the coral appears pink – blue and green wavelengths are strongly absorbed in comparison to red wavelengths.

• FWHM Bandwidth: Not determined
• Color: Red
• See Figures 6 and 7.

Acropora sp.

• Maximum Absorption (Acropora sp.): 400nm (violet) in the visible range.
• Full Width at Half Maximum (FWHM): Not determined
• FWHM Bandwidth: Not determined
• Color: Red
• Shibata (1969) reports that this pigment (CP-560) is found along with another chromoprotein (CP-590), and appears red. See Figure 8.

CP-562 – A “Kindling” Chromoprotein

• Maximum Absorption: 562nm (yellow-green)
  

  Figure 6. Absorption spectrum of a reddish chromoprotein from Stylophora pistillata. After Dove et al., 1995.

• Full Width at Half Maximum (FWHM): 60nm
• FWHM Bandwidth: 520-580nm
• Apparent Color: Reddish

This reddish chromoprotein is found in the anemone Anemonia sculata var.rufescens (specifically referred to asasCP-562). Figure 9. This is an interesting pigment – ‘green’ light (540-560nm – ‘yellow-green light according to our definition) transforms (‘kindles’) non-fluorescent CP-562 (which appears red) to a fluorescent state with an emission maxima of 595nm (which appears as red fluorescence due to the emission bandwidth). This pigment, along with CP-580, blows a neat hole in our tidy concepts of fluorescent and non-fluorescent pigments – these can be both! See http://www.advancedaquarist.com/2006/11/aafeature2 for further details.

In Closing

We have reviewed the absorbance of several non-fluorescent proteins found in anthozoans. The reader will have noticed that these chromoproteins appear red or pink in color. Next time, we’ll continue our examination of chromoproteins and review those responsible for purple, blue and violet coloration. We’ll also introduce the concept of reflectance, and what it tells us about not only color, but the health of the coral host and its symbiotic zooxanthellae.

I’m in the process of updating the reference list and will include that at the end of this series. Those interested in discussing coral pigments can reach me at [email protected]