Euphytica 36 (1987) 855-862

FLOWER COLOUR ANALYSES IN TULIP (TULIPA

L.). THE OCCURRENCE OF CAROTENOIDS AND
FLAVONOIDS IN TULIP TEPALS
J.P.VAN EIJK, M.NIEUWHOF, H.A.VAN KEULENandP.KEIJZER

Institute for Horticultural Plant Breeding (IVT), P.O. Box 16,670O AA Wageningen, the Netherlands

Received 23 June 1986; accepted in revisedform 17 February 1987

KEY WORDS

Tulipa, tulip, flower colour analysis, carotenoids, flavonoids, cultivars, species, mutants.

SUMMARY

Carotenoids, delphinidin, cyanidin, pelargonidin and flavonols were determined semi-quantitatively in over
500 cultivars, including about 50 species and species-hybrids, as part of a research programme for flower
colour breeding of tulip. For flavonols there was almost no variation. For delphinidin, cyanidin and pelar-
gonidin a moderate variation, and for carotenoids a rather large variation was found. White cultivars con-
tain only flavonols, or also low carotenoid concentrations. Yellow cultivars generally only contain caroten-
oids in addition to flavonols. Most pink and red cultivars have cyanidin and pelargonidin and no delphini-
din. In addition to anthocyanidin, orange cultivars mostly contain high carotenoid concentrations and
purple and violet cultivars delphinidin and cyanidin. Colour description seems impossible on the basis
of pigment determinations only. Sports showed decreased pigment synthesis. Synthesis of delphinidin and
pelargonidin appeared to be dependent on synthesis of cyanidin or of its precursor.

INTRODUCTION

In 1575, CLUSIUSintroduced yellow, white, red and purple coloured tulips from Thra-
cia (present-day Bulgaria, at that time part of the Great Ottoman Empire) into the
Netherlands. From that time on cross combinations have been achieved within the
crop.
Until 1900 cross pollination was entirely left to natural vectors (mainly insects).
In the 17th and 18th century the emphasis was primarily laid on flowers with broken
colours, so called ‘variegated tulips’. The ‘black tulip’ was also very popular.
In the first half of the 20th century breeders changed over to more ambitious crossing
programmes (DIx, 1974) which resulted in a wider variation in colour and shape.
In that period HOOG (Van Tubergen Co, Haarlem) introduced wild tulip species with
very attractive flower colours such as Tulipa eichleri Regel, T. fosteriana Hoog, T.
greigii Regel and T. kaufmunniunu Regel. Crosses of Darwin tulips (T. gesneriunu L.)
with T. fosteriunu ‘Mad. Lefeber’ (syn. ‘Red Emperor’) by LEFEBERat Lisse resulted
in the Darwin tulip-hybrids with bright red colours.
The most important factors determining flower colour are (HALEVY & ASEN, 1959;
HARBORNE, 1967; SHIBATA etal., 1959, 1960, 1961, 1967; WIERING &DE VLAMING,
855
 J. P. VAN EIJK ET AL.

1973,1977, 1984 and personal communications):

- Carotenoids. They cause the yellow colour.
- Flavonoids (glycosides). These are anthocyanins consisting of anthocyanidins and
flavonols. Both are linked with one or more glycose-units:
1. the anthocyanidins pelargonidin, cyanidin and delphinidin (and their methylated
forms) with one, two or three OH groups in the B-ring and colours from red,
magenta to purple respectively;
2. the flavonols quercetin and kaempferol, forming complexes with the anthocyan-
ins, resulting in a blue shading (copigmentation);
3. glycose-units (3-glucoside, 3-rhamno-glucoside, 3, 5-diglucoside and 3-rhamno-
glucoside-5-glucoside). Glucose on the 5-site has a strong blueing effect.
- Physiochemical factors:
1. the iron- and aluminium-complex. This causes bright bluish colours as in corn
flower (Centaurea cyanus L.), but probably does not play a role in tulip (WIERING,
pers. comm.).
2. the pH of the vacuoles of the epidermis cells. The higher the pH the bluer the
pigmentation. Generally speaking, the pH is not expected to influence tulip col-
our (WIERING, pers. comm.).
The research programme of IVT was limited to the determination of carotenoids,
anthocyanidins and flavonols, as these compounds are expected to provide sufficient
information for practical breeding programmes.
Flower colour was analysed in over 500 cultivars to provide breeders with basic
information. Besides, pigment analyses were carried out in a number of mutants to
ascertain to what extent colour and colour composition are subject to changes through
mutation.

MATERIALS AND METHODS

From 1964 to 1968, 503 old and modern cultivars were grown at IVT, most of them
belonging to T. gesneriana, but also about 50 species and specieshybrids were included.
At full bloom, tepals without claws were gathered for carotenoid and flavonoid
analyses according to semi-quantitative methods. Though in tulips the flavonoids only
occur as glycosides, the attached sugars were not scored. Total flavonol content was
determined.
In 1979, pigment analyses were made with the cultivars Apeldoorn and ‘Preludium’
and with mutants obtained from them. In this material kaempferol and quercetin were
determined separately.

Estimation of carotenoids. A sample of 0.2 g of fresh tepals was rubbed with a small
quantity of sand and extracted with 5 ml of a 3 : 2 petroleum ether-acetone mixture.
The decanted solutions were compared with a series of standard concentrations of
potassium chromate in small test tubes.
For the preparation of the standard solutions a 3% solution of potassium chromate
in water was assigned the number 5; a mixture with four times the quantity of water
was assigned 4, and those with 24,124 and 624 times the quantity of water were assigned
the numbers 3,2 and 1 respectively. The number 0 indicated absence of carotenoids.
856 Euphytica 36 (1987)