Phycobiliprotein Pigments

Inside a cyanobacterial cell or a red algal thallus, phycobiliproteins do one job: they catch photons that chlorophyll cannot reach and pass them downhill to the reaction centre. The relay is remarkably precise — light absorbed by phycoerythrin near 565 nm hops to phycocyanin around 620 nm, then to allophycocyanin at 650 nm, and finally lands on chlorophyll a just above 680 nm, losing a little energy as heat at each step so the transfer only ever flows one way. Bench scientists know the outcome better than the machinery: a fluorophore whose brightest member outshines roughly thirty fluorescein molecules, stable enough in water to be conjugated, sorted, imaged and quantified.

Eata Marisource supplies that relay as discrete, characterised components. C-phycocyanin from cultivated Arthrospira, B- and R-phycoerythrin from marine red microalgae and macroalgae, and allophycocyanin are offered across the full grade ladder — from food-colour enrichments through reagent preparations up to analytical-grade material whose absorbance ratios and emission positions are recorded lot by lot. Whether the destination is a natural blue beverage concept, a flow-cytometry conjugate or a phycobilisome study, the same question decides which grade you need, and the sections below are written to answer it.

01 Four Pigments, One Antenna System

The family divides by colour and by job. Two phycoerythrins sit at the antenna's rim where the shortest wavelengths arrive; phycocyanin and allophycocyanin sit deeper, passing the excitation onward. Their spectroscopic identities are the practical specification a buyer works from:

Two consequences follow from those numbers. First, the phycoerythrins carry 34 bilin chromophores apiece, which is why a single PE molecule behaves like a cluster of small-molecule dyes rather than one — brightness is structural, not incidental. Second, the absorbance maxima are what the purity arithmetic keys on, so a supplier who cannot state the pigment type precisely cannot state the ratio meaningfully either. Subunit masses reported in the purification literature run 17.0 and 19.0 kDa for the α and β chains of C-PC, and 15.0 / 17.0 kDa for APC; B-PE adds a 30 kDa γ subunit, giving roughly 265 kDa on gel filtration.

02 What the Purity Ratio Does — and Does Not — Tell You

Phycobiliprotein grading runs on a single arithmetic habit: absorbance at the pigment's visible maximum divided by absorbance at 280 nm, where every colourless protein contributes. For C-phycocyanin the convention is A620/A280; for R- and B-phycoerythrin, A565/A280; for allophycocyanin, A650/A280 (sometimes A620 is retained by older papers). The thresholds are broadly agreed:

• PC ≥ 0.7: ratio ≥ 0.7 — food-grade colourant territory;
• PC ≥ 1.5: the cosmetic-formulation band;
• PC ≈ 3.9: reagent grade, suitable for most assay development;
• PC ≥ 4.0: analytical and pharmaceutical grade.

Three caveats deserve a line on any certificate. The visible maximum itself drifts with pH and aggregation state, so a ratio taken at a wavelength the sample is not actually peaked at inflates nothing — it simply measures the wrong pigment. The 620 nm reading cannot distinguish C-phycocyanin from allophycocyanin on its own. And any chlorophyll riding along in the extract adds a shoulder near 680 nm that the ratio never sees, which is why the full 250–720 nm spectrum should accompany the number rather than replace it.

Where a matrix genuinely contains more than one phycobiliprotein, two-wavelength arithmetic resolves them. Yoshikawa and Belay's method reads 620 and 650 nm and back-calculates each pigment: c(C-PC) = A620 × 0.162 − A650 × 0.098 and c(APC) = A650 × 0.180 − A620 × 0.042 (mg/mL, 1 cm path). For B-PE in crude Porphyridium extracts, the Bermejo equations play the same role using 565, 620 and 650 nm simultaneously. We run the applicable set on every lot and print both the ratio and the resolved concentrations.

03 The Purification Ladder: Every Grade Point Buys Purity and Pays Yield

Phycobiliproteins are water-soluble, which makes extraction gentle and purification honest: there is no solvent trick that separates pigment from protein, because they are the same molecule. Published ladders for Arthrospira C-PC and Corallina APC show the trade explicitly:

Read one way, the table says chromatography is worth it — the ratio more than sextuples. Read the other way, the final polish costs more than half the mass, which is precisely why analytical-grade material prices the way it does and why a food-grade buyer should not be paying for ratios they will never use. On the process side, salt concentrations above roughly 1.28 mol/L ammonium sulfate drop C-PC selectively at ~0.7 purity; tangential-flow filtration retains about 79% on a 50 kDa membrane but surrenders over half through a 100 kDa cassette; and a chitosan clarification (0.01 g/L) followed by 1% activated carbon measurably cleans crude extract before any column sees it. Cell disruption method matters too — nitrogen cavitation with CHAPS/asolectin buffer recovered 85–88% of total phycobiliprotein from Synechococcus in comparative trials.

Ammonium sulfate crystals feeding into a blue phycobiliprotein extract during fractional precipitation.

Blue and magenta pigment bands separating down an ion-exchange column.

04 Where the Pigment Comes From

Marine supply splits along a clear line: microalgae give the phycoerythrins their highest documented titres, while macroalgae give R-PE its industrial volume. Reported figures span a wide range, which is itself the message — species, season and cultivation mode move the number more than any extraction tweak does.

Porphyridium earns its place for a second reason: the same biomass yields sulfated exopolysaccharides, EPA and arachidonic acid, so a phycoerythrin campaign can co-product the remainder rather than discard it. Macroalgal R-PE carries the opposite economics — cheaper raw material, but pigment content swings with harvest season and the polysaccharide-rich matrix slows solvent penetration, thickens extracts and fouls membranes. Microwave-assisted extraction has compressed Porphyridium processing dramatically: 73.7 µg PE per mg biomass in ten seconds at 40 °C, a three-orders-of-magnitude cut in contact time that keeps the protein out of trouble.

Fresh Porphyra fronds, a principal commercial source of R-phycoerythrin, laid out on a processing tray.

Arthrospira biomass beside its deep-blue phycocyanin extract, dried to a fine powder.

05 Keeping the Colour Alive

Everything that unfolds a protein strips the pigment, so stability is a formulation problem, not a storage footnote. The boundaries are sharp: near pH 7 solubility peaks; below pH 3 the pigment precipitates essentially on contact. Heat is the second antagonist — blue fades and protein drops out as temperature climbs, which is why sterilisation steps and phycocyanin rarely coexist happily. Light and oxygen erode both colour and fluorescence over weeks rather than days.

• Sugars and polyols: sucrose, trehalose and other polyhydric alcohols stabilise the protein safely and are the default excipients in our dried grades;
• Preservatives: in accelerated trials at 35 ± 5 °C, citric acid preserved C-PE colour better than sodium chloride, calcium chloride or sucrose alone;
• Salt penalty: sodium chloride trims PE colour intensity by roughly 10%, worth knowing before a brine-based matrix is specified;
• Laboratory additives: sodium azide and DTT stabilise analytical samples but are excluded from any food-bound material.

Finished lots leave us lyophilised with sugar excipients, or as ammonium sulfate suspensions for the conjugation bench; the dried powders are held at 2–8 °C and solutions aliquoted at or below −20 °C. Repeated freeze-thaw cycles are the quietest killer of fluorescence recovery, and the handling sheet that ships with every lot says so in the first paragraph.

R-phycoerythrin solutions at increasing concentration glowing magenta under backlight.

06 Two Markets, One Molecule

The fluorescence toolbox

R-PE reads cleanly on the 488 nm argon laser line and emits near 578 nm with a quantum yield of 0.82 that barely notices pH or temperature across physiological ranges — a tolerance built into the protein scaffold, which shelters the bilins from concentration quenching. Streptavidin–phycoerythrin conjugates resolve as few as one hundred bound antibodies per cell. The engineering gets better with tandems: crosslink B-PE to APC and excitation at 545 nm re-emerges at 660 nm, an effective Stokes shift past 100 nm; couple R-PE to a Cy5-type dye and the emission climbs to 660–665 nm for four-colour sorting panels. All of this depends on intact chromophore geometry, which is why fluorescence-grade material is shipped as ammonium sulfate suspension or lyophilisate with the spectrum on file.

Colour, cosmetics and formulation research

Pure C-phycocyanin is gentian blue; allophycocyanin leans turquoise; residual chlorophyll drags commercial extracts greenward, and ultrafiltration removes it. Blue on its own is only the start — blended with yellow colorants such as carthamin it spans a family of greens no synthetic lake quite matches. In cosmetic research, Porphyridium-derived PE rides alongside the alga's sulfated polysaccharides and zeaxanthin, and B-PE has documented activity in ORAC-type antioxidant assays, where its quantum yield above 0.9 makes it the integrity reporter itself. What the colour market must respect is the stability envelope above: phycocyanin colourants want neutral pH, gentle thermal budgets and opaque packaging, and they reward that care with a hue nothing synthetic replicates.

Phycoerythrin vials fluorescing bright orange-pink under blue excitation in a darkened bench.

07 Grades and Pack Forms

Every certificate carries: full 250–720 nm absorbance spectrum; ratio at the applicable wavelength pair; resolved C-PC / APC concentrations where the method applies; emission maximum and relative quantum yield for fluorescence grades; protein content; pH and conductivity of the reconstituted solution; and heavy metals. For cyanobacterial material we add microcystin screening on request — a formality for cultured Arthrospira, a necessity when the source is wild.

08 Custom Isolation and Development

Because the family spans four proteins, five grade bands and two very different end markets, most programmes start with a conversation rather than a catalogue page. We isolate to a target ratio rather than a target mass; we can transfer a purification from macroalgal R-PE to microalgal B-PE when a conjugate needs the extra quantum yield; we formulate sugar-protected powders for a specific pH window or thermal budget; and we prepare conjugation-activated intermediates so a downstream lab never touches the crosslinker chemistry. Co-product recovery from the same biomass — Porphyridium exopolysaccharide, EPA-rich fractions, Spirulina protein side-streams — is quoted as part of the campaign rather than after it. Tell us the wavelength you need to excite, the matrix the pigment must survive, and the grade band your budget allows; the isolation is designed backwards from there.

Eata Marisource ships phycobiliproteins worldwide for research and industrial programmes. Request the current lot spectra, a sample for bench validation, or a quotation on your next campaign — our team answers with numbers, not brochures.

For Research or Industrial Raw Materials, Not For Personal Medical Use!